ceruletide and Acute-Disease

Acute pancreatitis (AP) is one of the most common diseases in gastroenterology, affecting 2% of all hospitalized patients. Nevertheless, neither the etiology nor the pathophysiology of the disease is fully characterized, and no specific or effective treatment has been developed. Heparanase (Hpa) is an endoglycosidase that cleaves heparan sulfate (HS) side chains of heparan sulfate proteoglycans (HSPGs) into shorter oligosaccharides, activity that is highly implicated in cell invasion associated with cancer metastasis and inflammation. Given that AP is a typical inflammatory disease, we investigated whether Hpa plays a role in AP. Our results provide keen evidence that Hpa expression and activity are significantly increased following cerulein-induced AP in wild type mice. In parallel to the classic manifestations of AP, namely elevation of amylase and lipase levels, pancreas edema and inflammation as well as induction of cytokines and signaling molecules, have been detected in this experimental model of the disease. Noteworthy, these features were far more profound in transgenic mice overexpressing heparanase (Hpa-Tg), suggesting that these mice can be utilized as a model system to reveal the molecular mechanism by which Hpa functions in AP. Further support for the involvement of Hpa in the pathogenesis of AP emerged from our observation that treatment of experimental AP with PG545 or SST0001(= Ronepastat), two potent Hpa inhibitors, markedly attenuated the biochemical, histological and immunological manifestations of the disease. Hpa, therefore, emerges as a potential new target in AP, and Hpa inhibitors are hoped to prove beneficial in AP along with their promising efficacy as anti-cancer compounds.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Glucuronidase; Humans; Pancreatitis

Hydrogen sulfide (H2S) and substance P play a key role in inflammation. Using animal models of inflammation of different etiologies such as acute pancreatitis, sepsis, burns, and joint inflammation, studies have recently shown an important role of the proinflammatory action of H2S and substance P. Also, H2S contributes to inflammation in different conditions via substance P. This chapter reviews methods and key data that have led to our current understanding of the role of H2S and substance P in inflammation.

Topics: Acute Disease; Animals; Burns; Carrageenan; Ceruletide; Disease Models, Animal; Drug Synergism; Edema; Endotoxemia; Hydrogen Sulfide; Lipopolysaccharides; Lung Injury; Mice; Pancreatitis; Substance P

Melatonin, the main product of the pineal gland, is also released from the gastrointestinal endocrine-neurocrine (EE) cells. The concentrations of melatonin produced in the gut exceeds that originating from central nervous system. In spite of the presence of melatonin receptors in the pancreatic tissue little is known about the role of this indole in the pancreas. Our experimental studies have shown that exogenous melatonin, as well as this produced endogenously from its precursor; L-tryptophan, strongly stimulates pancreatic amylase secretion when given intraperitoneally, or into the gut lumen. This was accompanied by significant increases of CCK plasma level. Above pancreatostimulatory effects of luminal administration of melatonin, were completely reversed by bilateral vagotomy, capsaicin deactivation of sensory nerves or pretreatment of the rats with CCK1 receptor antagonist; tarazepide as well as serotonin antagonist; ketanserin. Melatonin, as well as its precursor; L-tryptophan, effectively protects the pancreas against the damage induced by caerulein overstimulation or ischemia/reperfusion. The beneficial effects of melatonin or L-tryptophan on acute pancreatitis could be related to the ability of melatonin to scavenge the free radicals, to activate antioxidative enzymes and to modulate the cytokine production.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cholecystokinin; Free Radical Scavengers; Gastrointestinal Tract; Melatonin; Pancreas; Pancreatitis; Receptor, Cholecystokinin A; Reperfusion Injury; Tryptophan

In acute pancreatitis, multiple organ failure in the early phase and infectious complications in the late phase are contributors to mortality. To analyze the mechanism of aggravation of acute pancreatitis is to investigate the mechanism of organ dysfunction and infection. As strategy to elucidate the mechanism, various animal experimental models are utilized. Caerulein-induced pancreatitis and bile salt-induced pancreatitis (duct injection model) are frequently employed for mild edematous pancreatitis and severe necrotizing pancreatitis, respectively. It is important to select an appropriate experimental model that corresponds to the purpose of study.

Topics: Acute Disease; Animals; Apoptosis; Arginine; Bacterial Translocation; Bile Acids and Salts; Ceruletide; Disease Models, Animal; Disease Progression; Duodenum; Ethionine; Humans; Ligation; Lipopolysaccharides; Macrophage Activation; Pancreatitis; Severity of Illness Index

Topics: Acute Disease; Animals; CD11b Antigen; CD11c Antigen; Ceruletide; Chemokines; Endothelium; Humans; Intercellular Adhesion Molecule-1; Liver; Neuropeptides; Pancreatitis; Reactive Oxygen Species; Respiratory Distress Syndrome; Selectins; Signal Transduction; Transcription, Genetic

Autodigestion by proteolytic enzymes is thought to represent the critical mechanism by which acute pancreatitis is initiated. Where and why pancreatic proteases, which are physiologically stored and secreted as inactive precursor zymogens, are activated within the pancreas has remained controversial. Here we present data which indicate that: the lysosomal protease cathepsin B can activate trypsinogen in vitro in a manner that is similar to trypsinogen activation by enterokinase; that cathepsin B colocalizes with trypsinogen in the secretory compartment of the rat pancreas and of the human pancreas; that trypsinogen activation begins in a secretory compartment that is distinct from mature zymogen granules; and that the inhibition of cathepsin B can either increase or decrease premature trypsinogen activation depending on the concentration of the inhibitor, its specificity and its site of action in the pancreatic acinar cell. These observations elucidate some of the complex relations between cysteine and serine proteases in the pancreas with respect to their mechanisms of activation, their subcellular sites of action, and their possible role in the onset of pancreatitis.

Topics: Acute Disease; Animals; Cathepsin B; Ceruletide; Cytoplasmic Granules; Dogs; Enteropeptidase; Enzyme Activation; Humans; Lysosomes; Mice; Pancreas; Pancreatitis; Rats; Trypsin; Trypsinogen

In the last decade, the role of oxidative stress has been extensively evaluated in different experimental models of acute pancreatitis. This review shows that there is strong evidence that this stress occurs as an early phenomenon in pancreatic tissue in the course of caerulein-induced acute pancreatitis. Oxidative stress was documented in pancreatic tissue by means of methods showing generation of reactive oxygen species (e.g., chemiluminescence) and accumulation of products of reactive oxygen species-mediated lipid peroxidation. with concomitant depletion of enzymatic and low molecular weight antioxidants. Features of acinar cell injury and inflammation, especially pancreatic edema, show a marked improvement following treatment with a broad spectrum of antioxidants, platelet activating factor antagonists, or donors of nitric oxide (NO). Unfortunately, in most cases these beneficial effects are temporary and generally restricted to an early phase of the disease. However, results of well-designed clinical trials should finally evaluate the importance of oxidative stress-oriented treatment in acute pancreatitis in humans.

Topics: Acute Disease; Animals; Ceruletide; Gastrointestinal Agents; Humans; Oxidative Stress; Pancreatitis

Topics: Acute Disease; Animals; Cathepsin B; Ceruletide; Disease Models, Animal; Enzyme Activation; Humans; In Vitro Techniques; Lysosomes; Pancreas; Pancreatitis; Trypsinogen

Circumstantial evidence suggests that gallstone-induced pancreatitis is triggered by obstruction of the pancreatic duct. In this report I will review the results of studies conducted during the last decade that have employed the diet-induced, secretagogue-induced, and duct obstruction-induced models of experimental pancreatitis to investigate the early events that lead to the development of acute pancreatitis. In each of these models, digestive enzyme zymogens and the lysosomal hydrolase cathepsin B were found to become colocalized. These observations have led to the hypothesis that intra-acinar cell activation of digestive enzyme zymogens by lysosomal hydrolases may be an important critical event in the development of acute pancreatitis. Recent morphologic studies evaluating the initial 24 hours after ligation of the opossum pancreatic duct indicate that the earliest lesions in this model of hemorrhagic pancreatitis occur in acinar cells.

Topics: Acute Disease; Amino Acids; Amylases; Animals; Ceruletide; Cholelithiasis; Choline Deficiency; Disease Models, Animal; Edema; Enzyme Precursors; Ethionine; Hydrolases; Mice; Pancreatitis; Protein Biosynthesis; Proteins; Rabbits

Intravenous infusion of the synthetic cholecystokinin analogue cerulein at a dose of 0.25 micrograms/kg/h causes maximal stimulation of pancreatic exocrine secretion. The infusion of supramaximal doses of cerulein (5 and 10 micrograms/kg/h) induces a significant increase in pancreatic enzymes in blood, and interstitial edema and inflammatory cell infiltration. This model of hormone-induced pancreatitis works in rats, mice, dogs and hamsters. Besides intravenous infusion, repeated intraperitoneal injections can also be used for induction of pancreatitis. In the early phase of cerulein-induced pancreatitis, large autophagic vacuoles result from fusion of zymogen granules within the acinar cell. This is accompanied by an increase in lysosomal enzyme activity and activation of trypsinogen which finally leads to cellular necrosis. All animals survive the induction of pancreatitis. The pancreas completely regenerates within 6 days after induction of pancreatitis. This model of experimental pancreatitis favors the analysis of intracellular events in the early phase of pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Humans; Infusions, Intravenous; Pancreatitis; Proteins; Regeneration

A membrane-bound system through which secretory and lysosomal proteins travel in a vectorial fashion is essential for the preserved integrity of pancreatic acinar cells. This system is composed of an ordered array of compartments, such as the rough endoplasmic reticulum, the Golgi complex, lysosomes, and secretory granules. As a principle, in acute pancreatitis the final steps of this transport seem to be disturbed. Caerulein-induced pancreatitis is a valuable experimental model for studying altered intracellular transport, and compartmentation of lysosomal and digestive enzymes. The formation of enlarged secretory vacuoles containing lysosomal and digestive enzymes is paralleled by the activation of lysosomes and degradation of cellular organelles in autophagosomes. On the level of secretory and autophagic vacuoles, activation of serine proteases occurs, which in addition to increasing lysosomal enzyme activities can represent the initial stage for acinar cell destruction and the development of pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Humans; Lysosomes; Pancreatitis

The authors examined the effect of long acting somatostatin analogue (Sandostatin, Sandoz) on acute experimental pancreatitis and on the subsequent regeneration. Acute injury to the pancreas was produced by an intraductal intervention (ligature of the bile duct and intraductal injection of taurocholic acid) and by a metabolic route (supramaximal dose of caerulein by repeated subcutaneous injections). The effect of the drug on the acute injury was examined at 6 and 24 hours following the intervention and the effect on regeneration was examined on day 3 and 5 in all cases by determination of plasma enzyme levels and examination of the pancreatic tissue. Long acting somatostatin analogue did not prove to be effective in the serious acute pancreatitis produced by the intraductal intervention. However, in the acute phase of the caerulein induced pancreatitis, it had a beneficial effect as seen by it's ability to moderate the serum enzyme levels. During the examination of pancreatic regeneration was found that in caerulein induced pancreatitis the weight of the pancreas decreases due to atrophy and that this was not affected by long acting somatostatin analogue. As a matter of fact, the somatostatin counteracted the caerulein induced DNA increase, and therefore acted against the reactive hyperplasia. Therefore, the favorable effect of long acting somatostatin analogue is witnessed only in the caerulein induced acute injury but it does not accelerate the rate of pancreatic regeneration following injury. Due to this fact, protracted administration of this agent can not be rationalized.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Drug Evaluation, Preclinical; Octreotide; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

The purpose of this study was to investigate the utility of repeated applications of ceruletid to reduce gallbladder volume and its feasibility as a means of prophylaxis of acute acalculous cholecystitis in intensive care patients. First, a dose-response curve of ceruletid was obtained in 20 mechanically ventilated patients of a surgical intensive care unit (SICU) not receiving enteral nutrition. An effective dose of ceruletid, defined by a 50% reduction of gallbladder volume was established and subsequently studied in 40 mechanically ventilated SICU patients on total parenteral nutrition in a prospective, randomized, controlled, triple-blind trial. Gallbladder volume, sludge formation and side effects were evaluated. A dose of 1.5 micrograms/kg body weight ceruletid was established as the effective dose, causing 50% reduction of gallbladder volume in all patients studied and reduction of gallbladder sludge in 95%. In 67.5% of patients side effects were observed, requiring therapeutic intervention in 68%. It is concluded that ceruletid is effective in stimulating gallbladder contraction and reducing sludge formation in severely ill patients on intensive care units. Its routine use as prophylaxis of acute acalculous cholecystitis, however, may be limited by the nature, severity and frequency of side effects.

Topics: Acute Disease; Adult; Aged; Aged, 80 and over; Ceruletide; Cholecystitis; Critical Care; Dose-Response Relationship, Drug; Female; Gallbladder; Gastrointestinal Agents; Humans; Male; Middle Aged; Multivariate Analysis; Prospective Studies

In the present study, we aimed to investigate the effects of Fenbufen treatment on the SAP model induced by caerulein and lipopolysaccharide.. Severe acute pancreatitis (SAP) is an extremely dangerous disease with high mortality, which is associated with inflammatory response and acinar cell death. The caspase family plays an important role in cell death, such as caspase-1 and caspase-11 in pyroptosis. In recent years, caspases have been shown to be a novel pharmacological target of Fenbufen.. Effects of Fenbufen on pancreatic tissue damage and serum levels of lipase and amylase in SAP in mice; Effect of Fenbufen on caspase-1 pathway in SAP in mice; Effect of Fenbufen on caspase-1/caspase-11-mediated pyroptosis of PACs in SAP in mice; Effect of Fenbufen on isolated PACs and caspase-1/caspase-11-mediated pyroptosis in vitro.. In vivo, eighteen female C57BL/6 mice were randomly divided into 3 groups: the NC group, the SAP group, and the Fenbufen +SAP group with 6 mice in each group. The SAP model was induced by intraperitoneal injection of caerulein and lipopolysaccharide. The pathological changes in pancreatic and the serum levels of lipase and amylase and the relative gene and protein expressions in each group were compared. In vitro, pancreatic acinar cells were assigned to 5 groups: medium group, SAP group, Fenbufen 100μM group, Fenbufen 200μM group, and Fenbufen 400μM group. The cell damage and the relative gene and protein expressions in each group were evaluated.. Our results showed that Fenbufen ameliorated the severity of SAP and decreased the serum levels of lipase and amylase. Meanwhile, the in vivo and in vitro data demonstrated that Fenbufen inhibited the activation of caspase-1 and caspase-11, decreasing the levels of IL-1β, IL-18, and GSDMD. In in vitro experiments, we found that by inhibiting the activation of caspase-1 and caspase-11, Fenbufen significantly reduced lactate dehydrogenase (LDH) excretion by acinar cells.. In general, our data showed that Fenbufen could protect the pancreatic acinar cell from injury by inhibiting pyroptosis.

Topics: Acute Disease; Amylases; Animals; Caspase 1; Caspases; Ceruletide; Female; Lipase; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Pancreatitis; Pyroptosis

Acute pancreatitis (AP) is an acute inflammatory condition of pancreas with high morbidity and mortality, which has no effective medical treatment. Chaiqin chengqi decoction (CQCQD) has been clinically used for AP for many years in China. However, the underlying mechanisms are still unknown.. To investigate the mechanism of CQCQD on gasdermin D (GSDMD) -mediated pyroptosis in AP.. In this study, network pharmacology was used to screen the potential mechanism of CQCQD protecting against AP and then we focused to investigate the mechanism of CQCQD on GSDMD mediated pyroptosis. Mouse models of AP were conducted by caerulein and L-arginine. In order to clarify the mechanism of CQCQD, two kinds of GSDMD gene knockout mice (Gsdmd. In the caerulein-induced AP model, CQCQD ameliorated pancreatic pathological injury, attenuated systemic inflammation and serum enzymatic levers. Moreover, network pharmacology analysis showed GSDMD mediated pyroptosis was one of the core targets of CQCQD protecting against AP. Additionally, CQCQD appreciably decreased the levels of pyroptosis-related proteins N-terminal GSDMD, nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3, and cleaved Caspase-1. Furthermore, the protective effect of CQCQD was neutralized in Gsdmd. CQCQD could reduce pancreatic necrosis and inflammatory response via inhibiting GSDMD-mediated pyroptosis in acinar cells of AP. Rhein may be the key active ingredient of CQCQD in suppressing pyroptosis.

Topics: Acute Disease; Animals; Ceruletide; Gasdermins; Humans; Mice; Molecular Docking Simulation; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreatitis; Pyroptosis

Gut microbiota dysbiosis induced by acute pancreatitis (AP) exacerbates pancreatic injury and systemic inflammatory responses. The alleviation of gut microbiota dysbiosis through faecal microbiota transplantation (FMT) is considered a potential strategy to reduce tissue damage and inflammation in many clinical disorders. Here, we aim to investigate the effect of gut microbiota and microbiota-derived metabolites on AP and further clarify the mechanisms associated with pancreatic damage and inflammation.. AP rat and mouse models were established by administration of caerulein or sodium taurocholate in vivo. Pancreatic acinar cells were exposed to caerulein and lipopolysaccharide in vitro to simulate AP.. Normobiotic FMT alleviated AP-induced gut microbiota dysbiosis and ameliorated the severity of AP, including mitochondrial dysfunction, oxidative damage and inflammation. Normobiotic FMT induced higher levels of NAD. Gut microbiota-derived NMN alleviates the severity of AP by activating the SIRT3-PRDX5 pathway. Normobiotic FMT could be served as a potential strategy for AP treatment.

Topics: Acute Disease; Animals; Ceruletide; Dysbiosis; Gastrointestinal Microbiome; Inflammation; Mice; NAD; Nicotinamide Mononucleotide; Pancreatitis; Rats; Sirtuin 3

Acute pancreatitis (AP) is an acute inflammatory abdominal disease frequently associated with intestinal barrier dysfunction. Biochanin A (BCA), a dietary isoflavone, has gained increasing interest with its pronounced biological activities. However, its potential beneficial effects on AP have not been demonstrated. Herein, we explored the protective effect of BCA on caerulein-induced AP in BALB/c mice and underlying mechanisms. BCA alleviated AP as evidenced by reduced serum amylase and lipase levels, pancreatic edema, pancreatic myeloperoxidase activity, and improved pancreatic morphology. Amelioration of pancreatic damage by BCA was associated with reduced levels of tumor necrosis factor-α, interleukin (IL)-1β, IL-6, and monocyte chemotactic protein-1 in both pancreas and colon. Moreover, BCA attenuated AP-associated barrier damage by upregulating the expression of tight junction proteins zonulin occluding (ZO)-1, ZO-2, occludin, and claudin-1. Concomitantly, the translocation of pathogenic bacteria Escherichia coli (E. coli) to pancreas was reduced by BCA. More importantly, reduction of E. coli dissemination by BCA inhibited the TLR4-MAPK/NF-κB signaling and NLRP3 inflammasome activation, thereby protecting against AP and related intestinal injury. Consistently, TLR4 inhibition by TAK-242 pre-treatment counteracted the anti-inflammatory effects of BCA in acinar cells. Taken together, our study extends beneficial effects of BCA to AP prevention, and dietary BCA supplement may be a potential strategy to safeguard AP.

Topics: Acute Disease; Animals; Ceruletide; Escherichia coli; Mice; NF-kappa B; Pancreatitis; Toll-Like Receptor 4

Thiopurine-induced acute pancreatitis (TIP) is one of the most common adverse events among inflammatory bowel disease patients treated with azathioprine (AZA), representing a significant clinical burden. Previous studies focused on immune-mediated processes, however, the exact pathomechanism of TIP is essentially unclear.. To model TIP in vivo, we triggered cerulein-induced experimental pancreatitis in mice receiving a daily oral dose of 1.5 mg/kg AZA. Also, freshly isolated mouse pancreatic cells were exposed to AZA ex vivo, and acinar cell viability, ductal and acinar Ca. We demonstrated that AZA treatment increases tissue damage in the early phase of cerulein-induced pancreatitis in vivo. Also, both per os and ex vivo AZA exposure impaired pancreatic fluid and ductal HCO. AZA impaired the ductal HCO

Topics: Acute Disease; Animals; Bicarbonates; Cell Membrane; Ceruletide; Cystic Fibrosis Transmembrane Conductance Regulator; Mice; Pancreatitis

Acute pancreatitis (AP) is a frequent abdominal inflammatory disease. Despite the high morbidity and mortality, the management of AP remains unsatisfactory. Disulfiram (DSF) is an FDA-proved drug with potential therapeutic effects on inflammatory diseases. In this study, we aim to investigate the effect of DSF on pancreatic acinar cell necrosis, and to explore the underlying mechanisms. Cell necrosis was induced by sodium taurocholate or caerulein, AP mice model was induced by nine hourly injections of caerulein. Network pharmacology, molecular docking, and molecular dynamics simulation were used to explore the potential targets of DSF in protecting against cell necrosis. The results indicated that DSF significantly inhibited acinar cell necrosis as evidenced by a decreased ratio of necrotic cells in the pancreas. Network pharmacology, molecular docking, and molecular dynamics simulation identified RIPK1 as a potent target of DSF in protecting against acinar cell necrosis. qRT-PCR analysis revealed that DSF decreased the mRNA levels of RIPK1 in freshly isolated pancreatic acinar cells and the pancreas of AP mice. Western blot showed that DSF treatment decreased the expressions of RIPK1 and MLKL proteins. Moreover, DSF inhibited NF-κB activation in acini. It also decreased the protein expression of TLR4 and the formation of neutrophils extracellular traps (NETs) induced by damage-associated molecular patterns released by necrotic acinar cells. Collectively, DSF could ameliorate the severity of mouse acute pancreatitis by inhibiting RIPK-dependent acinar cell necrosis and the following formation of NETs.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Disulfiram; Mice; Molecular Docking Simulation; Necrosis; Pancreatitis; Receptor-Interacting Protein Serine-Threonine Kinases

Acute pancreatitis can eventually lead to morbidity and mortality. The present study aimed to identify the differentially expressed microRNAs (miRNAs) that are related to acute pancreatitis and explore the in vitro functional role of miR-92b in acute pancreatitis.. Bioinformatics analysis was used to identify differentially expressed miRNAs in caerulein- induced acute pancreatitis samples when compared to normal controls. The role of miR-92b in acute pancreatitis was examined by in vitro functional assays.. MiRNA-network analysis revealed 12 miRNAs that function as "core regulatory miRNAs". Further validation studies revealed that six miRNAs (miR-216a, miR-216b, miR-217, miR- 92b, miR-375 and miR-148a) were differentially expressed in the serum samples from patients with acute pancreatitis. These six miRNAs have fair diagnostic potential for severe acute pancreatitis. Caerulein induced cell injury and inflammatory response and repressed miR-92b expression in AR42J cells. MiR-92b overexpression attenuated caerulein-induced cell injury and inflammatory responses in AR42J cells. Luciferase reporter assay showed that mitogen-activated protein kinase 4 (MAP2K4) was a direct target of miR-92b. MiR-92b overexpression repressed MAP2K4 expression, while caerulein up-regulated MAP2K4 expression in AR42J cells. The rescue experiments showed that enforced expression of MAP2K4 partially reversed the miR-92b-mediated protective effects on caerulein-induced AR42J cell injury.. In conclusion, we identified miR-216a, miR-216b, miR217, miR-92b, miR-375 and miR-148a as new candidate biomarkers for acute pancreatitis. Further in vitro functional studies revealed that miR-92b attenuated caerulein-induced cell injury and inflammatory responses in AJ42R cells partially via targeting MAP2K4.

Topics: Acute Disease; Biomarkers; Ceruletide; Humans; MicroRNAs; Pancreatitis

To explore the possible mechanism of Dachaihu Decoction (DCHD) in the treatment of AP, and use in vivo experiments to verify.. The targets and active ingredients of DCHD in the treatment of AP were obtained through network pharmacology, and the preliminary verification was carried out by molecular docking. Caerulein was used to develop the AP rat model. H&E staining was performed to observe variations in pancreatic tissue. Western blot and RT-qPCR were conducted to evaluate the associated proteins and mRNA.. The network pharmacology and molecular docking results showed that the key targets (EGFR, TNF, SRC, VEGFA and CTNNB1) and key active components (beta-sitosterol, stigmasterol, baicalein, quercetin, and kaempferol) of DCHD in the treatment of AP had good binding. H&E staining revealed that rat pancreatic tissues considerably damaged post caerulein intervention, and it has also been suggested that DCHD ameliorates damage to pancreatic tissue. Simultaneously, EGFR, TNF, SRC, VEGFA protein, and mRNA expression levels were increased in the model group compared to the blank group (. DCHD protects pancreatic tissues and improves symptoms in AP rats by upregulating CTNNB1 protein and mRNA while inhibiting EGFR, TNF, SRC, and VEGFA protein and mRNA expression.

Topics: Acute Disease; Animals; Artificial Intelligence; Ceruletide; Drugs, Chinese Herbal; ErbB Receptors; Molecular Docking Simulation; Pancreatitis; Rats; RNA, Messenger

Acute pancreatitis (AP) is a disease characterized by local and systemic inflammation with an increasing incidence worldwide. Receptor-interacting serine/threonine protein kinase 3 (RIPK3), mixed-lineage kinase domain-like protein (MLKL), and innate immune cell macrophages have been reported to be involved in the pathogenesis of AP. However, the mechanisms by which RIPK3 and MLKL regulate pancreatic injury, as well as the interactions between injured pancreatic acinar cells and infiltrating macrophages in AP, remain poorly defined. In the present study, experimental pancreatitis was induced in C57BL/6J, Ripk3

Topics: Acute Disease; Animals; Ceruletide; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreatitis; Protein Kinases; Receptor-Interacting Protein Serine-Threonine Kinases; Transcription Factors

Acute pancreatitis (AP) is one of the life-threatening diseases of the digestive system. MicroRNA has been asserted to be a regulator of AP. This paper explored the miR-374a-5p expression in AP patients and investigated the efficacy of AR42J cells. In this study, 60 healthy people, 58 MAP patients and 58 SAP patients were included, and the serum miR-374a-5p levels of the subjects were detected by RT-qPCR technology. The pancreatitis cell model was structured by stimulating AR42J cells with cerulein. Next, cell viability and apoptosis were detected by CCK-8 assay and flow cytometry. ELISA was used to measure the concentration of cytokines, such as TNF-α, IL-6, and IL-1β. The data showed that miR-374a-5p was downregulated in samples from AP patients, while showing discriminative power for AP populations. Attenuated miR-374a-5p were negatively bound up with patients' Ranson score and APACHE II score. Besides, miR-374a-5p was declined in cerulein-treated AR42J cells and forced elevation of miR-374a-5p was beneficial to increase cell viability, and inhibit cell apoptosis and inflammation. The present study found that miR-374a-5p was reduced in AP serum samples, and up-regulated expression level of miR-374a-5p in cell models had a protective effect on cerulein-induced inhibition of cell function and inflammatory response.

Topics: Acinar Cells; Acute Disease; Apoptosis; Ceruletide; Humans; MicroRNAs; Pancreatitis

Necroptosis is a necrotic form of regulated cell death, which is primarily mediated by the receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like (MLKL) pathway in a caspase-independent manner. Necroptosis has been found to occur in virtually all tissues and diseases evaluated, including pancreatitis. Celastrol, a pentacyclic triterpene extracted from the roots of Tripterygium wilfordii (thunder god vine), possesses potent anti-inflammatory and anti-oxidative activities. Yet, it is unclear whether celastrol has any effects on necroptosis and necroptotic-related diseases. Here we showed that celastrol significantly suppressed necroptosis induced by lipopolysaccharide (LPS) plus pan-caspase inhibitor (IDN-6556) or by tumor-necrosis factor-α in combination with LCL-161 (Smac mimetic) and IDN-6556 (TSI). In these in vitro cellular models, celastrol inhibited the phosphorylation of RIPK1, RIPK3, and MLKL and the formation of necrosome during necroptotic induction, suggesting its possible action on upstream signaling of the necroptotic pathway. Consistent with the known role of mitochondrial dysfunction in necroptosis, we found that celastrol significantly rescued TSI-induced loss of mitochondrial membrane potential. TSI-induced intracellular and mitochondrial reactive oxygen species (mtROS), which are involved in the autophosphorylation of RIPK1 and recruitment of RIPK3, were significantly attenuated by celastrol. Moreover, in a mouse model of acute pancreatitis that is associated with necroptosis, celastrol administration significantly reduced the severity of caerulein-induced acute pancreatitis accompanied by decreased phosphorylation of MLKL in pancreatic tissues. Collectively, celastrol can attenuate the activation of RIPK1/RIPK3/MLKL signaling likely by attenuating mtROS production, thereby inhibiting necroptosis and conferring protection against caerulein-induced pancreatitis in mice.

Topics: Acute Disease; Animals; Apoptosis; Caspases; Ceruletide; Mice; Necroptosis; Pancreatitis; Pentacyclic Triterpenes; Protein Kinases; Receptor-Interacting Protein Serine-Threonine Kinases

We aimed to show whether the serum level of Human Epididymitis Protein 4 increases in rats with an experimental acute pancreatitis model created by cerulein.. This study included 24 male Sprague-Dawley rats which were randomly divided into 4 groups each containing 6 rats.. the group treated with saline, Group 1: pancreatitis group created with cerulein at a total dose of 80 µg/kg, Group 2: pancreatitis group created with cerulein at a total dose of 120 µg/kg, Group 3: pancreatitis group created with cerulein at a total dose of 160 µg/kg.. There were statistically significant differences between edema, acinar necrosis, fat necrosis, and perivascular inflammation scores among the study groups. While the degree of all histopathological findings is lowest in the control group, pancreatic parenchyma damage increases as the amount of injected cerulein increases. There was no statistically significant difference between alanine aminotransferase, aspartate aminotransferase, and Human Epididymis Protein 4 values between study groups. On the other hand, there was a statistically significant difference between amylase and lipase values. The lipase value of the control group was significantly lower than the lipase value of the second and third groups. The amylase value of the control group was significantly lower than all other groups. The highest Human Epididymis Protein 4 value was measured as 104 pmol/L in the first pancreatitis group, where the severity of pancreatitis was mild.. In the present study, it was concluded that the Human Epididymis Protein 4 value increased in the case of mild pancreatitis, but there is no correlation between the severity of pancreatitis and the Human Epididymis Protein 4 value.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Epididymitis; Humans; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Rats, Wistar

Acute pancreatitis (AP) is an inflammatory disease with high mortality. Previous study has suggested that circular RNAs are dysregulated and involved in the regulation of inflammatory responses in AP. This study aimed to investigate the function and regulatory mechanism underlying mmu_circ_0000037 in caerulein-induced AP cellular model.. Caerulein-treated MPC-83 cells were used as an in vitro cellular model for AP. The expression levels of mmu_circ_0000037, microRNA (miR)-92a-3p, and protein inhibitor of activated STAT1 (Pias1) were detected by quantitative real-time polymerase chain reaction. Cell viability, amylase activity, apoptosis, and inflammatory response were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, Amylase Assay Kit, flow cytometry, and enzyme-linked immunosorbent assays. The protein level was quantified by western blot analysis. The target interaction between miR-92a-3p and mmu_circ_0000037 or Pias1 were predicted by StarbaseV3.0 and validated by dual-luciferase reporter assay and RNA immunoprecipitation assay.. Mmu_circ_0000037 and Pias1 levels were decreased, whereas miR-92a-3p expression was elevated in caerulein-induced MPC-83 cells. Overexpression of mmu_circ_0000037 protected MPC-83 cells from caerulein-induced the decrease of cell viability, as well as the promotion of amylase activity, apoptosis and inflammation. MiR-92a-3p was targeted by mmu_circ_0000037, and miR-92a-3p overexpression rescued the effect of mmu_circ_0000037 on caerulein-induced MPC-83 cell injury. Pias1 was confirmed as a target of miR-92a-3p and mmu_circ_0000037 regulated the expression of Pias1 by sponging miR-92a-3p.. Mmu_circ_0000037 relieves caerulein-induced inflammatory injury in MPC-83 cells by targeting miR-92a-3p/Pias1 axis, providing a theoretical basis for the treatment of AP.

Topics: Acute Disease; Amylases; Ceruletide; Humans; MicroRNAs; Pancreatitis; Protein Inhibitors of Activated STAT; RNA, Circular; Small Ubiquitin-Related Modifier Proteins

Hypertriglyceridemia (HTG) is one of the common causes of acute pancreatitis (AP). Hyperlipidemic acute pancreatitis (HTG-AP) is associated with higher mortality owing to its tendency for greater severity and rapid progression. The purpose of this study was to explore the mechanism of involvement of tumor necrosis factor receptor-related factor 6 (TRAF6) in pyroptosis during HTG-AP.. The HTG environment was simulated with palmitic acid treatment in vitro and a high-fat diet in vivo. Cerulein was used to establish the HTG-AP model, followed by genetic and pharmacological inhibition of TRAF6. Pyroptosis activation, inflammatory reaction, and the interaction between TRAF6 and pyroptosis in HTG-AP were assessed.. HTG was found to aggravate the development of pancreatitis, accompanied by increased pyroptosis and enhanced inflammatory response in HTG-AP models. Mechanistically, TRAF6 downregulation decreased the activation of pyroptosis in cerulein-induced HTG-AP.. Collectively, inhibition of TRAF6 improved HTG-AP and the associated inflammation by alleviating pyroptosis.

Topics: Acute Disease; Animals; Ceruletide; Hypertriglyceridemia; Inflammation; Pancreatitis; Pyroptosis; Rats; TNF Receptor-Associated Factor 6

Acute pancreatitis (AP) is an inflammatory process unexpectedly occurring in the pancreas, imposing a substantial burden on healthcare systems. Herein, we aimed to clarify the mechanism of action of phospholipase D2 (PLD2) in cerulein-treated AR42J cells, affording valuable insights into the treatment of AP.. The levels of PLD2, miR-5132-5p, inflammatory factors (interleukin [IL]-10, IL-6, and tumor necrosis factor-α), caspase-3 activity, and apoptosis-related proteins (Bax and Bcl-2) in cerulein-treated AR42J cells were detected using reverse transcription-quantitative polymerase chain, caspase-3 activity, and Western blot analysis. Protein levels of nuclear Factor erythroid 2-Related Factor 2 (Nrf2) and nuclear factor-k-gene binding (NF-κB) were detected by Western blot analysis. TargetScan predicted upstream microRNAs (miRNAs) of PLD2, and the interaction between miR-5132-5p and PLD2 was verified using a luciferase assay.. In cerulein-treated AR42J cells, PLD2 levels were downregulated, while miR-5132-5p expression was upregulated. Overexpression of PLD2 attenuated the cerulein-mediated facilitatory effect on inflammation and apoptosis in AR42J cells by regulating the Nrf2/NFκB pathway. Luciferase reporter analysis revealed that miR-5132-5p targeted PLD2, and miR-5132-5p negatively regulated PLD2. Upregulation of miR-5132-5p expression exacerbated inflammation and apoptosis and reversed the protective effect of PLD2 overexpression on AP.. PLD2 targeted by miR-5132-5p can attenuate cerulein-induced AP in AR42J cells via the Nrf2/NFκB pathway, providing therapeutic targets for patients with AP.

Topics: Acute Disease; Animals; Caspase 3; Ceruletide; Inflammation; MicroRNAs; NF-E2-Related Factor 2; NF-kappa B; Pancreatitis; Rats

Acute pancreatitis (AP) is an inflammatory condition of the pancreas characterized by oxidative stress and inflammation in its pathophysiology. Acetyl-11-keto-β-boswellic acid (AKBA) is an active triterpenoid with antioxidant activity. This article seeks to assess the impact of AKBA on AP and investigate its underlying mechanisms.. Our results confirm that AKBA exerts protective effects against AP in mice by inhibiting oxidative stress in macrophages through the Nrf2/HO-1 Pathway.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Lipase; Macrophages; Mice; Molecular Docking Simulation; NF-E2-Related Factor 2; Oxidative Stress; Pancreatitis; Reactive Oxygen Species

Acute pancreatitis (AP) is a severe disease with high morbidity and mortality in which inflammation and coagulation play crucial roles. The development of inflammation leads to vascular injury, endothelium and leukocytes stimulation, and an increased level of tissue factor, which results in the activation of the coagulation process. For this reason, anticoagulants may be considered as a therapeutic option in AP. Previous studies have shown that pretreatment with heparin, low-molecular-weight heparin (LMWH), or acenocoumarol inhibits the development of AP. The aim of the present study was to check if pretreatment with warfarin affects the development of edematous pancreatitis evoked by cerulein. Warfarin (90, 180, or 270 µg/kg/dose) or saline were administered intragastrically once a day for 7 days consecutively before the induction of AP. AP was evoked by the intraperitoneal administration of cerulein. The pre-administration of warfarin at doses of 90 or 180 µg/kg/dose reduced the histological signs of pancreatic damage in animals with the induction of AP. Additionally, other parameters of AP, such as an increase in the serum activity of lipase and amylase, the plasma concentration of D-dimer, and interleukin-1β, were decreased. In addition, pretreatment with warfarin administered at doses of 90 or 180 µg/kg/dose reversed the limitation of pancreatic blood flow evoked by AP development. Warfarin administered at a dose of 270 µg/kg/dose did not exhibit a preventive effect in cerulein-induced AP. Conclusion: Pretreatment with low doses of warfarin inhibits the development of AP evoked by the intraperitoneal administration of cerulein.

Topics: Acute Disease; Animals; Ceruletide; Heparin, Low-Molecular-Weight; Inflammation; Pancreatitis; Rats; Rats, Wistar; Warfarin

The pathogenesis of acute pancreatitis mainly involves NLRP3 inflammasome-mediated pancreatic cell injury, although regulators of this inflammasome machinery are still not fully identified. Membrane-associated RING-CH 9 (MARCH9) is a member of MARCH-type finger proteins, which regulates innate immunity through catalyzing polyubiquitination of critical immune factors. The aim of present research is to examine the function of MARCH9 in acute pancreatitis.. Cerulein-induced acute pancreatitis was established on pancreatic cell line AR42J and rat model. Reactive oxygen species (ROS) accumulation and NLRP3 inflammasome-dependent cell pyroptosis in pancreas were examined by flow cytometry.. MARCH9 was downregulated by cerulein, but overexpressing MARCH9 could inhibit NLRP3 inflammasome activation and ROS accumulation, thus suppressing pancreatic cell pyroptosis and mitigating pancreatic injury. We further uncovered that the mechanism underlying such an effect of MARCH9 is through mediating the ubiquitination of NADPH oxidase-2, whose deficiency reduces cellular ROS accumulation and inflammasome formation.. Our results suggested that MARCH9 suppresses NLRP3 inflammasome-mediated pancreatic cell injury through mediating the ubiquitination and degradation of NADPH oxidase-2, which compromises ROS generation and NLRP3 inflammasomal activation.

Topics: Acute Disease; Animals; Ceruletide; Inflammasomes; NADPH Oxidases; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreas; Pancreatic Hormones; Pancreatitis; Pyroptosis; Rats; Reactive Oxygen Species

Acute pancreatitis (AP), which is characterized by self-digestion of the pancreas by its own prematurely activated digestive proteases, is a major reason for hospitalization. The autodigestive process causes necrotic cell death of pancreatic acinar cells and the release of damage associated molecular pattern which activate macrophages and drive the secretion of pro-inflammatory cytokines. The MYD88/IRAK signaling pathway plays an important role for the induction of inflammatory responses. Interleukin-1 receptor associated kinase-3 (IRAK3) is a counter-regulator of this pathway. In this study, we investigated the role of MYD88/IRAK using Irak3-/- mice in two experimental animal models of mild and severe AP. IRAK3 is expressed in macrophages as well as pancreatic acinar cells where it restrains NFκB activation. Deletion of IRAK3 enhanced the migration of CCR2

Topics: Acute Disease; Adaptor Proteins, Signal Transducing; Animals; Ceruletide; Disease Models, Animal; Inflammation; Mice; Mice, Inbred C57BL; Myeloid Differentiation Factor 88; Necrosis; Pancreas; Pancreatitis; Patient Acuity; Signal Transduction; Systemic Inflammatory Response Syndrome

With the number of patients with acute pancreatitis (AP) increasing year by year, it is pressing to explore new key genes and markers for the treatment of AP. miR-455-3p/solute carrier family 2 member 1 (Slc2a1) obtained through bioinformatics analysis may participate in the progression of AP.. The C57BL/6 mouse model of AP was constructed for subsequent studies. Through bioinformatics analysis, the differentially expressed genes related to AP were screened and hub genes were identified. A caerulein-induced AP animal model was constructed to detect the pathological changes of mouse pancreas by HE staining. The concentrations of amylase and lipase were measured. Primary mouse pancreatic acinar cells were isolated and subjected to microscopy to observe their morphology. The enzymatic activities of trypsin and amylase were detected. The secretion of inflammatory cytokines in mouse were measured with the ELISA kits of TNF-. A total of five (Fyn, Gadd45a, Sdc1, Slc2a1, and Src) were identified by bioinformatics analysis, and miR-455-3p/Slc2a1 were further studied. HE staining results showed that the AP models were successfully established by caerulein induction. In mice with AP, the expression of miR-455-3p was reduced, while that of Slc2a1 was increased. In the caerulein-induced cell model, the expression of Slc2a1 was significantly reduced after intervention of miR-455-3p mimics, whereas increased after miR-455-3p inhibitor treatment. miR-455-3p decreased the secretion of inflammatory cytokines in the cell supernatant, reduced the activity of trypsin and amylase, and alleviated the cell damage induced by caerulein. In addition, Slc2a1 3'UTR region was bound by miR-455-3p, and its protein expression was also regulated.. miR-455-3p alleviated caerulein-induced mouse pancreatic acinar cell damage by regulating the expression of Slc2a1.

Topics: Acinar Cells; Acute Disease; Amylases; Animals; Ceruletide; Cytokines; Mice; Mice, Inbred C57BL; MicroRNAs; Pancreatitis; Trypsin

This research discussed the specific mechanism by which PIAS1 affects acute pancreatitis (AP).. PIAS1, Foxa2, and FTO expression was assessed in Cerulein-induced AR42J cells and mice. Loss- and gain-of-function assays and Cerulein induction were conducted in AR42J cells and mice for analysis. The relationship among PIAS1, Foxa2, and FTO was tested. Cell experiments run in triplicate, and eight mice for each animal group.. Cerulein-induced AP cells and mice had low PIAS1 and Foxa2 and high FTO. Cerulein induced pancreatic injury in mice and inflammation and oxidative stress in pancreatic tissues, which could be reversed by PIAS1 or Foxa2 upregulation or FTO downregulation. PIAS1 elevated SUMO modification of Foxa2 to repress FTO transcription. FTO upregulation neutralized the ameliorative effects of PIAS1 or Foxa2 upregulation on Cerulein-induced AR42J cell injury, inflammation, and oxidative stress.. PIAS1 upregulation diminished FTO transcription by increasing Foxa2 SUMO modification, thereby ameliorating Cerulein-induced AP.

Topics: Acute Disease; Alpha-Ketoglutarate-Dependent Dioxygenase FTO; Animals; Ceruletide; Down-Regulation; Hepatocyte Nuclear Factor 3-beta; Inflammation; Mice; Pancreatitis; Sumoylation; Up-Regulation

Acute pancreatitis (AP) is an inflammatory disease of the exocrine pancreas. Disruptions in organelle homeostasis, including macroautophagy/autophagy dysfunction and endoplasmic reticulum (ER) stress, have been implicated in human and rodent pancreatitis. Syntaxin 17 (STX17) belongs to the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) subfamily. The Qa-SNARE STX17 is an autophagosomal SNARE protein that interacts with SNAP29 (Qbc-SNARE) and the lysosomal SNARE VAMP8 (R-SNARE) to drive autophagosome-lysosome fusion. In this study, we investigated the role of STX17 in the pathogenesis of AP in male mice or rats induced by repeated intraperitoneal injections of cerulein. We showed that cerulein hyperstimulation induced AP in mouse and rat models, which was characterized by increased serum amylase and lipase activities, pancreatic edema, necrotic cell death and the infiltration of inflammatory cells, as well as markedly decreased pancreatic STX17 expression. A similar reduction in STX17 levels was observed in primary and AR42J pancreatic acinar cells treated with CCK (100 nM) in vitro. By analyzing autophagic flux, we found that the decrease in STX17 blocked autophagosome-lysosome fusion and autophagic degradation, as well as the activation of ER stress. Pancreas-specific STX17 knockdown using adenovirus-shSTX17 further exacerbated pancreatic edema, inflammatory cell infiltration and necrotic cell death after cerulein injection. These data demonstrate a critical role of STX17 in maintaining pancreatic homeostasis and provide new evidence that autophagy serves as a protective mechanism against AP.

Topics: Acute Disease; Animals; Autophagy; Ceruletide; Disease Models, Animal; Edema; Humans; Male; Mice; Pancreatitis; Rats; SNARE Proteins

Ursolic acid (UA) is found in many plants, and has been reported to have anti-protease, antioxidant, anti-inflammatory, antimicrobial, nephroprotective, hepatoprotective, and cardioprotective effects.. The purpose of this study was to investigate the effects of ursolic acid in cerulein-induced acute pancreatitis (AP).. Thirty-two Wistar albino rats were randomly assigned to 4 equal groups: Sham, acute pancreatitis, treatment, and ursolic acid group.. Serum amylase levels in the AP and treatment groups were significantly higher than in the others (p < 0.05). In addition, serum IL-1β, IL-6, and TNF-α levels were significantly higher in the AP group in comparison with the treatment group. Although pancreatic tissue total oxidant activity in the AP and treatment groups was similar, pancreatic tissue total antioxidant capacity was significantly higher in the treatment group than in the AP group.. Damage to the pancreas and remote organs in AP was observed to be reduced by UA. In addition, oxidative stress was observed to be decreased by the effect of UA.. El ácido ursólico se encuentra en numerosas plantas y se ha informado que tiene efectos antiproteasas, antioxidantes, antiinflamatorios, antimicrobianos, nefroprotectores, hepatoprotectores y cardioprotectores.. Este estudio tuvo como objetivo investigar los efectos del ácido ursólico en la pancreatitis aguda inducida por ceruleína.. Treinta y dos ratas albinas Wistar fueron asignadas aleatoriamente a cuatro grupos iguales: grupo simulado, grupo de pancreatitis aguda, grupo de tratamiento y grupo de ácido ursólico.. Los niveles de amilasa sérica en los grupos de pancreatitis aguda y de tratamiento fueron significativamente más altos que en los otros grupos (p < 0.05). Además, los niveles séricos de IL-1β, IL-6 y TNF-α fueron significativamente más altos en el grupo de pancreatitis aguda en comparación con el grupo de tratamiento. Aunque la actividad oxidante total del tejido pancreático en ambos grupos fue similar, la capacidad antioxidante total del tejido pancreático en el grupo de tratamiento fue significativamente mayor.. Se observó que el ácido ursólico reduce el daño al páncreas y órganos remotos en la pancreatitis aguda, al igual que el estrés oxidativo.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Pancreatitis; Rats; Rats, Wistar; Triterpenes; Ursolic Acid

Chronic pancreatitis (CP) is associated with serious complications and reduced quality of life. Kidney failure is a frequent complication of acute pancreatitis (AP), however limited information is available regarding the impact of CP on this condition. In the kidney, 9 aquaporins (AQPs) are expressed to maintain body water homeostasis and concentrate urine. The purpose of this study was to morphologically assess and analyze the location and expression of AQP2, AQP3 and AQP4 and determine whether CP affects renal structure and expression of AQPs in collecting duct (CD) principal cells.. CP was induced in domestic pigs through intramuscular injections of cerulein (1 ​μg/kg ​bw/day for 6 days; n ​= ​5); pigs without CP (n ​= ​5) were used as a control group. Kidney samples were collected 6 weeks after the last injection and subjected to histological examination. Expression of AQPs was determined by immunohistochemistry and Western blot.. The kidneys of animals with CP exhibited moderate changes, including glomerular enlargement, increased collagen percentage, numerous stromal erythrorrhages and inflammatory infiltrations compared to control group. Although the total abundance of AQP2 in the CD decreased in pigs after cerulein administration, the difference was not statistically significant. Expression of AQP3 and AQP4 was limited to the basolateral membrane of the CD cells. AQP4 abundance remained relatively stable in both groups, while AQP3 expression increased nearly three-fold in pigs with CP.. This study identified morphological alterations and a statistically significant increase in the expression of renal AQP3 when pigs developed CP.

Topics: Acute Disease; Animals; Aquaporin 2; Aquaporin 3; Ceruletide; Kidney; Pancreatitis, Chronic; Quality of Life; Swine

Nafamostat mesilate (NM), a synthetic broad-spectrum serine protease inhibitor, has been commonly used for treating acute pancreatitis (AP) and other inflammatory-associated diseases in some East Asia countries. Although the potent inhibitory activity against inflammation-related proteases (such as thrombin, trypsin, kallikrein, plasmin, coagulation factors, and complement factors) is generally believed to be responsible for the anti-inflammatory effects of NM, the precise target and molecular mechanism underlying its anti-inflammatory activity in AP treatment remain largely unknown.. The protection of NM against pancreatic injury and inhibitory effect on the NOD-like receptor protein 3 (NLRP3) inflammasome activation were investigated in an experimental mouse model of AP. To decipher the molecular mechanism of NM, the effects of NM on nuclear factor kappa B (NF-κB) activity and NF-κB mediated NLRP3 inflammasome priming were examined in lipopolysaccharide (LPS)-primed THP-1 cells. Additionally, the potential of NM to block the activity of histone deacetylase 6 (HDAC6) and disrupt the association between HDAC6 and NLRP3 was also evaluated.. NM significantly suppressed NLRP3 inflammasome activation in the pancreas, leading to a reduction in pancreatic inflammation and prevention of pancreatic injury during AP. NM was found to interact with HDAC6 and effectively inhibit its function. This property allowed NM to influence HDAC6-dependent NF-κB transcriptional activity, thereby blocking NF-κB-driven transcriptional priming of the NLRP3 inflammasome. Furthermore, NM exhibited the potential to interfere the association between HDAC6 and NLRP3, impeding HDAC6-mediated intracellular transport of NLRP3 and ultimately preventing NLRP3 inflammasome activation.. Our current work has provided valuable insight into the molecular mechanism underlying the immunomodulatory effect of NM in the treatment of AP, highlighting its promising application in the prevention of NLRP3 inflammasome-associated inflammatory pathological damage.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Ceruletide; Histone Deacetylase 6; Inflammasomes; Inflammation; Mice; NF-kappa B; NLR Family, Pyrin Domain-Containing 3 Protein; NLR Proteins; Pancreatitis

Various organosulfur compounds, such as dimethyl trisulfide (DMTS), display anti-inflammatory properties. We aimed to examine the effects of DMTS on acute pancreatitis (AP) and its mechanism of action in both in vivo and in vitro studies. AP was induced in FVB/n mice or Wistar rats by caerulein, ethanol-palmitoleic acid, or L-ornithine-HCl. DMTS treatments were administered subcutaneously. AP severity was assessed by pancreatic histological scoring, pancreatic water content, and myeloperoxidase activity measurements. The behaviour of animals was followed. Pancreatic heat shock protein 72 (HSP72) expression, sulfide, and protein persulfidation were measured. In vitro acinar viability, intracellular Ca

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Mice; Pancreas; Pancreatitis; Rats; Rats, Wistar; Sulfides

Acute pancreatitis (AP) is an inflammatory disease of the pancreas. Iron is an essential element for life and is involved in many metabolic processes. Ferroptosis is a type of regulated cell death that is triggered by iron and oxidative stress. A well-established mouse AP model was adopted to study the role of iron and ferroptosis in the pathogenesis of pancreatitis. Mice were injected with cerulein to induce AP, and pancreatic tissue samples were analyzed to determine the pathology, cell death, iron deposition, expression of iron transporters, and lipid peroxidation. The role of iron was studied by giving mice extra iron or iron chelator. In vitro studies with acinar cells with ferroptosis activator and inhibitor were also performed to assess the inflammatory response. Iron was found accumulated in the pancreatic tissue of mice who suffered cerulein-induced pancreatitis. Cell death and lipid peroxidation increased in these tissues and could be further modulated by iron dextran or iron chelator. Mice given Hemin through gavage had reduced levels of GSH in pancreatic tissue and increased inflammatory response. Studies with acinar cells showed increased levels of lipid peroxidation and ferroptosis-specific mitochondrial damage when treated with ferroptosis inducer and inflammatory cytokines.

Topics: Acute Disease; Animals; Ceruletide; Ferroptosis; Iron; Iron Chelating Agents; Mice; Pancreatitis

Ubiquitin-binding associated protein 2 (UBAP2) is reported to promote macropinocytosis and pancreatic adenocarcinoma (PDAC) growth, however, its role in normal pancreatic function remains unknown. We addressed this knowledge gap by generating UBAP2 knockout (U2KO) mice under a pancreas-specific Cre recombinase (Pdx1-Cre). Pancreatic architecture remained intact in U2KO animals, but they demonstrated slight glucose intolerance compared to controls. Upon cerulein challenge to induce pancreatitis, U2KO animals had reduced levels of several pancreatitis-relevant cytokines, amylase and lipase in the serum, reduced tissue damage, and lessened neutrophil infiltration into the pancreatic tissue. Mechanistically, cerulein-challenged U2KO animals revealed reduced NF-κB activation compared to controls. In vitro promoter binding studies confirmed the reduction of NF-κB binding to its target molecules supporting UBAP2 as a new regulator of inflammation in pancreatitis and may be exploited as a therapeutic target in future to inhibit pancreatitis.

Topics: Acute Disease; Adenocarcinoma; Animals; Ceruletide; Glucose; Inflammation; Mice; NF-kappa B; Pancreas; Pancreatic Neoplasms; Pancreatitis

Acute pancreatitis (AP) is the most common gastrointestinal disease leading to hospitalizations and unexpected deaths. The development of AP leads to damage of the pancreatic microcirculation with a cascade of subsequent events resulting, among others, in coagulopathy. Previous research showed that anticoagulants can be important therapeutic agents. Heparin and acenocoumarol can alleviate the course of AP, as well as accelerate healing and post-inflammatory regeneration of the pancreas. The aim of this study was to determine whether warfarin, a drug with more stable effects than acenocoumarol, affects the healing and regeneration of the pancreas in the cerulein-induced AP. AP was evoked in Wistar male rats by intraperitoneal administration of cerulein. The first dose of warfarin (45, 90 or 180 μg/kg) was administered 24 hours after the first dose of cerulein and the doses of warfarin were repeated once a day in subsequent 10 days. The severity of AP was assessed immediately after the last dose of cerulein, as well as at days 1, 2, 3, 5, and 10 after AP induction. Treatment with warfarin dose-dependently increased international normalized ratio (INR) and attenuated the severity of pancreatitis in histological examination and accelerated pancreatic recovery. These effects were accompanied with a faster reduction in the AP-evoked increase in serum activity of amylase and lipase, the serum concentration of pro-inflammatory interleukin-1β, and the plasma level of D-Dimer. In addition, treatment with warfarin decreased pancreatic weight (an index of pancreatic edema) and improved pancreatic blood flow in rats with AP. The therapeutic effect was particularly pronounced after the administration of warfarin at a dose of 90 μg/kg. We conclude that treatment with warfarin accelerated regeneration of the pancreas and recovery in the course of cerulein-induced mild-edematous acute pancreatitis.

Topics: Acenocoumarol; Acute Disease; Animals; Ceruletide; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Warfarin

Recent clinical studies have shown that serum high-density lipoprotein (HDL) levels are correlated with acute pancreatitis (AP) severity. We aimed to investigate the role of HDL in pancreatic necrosis in AP.. ApoA-I is the main constitution and function component of HDL. The roles of healthy human-derived HDL and apoA-I mimic peptide D4F were demonstrated in AP models in vivo and in vitro. Constitutive Apoa1 genetic inhibition on AP severity, especially pancreatic necrosis was assessed in both caerulein and sodium taurocholate induced mouse AP models. In addition, constitutive (Casp1. Apoa1 knockout dramatically aggravated pancreatic necrosis. Human-derived HDL protected against acinar cell death in vivo and in vitro. We found that mimic peptide D4F also protected against AP very well. Constitutive Casp1 or acinar cell-conditional Nlrp3 and Gsdmd genetic inhibition could counteract the protective effects of HDL, implying HDL may exert beneficial effects on AP through inhibiting acinar cell pyroptosis.. This work demonstrates the protective role of HDL and apoA-I in AP pathology, potentially driven by the inhibition of NLRP3 inflammasome signaling and acinar cell pyroptosis. Mimic peptides have promise as specific therapies for AP.

Topics: Acinar Cells; Acute Disease; Animals; Apolipoprotein A-I; Caspase 1; Ceruletide; Humans; Inflammasomes; Mice; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreatitis, Acute Necrotizing; Pyroptosis

Acute pancreatitis is a common and serious inflammatory condition currently lacking disease modifying therapy. The cholinergic anti-inflammatory pathway (CAP) is a potent protective anti-inflammatory response activated by vagus nerve-dependent α7 nicotinic acetylcholine receptor (α7nAChR) signaling using splenic CD4. The effect of galantamine (1-6 mg/kg-body weight) on caerulein-induced acute pancreatitis was evaluated in mice. Two hours following 6 hourly doses of caerulein (50 µg/kg-body weight), organ and serum analyses were performed with accompanying pancreatic histology. Experiments utilizing vagotomy, gene knock out (KO) technology and the use of nAChR antagonists were also performed.. Galantamine attenuated pancreatic histologic injury which was mirrored by a reduction in serum amylase and pancreatic inflammatory cytokines and an increase the anti-inflammatory cytokine IL-10 in the serum. These beneficial effects were not altered by bilateral subdiaphragmatic vagotomy, KO of either choline acetyltransferase. Galantamine improves acute pancreatitis via a mechanism which does not involve previously established physiological and molecular components of the CAP. As galantamine is an approved drug in widespread clinical use with an excellent safety record, our findings are of interest for further evaluating the potential benefits of this drug in patients with acute pancreatitis.

Topics: Acetylcholinesterase; Acute Disease; alpha7 Nicotinic Acetylcholine Receptor; Animals; Anti-Inflammatory Agents; Body Weight; Ceruletide; Cytokines; Galantamine; Humans; Mice; Pancreatitis

Circular RNA hsa_circ_0073748 (circ_0073748) is upregulated in patients with acute pancreatitis (AP), a clinically common sudden inflammatory response. MicroRNA (miR)-132-3p is a stress-induced factor with high conservation between species. Herein, expression and role of circ_0073748 and miR-132-3p in caerulein-induced pancreatitis were studied. Expression levels of circ_0073748, miR-132-3p, TNF receptor associated factor 3 (TRAF3), Bcl-2 and Bcl-2-associated X protein (Bax) were examined by reverse transcription-quantitative PCR and Western blotting. Cell proliferation was measured by MTS and EdU assays. Flow cytometry and assay kits detected apoptosis, inflammatory, and oxidative responses. Western blotting detected nuclear factor (NF)-κB signaling pathway. Circ_0073748 was upregulated and miR-132-3p was downregulated in AP patients' plasma and human pancreatic ductal HPDE6-C7 cells with caerulein induction. Interfering circ_0073748 and reinforcing miR-132-3p improved cell viability, EdU incorporation, and superoxide dismutase (SOD) activity of caerulein-treated HPDE6-C7 cells but suppressed malonaldehyde (MDA), IL-6 and TNF-α levels and apoptosis rate. Moreover, TRAF3 downregulation was allied with circ_0073748 silencing and miR-132-3p overexpression in caerulein-induced HPDE6-C7 cells. Mechanically, circ_0073748 was identified as a sponge for miR-132-3p to modulate TRAF3 expression, thus establishing a competitive endogenous RNA (ceRNA) regulation model. Notably, circ_0073748 blockage could suppress expressions of phosphorylated P65 (p-P65) and p-IκB in caerulein-induced HPDE6-C7 cells by promoting miR-132-3p and inhibiting TRAF3. Silencing circ_0073748 and upregulating miR-132-3p could alleviate caerulein-induced HPDE6-C7 injury and inactivate canonical NF-κB signal by inhibiting TRAF3. Circ_0073748/miR-132-3p/TRAF3 ceRNA pathway might be one underlying mechanism and therapeutic target of caerulein-induced AP.

Topics: Acute Disease; Apoptosis; Ceruletide; Humans; MicroRNAs; NF-kappa B; Pancreatitis; TNF Receptor-Associated Factor 3

T7K24R mice carry mutation p.K24R in mouse cationic trypsinogen (isoform T7), which is analogous to the human hereditary pancreatitis-associated mutation p.K23R. The mutation renders trypsinogen more prone to autoactivation. We recently reported that T7K24R mice exhibit increased severity of acute pancreatitis induced by repeated cerulein injections. The objective of the present study was to test whether trypsinogen mutant mice are prone to develop chronic pancreatitis, as observed in patients. We characterized the natural course of cerulein-induced pancreatitis in T7K24R mice and the C57BL/6N parent strain from the acute episode to 3 months post-attack. As expected, an acute episode of pancreatitis in C57BL/6N mice was followed by rapid recovery and histological restitution. In stark contrast, T7K24R mice developed progressive chronic pancreatitis with acinar cell atrophy, persistent macrophage infiltration, and diffuse fibrosis. The nadir of pancreas damage occurred on days 5-6 after the acute episode and was accompanied by digestive dysfunction. Remarkably, histological recovery was markedly delayed and permanent, chronic changes were still detectable 1-3 months after the acute pancreatitis episode. We conclude that during cerulein-induced acute pancreatitis in T7K24R mice, trypsin triggers an autonomous inflammatory program resulting in chronic disease progression, even after the cessation of cerulein-mediated injury. We propose that this uniquely trypsin-dependent mechanism explains the development of hereditary chronic pancreatitis in humans. Trypsin inhibition during acute attacks should prevent or delay progression to chronic disease.

Topics: Acute Disease; Animals; Ceruletide; Humans; Mice; Mice, Inbred C57BL; Pancreas; Pancreatitis; Trypsinogen

The anti-inflammatory and anti-apoptotic properties of crocetin have been widely demonstrated in numerous diseases. However, the exact role and mechanism of crocetin in acute pancreatitis have not been elucidated. Thus, this paper aims at exploring whether crocetin could be used to alleviate acute pancreatitis and further demonstrating the underlying mechanisms. Cell viability of caerulein-induced pancreatic exocrine cell line AR42J treated with crocetin was determined by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT). Apoptosis and inflammation of these treated cells were detected by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL), western blot and enzyme linked immunosorbent assay (ELISA). The expression of sirtuin-1 (SIRT1) was quantified by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot. After knockdown of SIRT1, cell viability, apoptosis and inflammation were measured again by corresponding kits. Finally, the NF-κB nuclear translocation and proteins in the NF-κB signaling were examined. Crocetin remarkably suppressed the apoptosis and inflammation of caerulein-induced AR42J cells. The decreased expression of SIRT1 was increased in caerulein-induced AR42J cells after exposure to crocetin. After knockdown of SIRT1, the alleviative effects of crocetin were found to be canceled in these cells. Furthermore, SIRT1 knockdown promoted the NF-κB signal transduction. On the whole, we presented the first evidence for the importance of SIRT1-NF-κB axis in acute pancreatitis and proposed that crocetin alleviates the caerulein-induced apoptosis and inflammation in AR42J cells by activating SIRT1 via NF-κB.

Topics: Acute Disease; Apoptosis; Carotenoids; Ceruletide; Humans; Inflammation; NF-kappa B; Pancreatitis; Sirtuin 1; Vitamin A

Acute pancreatitis (AP) is an acute inflammatory disease that can lead to death. Mir-325-3p is strongly and abnormally expressed in many diseases, necessitating exploration of its function and mechanism in AP.. Blood samples from AP patients and mice were analyzed. The expression levels of miR-325-3p in AP patients and mouse were detected. Whether miR-325-3p targets RIPK3 gene was predicted by TargetScan online database and dual luciferase reporter assay. In vitro experiments verified the effect of miR-325-3p overexpression on caerulein-induced MPC83 pancreatic acinar cancer cell line. In vivo experiments verified the effect of overexpression of miR-325-3p on the disease degree of pancreatic tissues in AP mice.. Analysis of blood samples from AP patients and experiments in mice demonstrated that expression of miR-325-3p was significantly reduced during the process of AP in humans and mice. Predicted using the TargetScan online database and through dual luciferase reporter assay detection, miR-325-3p directly targets the RIPK3 gene. In vitro experiments revealed that overexpression of miR-325-3p reversed caerulein-induced apoptosis and necroptosis in MPC83 pancreatic acinar cancer cell line. We used Z-VAD-FMK to assess necroptosis and demonstrated that miR-325-3p targets necroptosis to reduce cell damage. In subsequent experiments in mice, we verified that overexpression of miR-325-3p reduces inflammation, edema, hemorrhage, and necrosis in acute pancreatitis. Characteristic western blot, immunohistochemistry, and transmission electron microscopy results revealed that overexpression of miR-325-3p reduces the severity of acute pancreatitis by inhibiting pancreatic necroptosis in AP mice.. The current research results indicate that miR-325-3p directly targets RIPK3 and exerts a protective role in mouse AP. Necroptosis is still the primary mechanism of RIPK3 regulation. MiR-325-3p inhibits acute pancreatitis by targeting RIPK3-dependent necroptosis, which may represent a novel treatment method for acute pancreatitis.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Humans; Mice; MicroRNAs; Pancreatitis; Receptor-Interacting Protein Serine-Threonine Kinases

Triptolide (TP), the main active ingredient of

Topics: Acute Disease; Animals; Antioxidants; Ceruletide; China; Disease Models, Animal; Diterpenes; Epoxy Compounds; Gene Expression Regulation; Hep G2 Cells; Humans; Inflammation; Male; Mice; Mice, Inbred ICR; NF-E2-Related Factor 2; NF-kappa B; Oxidative Stress; Pancreas; Pancreatitis; Phenanthrenes; Reactive Oxygen Species

The excessive inflammatory response mediated by macrophage is one of the key factors for the progress of acute pancreatitis (AP). Paeonol (Pae) was demonstrated to exert multiple anti-inflammatory effects. However, the role of Pae on AP is not clear. In the present study, we aimed to investigate the protective effect and mechanism of Pae on AP in vivo and vitro. In the caerulein-induced mild acute pancreatitis (MAP) model, we found that Pae administration reduced serum levels of amylase, lipase, IL-1β and IL-6 and alleviated the histopathological manifestations of pancreatic tissue in a dose-dependent manner. And Pae decrease the ROS generated, restore mitochondrial membrane potential (ΔΨm), inhibit M1 macrophage polarization and NLRP3 inflammasome in bone marrow-derived macrophages (BMDMs) in vitro. In addition, specific NLRP3 inhibitor MCC950 eliminated the protective effect of Pae on AP induced by caerulein in mice. Correspondingly, the inhibitory effect of Pae on ROS generated and M1 polarization was not observed in BMDMs with MCC950 in vitro. Taken together, our datas for the first time confirmed the protective effects of Pae on AP via the NLRP3 inflammasomes Pathway.

Topics: Acetophenones; Acute Disease; Animals; Ceruletide; Inflammasomes; Macrophages; Mice; Mice, Inbred C57BL; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreatitis; Reactive Oxygen Species

Proper mitochondrial function and adequate cellular ATP are necessary for normal pancreatic protein synthesis and sorting, maintenance of intracellular organelles and enzyme secretion. Inorganic phosphate is required for generating ATP and its limited availability may lead to reduced ATP production causing impaired Ca

Topics: Acute Disease; Adenosine Triphosphate; Animals; Ceruletide; Cholecystokinin; Hypophosphatemia; Ion Channels; Mice; Pancreas; Pancreatitis; Phosphates

Mounting evidence suggests that long non-coding RNAs (lncRNAs) and microRNAs exert a critical regulatory role in acute pancreatitis. The present study aimed to explore the role of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in acute pancreatitis (AP) that was induced by caerulein in rat pancreatic acinar cells (AR42J). The potential target sites of lncRNA NEAT1 and miR-365a-3p were predicted using starBase and were confirmed using dual-luciferase reporter assay. Reverse transcription-quantitative polymerase chain reaction was performed to assess lncRNA NEAT1 and miR-365a-3p expression levels in AP induced by caerulein. Cell Counting Kit-8 and flow cytometry assays were performed to assess AR42J cell viability. Western blotting was performed to evaluate the expression of apoptosis-related proteins. Interleukin (IL)-1β, IL-6, and tumor necrosis factor-α levels were detected by ELISA. The results of the dual-luciferase reporter assay confirmed that miR-365a-3p could bind to NEAT1. LncRNA NEAT1 was upregulated in AR42J cells treated with 10 nmol/l caerulein, and miR-365a-3p was expressed at low levels in an AP model. Overexpression of miR-365a-3p suppressed the apoptosis and inflammatory response of AR42J cells induced by caerulein. Importantly, inhibition of lncRNA NEAT1 decreased apoptosis and inflammation in caerulein-treated AR42J cells, while these effects were reverted upon co-transfection with a miR-365a-3p inhibitor. In conclusion, lncRNA NEAT1 was involved in AP progression by sponging miR-365a-3p and may thus be a novel target for treating patients with AP.

Topics: Acute Disease; Animals; Apoptosis; Ceruletide; Down-Regulation; MicroRNAs; Pancreatitis; Rats; RNA, Long Noncoding

Innate immunity and metabolites link to the pathogenesis and severity of acute pancreatitis (AP). However, liver metabolism and its role in immune response and AP progression remain elusive. We investigated the function of liver metabolism in the pathogenesis of AP.. Circulating ketone body β-hydroxybutyrate (βOHB) levels were determined in AP clinical cohorts and caerulein-induced AP (CER-AP) mouse models receiving seven (Cer*7) or twelve (Cer*12) injection regimens at hourly intervals. Liver transcriptomics and metabolomics were compared between CER-AP (Cer*7) and CER-AP (Cer*12). Inhibition of fatty acid β-oxidation (FAO)-ketogenesis, or supplementation of βOHB was performed in mouse models of AP. The effect and mechanism of βOHB were examined in vitro.. Elevated circulating βOHB was observed in patients with non-severe AP (SAP) but not SAP. These findings were replicated in CER-AP (Cer*7) and CER-AP (Cer*12), which manifested as limited and hyperactive immune responses, respectively. FAO-ketogenesis was activated in CER-AP (Cer*7), while impaired long-chain FAO and mitochondrial function were observed in the liver of CER-AP (Cer*12). Blockage of FAO-ketogenesis (Cpt1a antagonism or Hmgcs2 knockdown) worsened, while supplementation of βOHB or its precursor 1,3-butanediol alleviated the severity of CER-AP. Mechanistically, βOHB had a discernible effect on pancreatic acinar cell damage, instead, it greatly attenuated the activation of pancreatic and systemic proinflammatory macrophages via class I histone deacetylases.. Our findings reveal that hepatic ketogenesis is activated as an endogenous protective programme to restrain AP progression, indicating its potential therapeutic value.. This work was supported by the National Natural Science Foundation of China, Shanghai Youth Talent Support Programme, and Shanghai Municipal Education Commission-Gaofeng Clinical Medicine Grant.

Topics: 3-Hydroxybutyric Acid; Acute Disease; Adolescent; Animals; Ceruletide; China; Disease Models, Animal; Humans; Ketone Bodies; Macrophage Activation; Mice; Pancreatitis

A hybrid allele that originated from homologous recombination between CEL and its pseudogene (CELP), CEL-HYB1 increases the risk of chronic pancreatitis (CP). Although suggested to cause digestive enzyme misfolding, definitive in vivo evidence for this postulate has been lacking.. CRISPR-Cas9 was used to generate humanized mice harboring the CEL-HYB1 allele on a C57BL/6J background. Humanized CEL mice and C57BL/6J mice were used as controls. Pancreata were collected and analyzed by histology, immunohistochemistry, immunoblotting, and transcriptomics. Isolated pancreatic acini were cultured in vitro to measure the secretion and aggregation of CEL-HYB1 protein. Mice were given caerulein injections to induce acute pancreatitis (AP) and CP.. Pancreata from mice expressing CEL-HYB1 developed pathological features characteristic of focal pancreatitis that included acinar atrophy and vacuolization, inflammatory infiltrates, and fibrosis in a time-dependent manner. CEL-HYB1 expression in pancreatic acini led to decreased secretion and increased intracellular aggregation and triggered endoplasmic reticulum stress compared with CEL. The autophagy levels of pancreata from mice expressing CEL-HYB1 changed at different developmental stages; some aged CEL-HYB1 mice exhibited an accumulation of large autophagic vesicles and impaired autophagy in acinar cells. Administration of caerulein increased the severity of AP/CP in mice expressing CEL-HYB1 compared with control mice, accompanied by higher levels of endoplasmic reticulum stress.. Expression of a humanized form of CEL-HYB1 in mice promotes endoplasmic reticulum stress and pancreatitis through a misfolding-dependent pathway. Impaired autophagy appears to be involved in the pancreatic injury in aged CEL-HYB1 mice. These mice have the potential to be used as a model to identify therapeutic targets for CP.

Topics: Acute Disease; Alleles; Animals; Ceruletide; Mice; Mice, Inbred C57BL; Pancreatitis, Chronic

The expression of amino acid transporters is known to vary during acute pancreatitis (AP) except for LAT1 (. To evaluate the role of LAT1 in the development of and recovery from AP.. AP was induced with caerulein (cae) injections in female and male mice expressing LAT1 or after its knockout (LAT1 Cre/LoxP). The development of the initial AP injury and its recovery were followed for seven days after cae injections by daily measuring body weight, assessing microscopical tissue architecture, mRNA and protein expression, protein synthesis, and enzyme activity levels, as well as by testing the recruitment of immune cells by FACS and ELISA.. The initial injury, evaluated by measurements of plasma amylase, lipase, and trypsin activity, as well as the gene expression of dedifferentiation markers, did not differ between the groups. However, early metabolic adaptations that support regeneration at later stages were blunted in LAT1 knockout mice. Especially in females, we observed less mTOR reactivation and dysfunctional autophagy. The later regeneration phase was clearly delayed in female LAT1 knockout mice, which did not regain normal expression of the pancreas-specific differentiation markers recombining binding protein suppressor of hairless-like protein (rbpjl) and basic helix-loop-helix family member A15 (mist1). Amylase mRNA and protein levels remained lower, and, strikingly, female LAT1 knockout mice presented signs of fibrosis lasting until day seven. In contrast, pancreas morphology had returned to normal in wild-type littermates.. LAT1 supports the regeneration of acinar cells after AP. Female mice lacking LAT1 exhibited more pronounced alterations than male mice, indicating a sexual dimorphism of amino acid metabolism.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Female; Large Neutral Amino Acid-Transporter 1; Male; Mammals; Mice; Mice, Knockout; Pancreas; Pancreatitis; RNA, Messenger; TOR Serine-Threonine Kinases

Severe acute pancreatitis (SAP), as a typical acute inflammatory injury disease, is one of the acute gastrointestinal diseases with a remarkable mortality rate. Macrophages, typical inflammatory cells involved in SAP, play an important role in the pathogenesis of SAP, which are separated into proinflammation M1 and antiinflammation M2. Growth and differentiation factor 11 (GDF11), as a member of the TGF-β family also called BMP-11, has been discovered to suppress inflammation. However, the mechanism by which GDF11 inhibits inflammation and whether it can ameliorate SAP are still elusive. The present research aimed to investigate the roles of GDF11 in SAP and the potential immunomodulatory effect of macrophage polarization. The mouse and rat SAP model were constructed by caerulein and retrograde injection of sodium taurocholate respectively. The effects of GDF11 on SAP were observed by serology, histopathology and tissue inflammation, and the effects of GDF11 on the polarization of macrophages in vivo were observed. Raw264.7 and THP1 crells were used to study the effect of GDF11 on macrophage polarization in vitro. To further investigate the causal link underneath, our team first completed RNA and proteome sequencing, and utilized specific suppressor to determine the implicated signal paths. Herein, we discovered that GDF11 alleviated the damage of pancreatic tissues in cerulein induced SAP mice and SAP rats induced by retrograde injection of sodium taurocholate, and further found that GDF11 facilitated M2 macrophage polarization and diminished M1 macrophage polarization in vivo and in vitro. Subsequently, we further found that the regulation of GDF11 on macrophage polarization through TGFβR1/smad2 pathway. Our results revealed that GDF11 ameliorated SAP and diminished M1 macrophage polarization and facilitated M2 macrophage polarization. The Role of GDF11 in modulating macrophage polarization might be one of the mechanisms by which GDF11 played a protective role in pancreatic tissues during SAP.

Topics: Acute Disease; Animals; Ceruletide; Growth Differentiation Factors; Humans; Inflammation; Macrophage Activation; Macrophages; Mice; Pancreatitis; Rats; RAW 264.7 Cells; Receptor, Transforming Growth Factor-beta Type I; Smad2 Protein; Taurocholic Acid; THP-1 Cells

Pancreatitis occurs in two forms defined by its chronicity. Acute pancreatitis (AP) occurs suddenly and only lasts for several days. Consequently, most patients with AP recover without permanent damage to the pancreas, and about 20% of patients with AP have severe disease. In contrast, chronic pancreatitis (CP) is a long-lasting inflammation that causes permanent damage to pancreatic tissue; consequently, this form is marked by the emergence of persistent endocrine and exocrine pancreatic insufficiency. Despite these differences, AP and CP share central mechanisms of disease: in both forms, inflammation is initiated and/or sustained by the intrapancreatic activation of pancreatic digestive enzymes followed by the autodigestion of pancreatic tissues. In addition, in both forms enzymatic damage is accompanied by changes in intestinal permeability and entry of commensal organisms into the pancreas where they elicit innate immune responses that ultimately dominate and define pancreatic inflammation. In the murine models of AP and CP described here, both of these elements of pancreatitis pathogenesis are taken into account. Thus, in one approach mice are administered high doses of cerulein, a cholecystokinin analog with the ability at this dose to induce excessive activation of the cholecystokinin receptor expressed in pancreatic acinar cells and the release of active trypsin that causes both direct and indirect acinar damages due to entry of commensal organisms and stimulation of innate immune responses. In a second approach mice are administered low doses of cerulein, which causes little or no damage to the pancreas unless given along with nucleotide-binding oligomerization domain 1 (NOD1) ligand, which in the presence of low-dose cerulein administration induces a pathologic innate immune response mediated by NOD1. These approaches are adopted to produce AP when cerulein or cerulein plus NOD1 ligand is applied only once or to produce CP when a similar regimen is applied multiple times. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Cerulein-induced acute pancreatitis Alternate Protocol 1: Acute pancreatitis induced by cerulein and NOD1 ligand Basic Protocol 2: Cerulein-induced chronic pancreatitis Alternate Protocol 2: Chronic pancreatitis induced by cerulein and NOD1 ligand Support Protocol: Isolation of pancreatic mononuclear cells.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Humans; Inflammation; Ligands; Mice; Pancreatitis, Chronic

This study was to investigate the changes of autophagy in pancreatic tissue cells from hyperlipidemic acute pancreatitis (HLAP) rats and the molecular mechanism of autophagy to induce inflammatory injury in pancreatic tissue cells. Male Sprague Dawley (SD) rats were intraperitoneally injected with caerulein to establish acute pancreatitis (AP) model and then given a high fat diet to further prepare HLAP model. The HLAP rats were treated with autophagy inducer rapamycin or inhibitor 3-methyladenine. Pancreatic acinar (AR42J) cells were treated with caerulein to establish HLAP cell model. The HLAP cell model were treated with rapamycin or transfected with vascular endothelial growth factor (VEGF) siRNA. The inflammatory factors in serum and cell culture supernatant were detected by ELISA method. The histopathological changes of pancreatic tissue were observed by HE staining. The changes of ultrastructure and autophagy in pancreatic tissue were observed by electron microscopy. The expression levels of Beclin-1, microtubule- associated protein light chain 3-II (LC3-II), mammalian target of rapamycin complex 1 (mTORC1), and VEGF were measured by immunohistochemistry and Western blot. The results showed that, compared with control group, the autophagy levels and inflammatory injury of pancreatic tissue cells from HLAP model rats were obviously increased, and these changes were aggravated by rapamycin treatment, but alleviated by 3-methyladenine treatment. In HLAP cell model, rapamycin aggravated the autophagy levels and inflammatory injury, whereas VEGF siRNA transfection increased mTORC1 protein expression, thus alleviating the autophagy and inflammatory injury of HLAP cell model. These results suggest that VEGF-induced autophagy plays a key role in HLAP pancreatic tissue cell injury, and interference with VEGF-mTORC1 pathway can reduce the autophagy levels and alleviate the inflammatory injury. The present study provides a new target for prevention and treatment of HLAP.

Topics: Acute Disease; Animals; Autophagy; Ceruletide; Male; Mammals; Mechanistic Target of Rapamycin Complex 1; Microtubule-Associated Proteins; Pancreatitis; Rats; Rats, Sprague-Dawley; RNA, Small Interfering; Sirolimus; Vascular Endothelial Growth Factor A

Chronic pancreatitis (CP) is an irreversible fibro-inflammatory disease of the pancreas with no current targeted therapy. Pirfenidone, an anti-fibrotic and anti-inflammatory drug, is FDA approved for treatment of Idiopathic Pulmonary Fibrosis (IPF). Its efficacy in ameliorating CP has never been evaluated before. We recently reported that pirfenidone improves acute pancreatitis in mouse models. The aim of the current study was to evaluate the therapeutic efficacy of pirfenidone in mouse models of CP. We used caerulein and L-arginine models of CP and administered pirfenidone with ongoing injury, or in well-established disease. We evaluated for fibrosis by Sirius-red staining for collagen, immunohistochemistry, western blotting, and qPCR for fibrosis markers to show the salutary effects of pirfenidone in CP. Our results suggest that treatment with pirfenidone ameliorated CP related changes in the pancreas (i.e., atrophy, acinar cell loss, fibrosis, and inflammation) not only when administered with ongoing injury, but also in well-established models of caerulein as well as L-arginine induced CP. It reduces the pro-fibrotic phenotype of macrophages (in-vivo and in-vitro), reduces macrophage infiltration into the pancreas and alters the intra-pancreatic cytokine milieu preceding changes in histology. The therapeutic effect of pirfenidone is abrogated in absence of macrophages. Furthermore, it reduces collagen secretion, cytokine levels and fibrosis markers in pancreatic stellate cells in-vitro. As it is FDA approved, our findings in mouse models simulating clinical presentation of patients to the clinic, can be used as the basis of a clinical trial evaluating the efficacy of this drug as a therapeutic agent for CP.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Collagen; Cytokines; Disease Models, Animal; Fibrosis; Humans; Mice; Pancreatitis, Chronic; Pyridones

Circular RNAs (circRNAs) have been found to be involved in the progression of acute pancreatitis (AP). The objective of our study was to investigate the effects of circ_0000284 on caerulein-induced AR42J cell injury. To mimic AP in vitro, rat pancreatic acinar AR42J cells were treated with caerulein. The expression of circ_0000284 and miR-10a-5p was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Enzyme-linked immunosorbent assay (ELISA) was employed to determine the content of inflammatory cytokines interleukin (IL)-1β, IL-6, IL-8 and tumor necrosis factor α (TNF-α). Western blotting was applied to analyze the levels of Wnt/β-catenin pathway-related and apoptosis-related proteins. Cell viability and apoptosis were monitored by Counting Kit-8 (CCK-8) assay and flow cytometry, respectively. The target connection between circ_0000284 and miR-10a-5p was verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. AP induced inflammation in patients, and caerulein treatment increased apoptosis and inflammation in AR42J cells. Circ_0000284 was upregulated in serum of AP patients and caerulein-induced AR42J cells, while Wnt/β-catenin pathway was inactivated. Knockdown of circ_0000284 could decrease apoptosis and inflammation in caerulein-induced AR42J cells, which was attenuated by miR-10a-5p inhibition or Wnt signaling pathway antagonist Dickkopf-related protein 1 (DKK1). MiR-10a-5p was sponged by circ_000028 and was downregulated in caerulein-induced AR42J cells. Circ_0000284 depletion could protect caerulein-induced AR42J cells from apoptosis and inflammation by upregulating miR-10a-5p expression and activating Wnt/β-catenin pathway, underscoring a potential target for AP therapy.

Topics: Acute Disease; Animals; beta Catenin; Ceruletide; Humans; Inflammation; MicroRNAs; Pancreatitis; Rats; Wnt Signaling Pathway

Necroptosis is a form of regulated necrosis mainly controlled by receptor-interacting protein kinases 3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL). Necroptosis has important roles in defensing against pathogenic infections, but it is also implicated in various inflammatory diseases including pancreatitis. Baicalin, a flavonoid from Scutellaria baicalensis Georgi, has been shown to possess anti-inflammatory and anti-pyroptosis properties, yet it is unclear whether baicalin can inhibit necroptosis and confer protection against necroptosis-related diseases. Here we reported that baicalin significantly inhibited necroptosis in macrophages induced by lipopolysaccharide plus pan-caspase inhibitor (IDN-6556), or by tumor-necrosis factor-α in combination with LCL-161 (Smac mimetic) and IDN-6556 (TSI). Mechanistically, baicalin did not inhibit the phosphorylation of RIPK1, RIPK3 and MLKL, nor membrane translocation of p-MLKL, during necroptotic induction, but instead inhibited p-MLKL oligomerization that is required for executing necroptosis. As intracellular reactive oxygen species (ROS) has been reported to be involved in p-MLKL oligomerization, we assessed the effects of N-acetyl-L-cysteine (NAC), an ROS scavenger, on necroptosis and found that NAC significantly attenuated TSI-induced necroptosis and intracellular ROS production concomitantly with reduced levels of oligomerized p-MLKL, mirroring the effect of baicalin. Indeed, inhibitory effect of baicalin was associated with reduced TSI-induced superoxide (indicating mitochondrial ROS) production and increased mitochondrial membrane potential within cells during necroptosis. Besides, oral administration of baicalin significantly reduced the severity of caerulein-induced acute pancreatitis in mice, an animal model of necroptosis-related disease. Collectively, baicalin can inhibit necroptosis through attenuating p-MLKL oligomerization and confers protection against caerulein-induced pancreatitis in mice.

Topics: Acute Disease; Animals; Apoptosis; Ceruletide; Flavonoids; Mice; Necroptosis; Necrosis; Pancreatitis; Protein Kinases; Reactive Oxygen Species; Receptor-Interacting Protein Serine-Threonine Kinases

Hinokitiol is a natural bio-active tropolone derivative with promising antioxidant and anti-inflammatory properties. This study was conducted to evaluate the ameliorative effects of hinokitiol against acute pancreatitis induced by cerulein. Mice were pre-treated with hinokitiol intraperitoneally for 7 days (50 and 100 mg/kg), and on the final day of study, cerulein (6 × 50 μg/kg) was injected every hour for six times. Six hours after the last dose of cerulein, blood was collected from the mice through retro-orbital plexus for biochemical analysis. After blood collection, mice were euthanized and the pancreas was harvested for studying effects on oxidative stress, pro-inflammatory cytokines, immunohistochemistry and histopathology of tissue sections. Hinokitiol treatment significantly reduced edema of the pancreas and reduced the plasma levels of lipase and amylase in mice with cerulein-induced acute pancreatitis. It also attenuated the oxidative and nitrosative stress related damage as evident from the reduced malondialdehyde (MDA) and nitrite levels, which were significantly increased in the mice with acute pancreatitis. Furthermore, hinokitiol administration significantly reduced the pancreatitis-evoked decrease in the activity of catalase, glutathione (GSH) and superoxide dismutase (SOD) in the pancreatic tissue. Pre-treatment with hinokitiol significantly reduced the elevated levels of pro-inflammatory cytokines like interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor-alpha (TNF-α) as well as increased the levels of anti-inflammatory cytokine interleukin-10 (IL-10) in the pancreatic tissue of mice with acute pancreatitis. The immunohistochemical expression of nuclear factor kappa light chain enhancer of activated B cells (NF-κB), cyclooxygenase (COX-2) and TNF-α were significantly decreased by hinokitiol in mice with cerulein-induced acute pancreatitis. In conclusion, the results of the present study demonstrate that hinokitiol has significant potential to prevent cerulein-induced acute pancreatitis.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Ceruletide; Cytokines; Disease Models, Animal; Mice; Monoterpenes; NF-kappa B; Pancreas; Pancreatitis; Tropolone; Tumor Necrosis Factor-alpha

Acute pancreatitis (AP) is a frequent cause for hospitalization. However, molecular determinants that modulate severity of experimental pancreatitis are only partially understood.. To investigate the role of secreted protein acidic and rich in cysteine (SPARC) during cerulein-induced AP in mice.. AP was induced by repeated cerulein injections in SPARC knock-out mice (SPARC. Upon cerulein induction, SPARC expression was robustly induced in pancreatic stellate cells (PSCs) but not in acinar cells. Genetic SPARC ablation resulted in attenuated severity of AP with significantly reduced levels of pancreatic necrosis, apoptosis, immune cell infiltration, and reduced fibrosis upon chronic stimulation. However, the release of amylase upon cerulein stimulation in primary acinar cell culture from SPARC. SPARC mediates the severity of AP. The potential link between SPARC and the CCL2 axis could open new avenues for tailored therapeutic interventions in AP patients and warrants further investigations.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cysteine; Mice; Osteonectin; Pancreatitis

Cerulein‑induced pancreatitis resembles human acute pancreatitis in terms of pathological events, such as enzymatic activation and inflammatory cell infiltration in the pancreas. Cerulein is a cholecystokinin analog that increases levels of reactive oxygen species (ROS) and interleukin‑6 (IL‑6) expression level in pancreatic acinar cells. Serum levels of resistin, which is secreted from adipocytes, are reportedly higher in patients with acute pancreatitis than in healthy individuals. Previously, it was shown that the adipokine resistin can aggravate the cerulein‑induced increase in ROS levels and IL‑6 expression level in pancreatic acinar cells. Peroxisome proliferator‑activated receptor‑gamma (PPAR‑γ) is a key regulator of the transcription and expression of antioxidant enzymes, including heme oxygenase 1 (HO‑1) and catalase. α‑lipoic acid, a naturally occurring dithiol antioxidant, can prevent cerulein‑induced pancreatic damage in rats. In the present study, it was aimed to investigate whether α‑lipoic acid can attenuate the cerulein/resistin‑induced increase in IL‑6 expression and ROS levels via PPAR‑γ activation in pancreatic acinar AR42J cells. The anti‑inflammatory mechanism of α‑lipoic acid was determined using reverse transcription‑quantitative PCR, western blot analysis, enzyme‑linked immunosorbent assay, immunofluorescence staining and fluorometry. Treatment with cerulein and resistin increased ROS levels and IL‑6 expression level, which were inhibited by α‑lipoic acid in pancreatic acinar cells. α‑lipoic acid increased the nuclear translocation and expression level of PPAR‑γ and the expression levels of its target genes: HO‑1 and catalase. The PPAR‑γ antagonist GW9662 and HO‑1 inhibitor zinc protoporphyrin reversed the inhibitory effect of α‑lipoic acid on cerulein/resistin‑induced increase in ROS and IL‑6 levels. In conclusion, α‑lipoic acid inhibits the cerulein/resistin‑induced increase in ROS production and IL‑6 expression levels by activating PPAR‑γ and inducing the expression of HO‑1 and catalase in pancreatic acinar cells.

Topics: Acinar Cells; Acute Disease; Animals; Antioxidants; Catalase; Ceruletide; Humans; Interleukin-6; Pancreas; Pancreatitis; PPAR gamma; Rats; Reactive Oxygen Species; Resistin; Thioctic Acid

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Humans; Mice; Pancreas; Pancreatitis

Epidemiological studies have demonstrated a strong association between cigarette smoking (CS) and chronic pancreatitis (CP); however, the exact mechanisms of this phenomenon remains unknown. The authors have previously shown that increased Ras expression activates the NF-κB mediated pathway and promotes development of CP. However, it is unclear whether a similar phenomenon occurs in CS-induced CP. Therefore, the aim of the study was to determine whether CS increases the expression of K-Ras, and promotes the development of CP in mice exposed to repeated episodes of acute pancreatitis (AP).. C57BL6/cmdb mice were exposed to CS or a sham treatment for 12 weeks. After one week of exposure, half of the animals from both groups were additionally subjected to repeated cerulein treatment (once a week, for 10 consecutive weeks) to mimic recurrent episodes of AP. Extension of pancreatic damage was determined histologically by H&E and Trichrome staining. The expression of K-Ras protein and downstream components (NF-κB, Cox-2, TGF-β) was evaluated by immunohistochemistry.. C57BL6/cmdb mice exposed to CS or cerulein alone did not develop any chronic pancreatic damage. However, concomitant treatment with both of these agents caused focal acinar atrophy, with slight intralobular and perivascular areas of fibrosis, and inflammatory cells infiltration resembling mild CP. Moreover, immunohistochemistry examinations revealed increased pancreatic expression of K-Ras and NF-κB only in mice treated both with CS and cerulein.. CS promotes development of CP in mice exposed to repeated episodes of AP. This process may be, at least partially, related to increased expression of K-Ras and NF-κB protein.

Topics: Acute Disease; Animals; Ceruletide; Cigarette Smoking; Disease Models, Animal; Mice; Mice, Inbred C57BL; NF-kappa B; Pancreatitis, Chronic; Proto-Oncogene Proteins p21(ras)

Epidemiological studies have established alcohol and smoking as independent risk factors for recurrent acute pancreatitis and chronic pancreatitis. However, the molecular players responsible for the progressive loss of pancreatic parenchyma and fibroinflammatory response are poorly characterized.. Tandem mass tag-based proteomic and bioinformatics analyses were performed on the pancreata of mice exposed to alcohol, cigarette smoke, or a combination of alcohol and cigarette smoke. Biochemical, immunohistochemistry, and transcriptome analyses were performed on the pancreatic tissues and primary acinar cells treated with cerulein in combination with ethanol (50 mmol/L) and cigarette smoke extract (40 μg/mL) for the mechanistic studies.. A unique alteration in the pancreatic proteome was observed in mice exposed chronically to the combination of alcohol and cigarette smoke (56.5%) compared with cigarette smoke (21%) or alcohol (17%) alone. The formation of toxic metabolites (P < .001) and attenuated unfolded protein response (P < .04) were the significantly altered pathways on combined exposure. The extracellular matrix (ECM) proteins showed stable malondialdehyde-acetaldehyde (MAA) adducts in the pancreata of the combination group and chronic pancreatitis patients with a history of smoking and alcohol consumption. Interestingly, MAA-ECM adducts significantly suppressed expression of X-box-binding protein-1, leading to acinar cell death in the presence of alcohol and smoking. The stable MAA-ECM adducts persist even after alcohol and smoking cessation, and significantly delay pancreatic regeneration by abrogating the expression of cyclin-dependent kinases (CDK7 and CDK5) and regeneration markers.. The combined alcohol and smoking generate stable MAA-ECM adducts that increase endoplasmic reticulum stress and acinar cell death due to attenuated unfolded protein response and suppress expression of cell cycle regulators. Targeting aldehyde adducts might provide a novel therapeutic strategy for the management of recurrent acute pancreatitis and chronic pancreatitis.

Topics: Acetaldehyde; Acute Disease; Aldehydes; Animals; Ceruletide; Cyclin-Dependent Kinases; Ethanol; Extracellular Matrix Proteins; Malondialdehyde; Mice; Pancreatitis, Chronic; Proteome; Proteomics; Smoking; Unfolded Protein Response

Severe acute pancreatitis (SAP) is the most serious subtype of acute pancreatitis, manifested as multiple-organ failure resulting in high morbidity and mortality. Based on the role of tripartite motif-containing protein 29 (TRIM29) in immune responses, we aimed to explore its effect on SAP.. Peripheral blood monocyte cells from the SAP or non-SAP patients, as well as bone marrow-derived macrophages from wild-type, TRIM29 -/- , or stimulator of interferon genes (STING) -/- mice after injecting 50 mg/kg of cerulein to induce SAP, were isolated to analyze the role of TRIM29 and STING in the SAP.. Tripartite motif-containing protein 29 was significantly reduced in SAP patients. Compared with wild-type mice, TRIM29 deficiency mice displayed more severe symptom of acute pancreatitis after cerulein injection, which were lost in TRIM29 -/- STING -/- mice. Moreover, interferon and its related genes, as well as STING degradation, were decreased in TRIM29 -/- mice.. Our study demonstrated that TRIM29 negatively regulated the severity of SAP by degrading STING at its downstream, suggesting that TRIM29 and STING might serve as therapeutic targets for SAP.

Topics: Acute Disease; Animals; Ceruletide; Interferons; Macrophages; Mice; Pancreatitis; Transcription Factors

Acute pancreatitis (AP) was initiated within pancreatic parenchymal cells and sustained by uncontrolled inflammatory responses. AXL and MERTK receptor tyrosine kinases play a crucial role in negatively regulating the innate immunity. Therefore, this study aimed to investigate the role and underlying mechanism of AXL and MERTK in AP.. Experimental AP was induced by ten hourly intraperitoneal administration of caerulein in global, hematopoietic- and pancreas-specific Axl and Mertk deficient mice. Pancreatitis severity was assessed biochemically and histologically. Pancreatic transcriptomics and pancreatic infiltrating immune cells were profiled. Some mice were given R428, an antagonist of AXL and MERTK. AXL and MERTK in peripheral leukocytes were measured by flow cytometry.. The levels of AXL and MERTK in pancreatic tissue and pancreatic CD45. Our findings reveal that specific AXL/MERTK antagonist may be a novel and potential early treatment for AP and the levels of MERTK in peripheral leukocytes may be a promising biomarker for predicting pancreatic severity in patients with AP.. National Natural Science Foundation of China, Shanghai Natural Science Foundation, a Shanghai Young Talent Award and a Shanghai Young Orient Scholar Award.. Evidence before this study Acute pancreatitis (AP) is a common inflammatory disorder of the exocrine pancreas, the severity of which was determined by the extent of pancreatic necrosis, with no targeted therapy. AP was initiated by signals within pancreatic parenchymal cells and sustained by uncontrolled innate immune responses. One of the three crucial regulatory roles for AXL and MERTK is to negatively regulate innate immune responses. Added value of this study Global and hematopoietic-, but not pancreas-specific Axl and Mertk deficiency protected against pancreatitis, primarily pancreatic necrosis. Deletion of Axl and Mertk selectively inhibited pancreatic neutrophil infiltration that was related to CXCL2 secreted by pro-inflammatory macrophages. AXL and MERTK antagonist similarly reduced pancreatitis severity via limiting CXCL2-mediated pancreatic neutrophil infiltration. Higher levels of MERTK, but not AXL in peripheral leukocytes were correlated with more severe form of acute pancreatitis. Implications of all the available evidence A specific AXL/MERTK antagonist may be a novel and potential early treatment for AP. The level of MERTK on peripheral leukocytes may be a promising biomarker for predicting disease severity in patients with AP.

Topics: Acute Disease; Animals; c-Mer Tyrosine Kinase; Ceruletide; Chemokine CXCL2; China; Mice; Mice, Inbred C57BL; Neutrophil Infiltration; Pancreatitis, Acute Necrotizing; Trypsinogen; Tyrosine

hyperlipidemia acute pancreatitis (HTG-AP) is a major hidden danger affecting human health, however, whether there is a protective effect of resveratrol on HTG-AP is unclear. Therefore our study was aimed to investigate the preventive effect and the underlying mechanism of resveratrol in the HTG-AP mice model.. This research was divided into two parts. In the first part, mice were adaptively fed with normal chow or HFD for 6 weeks. From the second week, resveratrol-treated mice were in intragastric administration with resveratrol (45 mg/kg/d) for 4 weeks. In the second part, the procedures were the same as the first part. After the last intragastric administration with resveratrol, all mice were intraperitoneal injections of cerulean.. We found resveratrol effectively inhibited pancreatic pathological injury in the HFD, AP, and HTG-AP mice. Resveratrol reduced the LPS, IL-6, TNF-α, and MCP-1 expressions in the HFD mice. Resveratrol also reduced TNF-α, MDA, and MCP-1 expressions and increased SOD and T-AOC expressions in the AP and HTG-AP mice. Furthermore, resveratrol suppressed the NF-κB pro-inflammatory signaling pathway in pancreatic tissues in the AP and HTG-AP mice. Moreover, resveratrol improved the gut microbiota in the HFD mice.. The resveratrol pre-treatment could attenuate pancreas injury, inflammation, and oxidative stress in the HTG-AP mice, via restraining the NF-κB signaling pathway and regulating gut microbiota. Therefore, Our study proved that the resveratrol pre-treatment had a preventive effect on HTG-AP.

Topics: Acute Disease; Animals; Ceruletide; Diet, High-Fat; Gastrointestinal Microbiome; Humans; Mice; NF-kappa B; Pancreatitis; Resveratrol; Signal Transduction; Tumor Necrosis Factor-alpha

Acute pancreatitis (AP) is an inflammatory process of the pancreas resulting from biliary obstruction or alcohol consumption. Approximately, 10-20% of AP can evolve into severe AP (SAP). In this study, we sought to explore the physiological roles of the transcription factor serum response factor (SRF), annexin A2 (ANXA2), and nuclear factor-kappaB (NF-κB) in SAP.. C57BL/6 mice and rat pancreatic acinar cells (AR42J) were used to establish an AP model in vivo and in vitro by cerulein with or without lipopolysaccharide (LPS). Production of pro-inflammatory cytokines (IL-1β and TNF-α) were examined by ELISA and immunoblotting analysis. Hematoxylin and eosin (HE) staining and TUNEL staining were performed to evaluate pathological changes in the course of AP. Apoptosis was examined by flow cytometric and immunoblotting analysis. Molecular interactions were tested by dual luciferase reporter, ChIP, and Co-IP assays.. ANXA2 was overexpressed in AP and correlated to the severity of AP. ANXA2 knockdown rescued pancreatic acinar cells against inflammation and apoptosis induced by cerulein with or without LPS. Mechanistic investigations revealed that SRF bound with the ANXA2 promoter region and repressed its expression. ANXA2 could activate the NF-κB signaling pathway by inducing the nuclear translocation of p50. SRF-mediated transcriptional repression of ANXA2-protected pancreatic acinar cells against AP-like injury through repressing the NF-κB signaling pathway.. Our study highlighted a regulatory network consisting of SRF, ANXA2, and NF-κB that was involved in AP progression, possibly providing some novel targets for treating SAP.

Topics: Acute Disease; Animals; Annexin A2; Ceruletide; Lipopolysaccharides; Mice; Mice, Inbred C57BL; NF-kappa B; Pancreas; Pancreatitis; Rats; Serum Response Factor; Signal Transduction

Acute pancreatitis (AP) is a common cause of a clinically acute abdomen. Crosstalk between acinar cells and leukocytes (especially macrophages) plays an important role in the development of AP. However, the mechanism mediating the interaction between acinar cells and macrophages is still unclear. This study was performed to explore the role of acinar cell extracellular vesicles (EVs) in the crosstalk between acinar cells and macrophages involved in the pathogenesis of AP. EVs derived from caerulein-treated acinar cells induced macrophage infiltration and aggravated pancreatitis in an AP rat model. Further research showed that acinar cell-derived EV miR-183-5p led to M1 macrophage polarization by downregulating forkhead box protein O1 (FoxO1), and a dual-luciferase reporter assay confirmed that FoxO1 was directly inhibited by miR-183-5p. In addition, acinar cell-derived EV miR-183-5p reduced macrophage phagocytosis. Acinar cell-derived EV miR-183-5p promoted the pancreatic infiltration of M1 macrophages and increased local and systemic damage

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Down-Regulation; Extracellular Vesicles; Macrophages; MicroRNAs; Nerve Tissue Proteins; Pancreatitis; Rats

Severe acute pancreatitis can easily lead to systemic inflammatory response syndrome and death. Macrophages are known to be involved in the pathophysiology of acute pancreatitis (AP), and macrophage activation correlates with disease severity. In this study, we examined the role of ubiquitin-specific protease 25, a deubiquitinating enzyme and known regulator of macrophages, in the pathogenesis of AP.. We used L-arginine, cerulein, and choline-deficient ethionine-supplemented diet-induced models of AP in Usp25. Our results show that Usp25 deficiency exacerbates pancreatic and lung injury, neutrophil and macrophage infiltration, and systemic inflammatory responses in L-arginine, cerulein, and choline-deficient ethionine-supplemented diet-induced models of AP. Bone marrow Usp25. Usp25 deficiency in macrophages enhances TBK1-NF-κB signaling, and the induction of inflammatory chemokines and type I interferon-related genes exacerbates pancreatic and lung injury in AP.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Choline; Cytokines; Deubiquitinating Enzymes; Disease Models, Animal; Ethionine; Interferon Type I; Lung Injury; Macrophages; Mice; Mice, Inbred C57BL; NF-kappa B; Pancreatitis; Signal Transduction; Ubiquitin Thiolesterase; Ubiquitin-Specific Proteases

Acute pancreatitis is a severe inflammatory disease of the pancreas. In experimental acute pancreatitis, cerulein induces the expression of interleukin‑6 (IL‑6) by activating Janus kinase (JAK) 2/signal transducer and activator of transcription (STAT) 3 in pancreatic acinar cells. Ligands of peroxisome proliferator activated receptor‑γ (PPAR‑γ) and suppressor of cytokine signaling (SOCS) 3 inhibit IL‑6 expression by suppressing JAK2/STAT3 in cerulein‑stimulated pancreatic acinar AR42J cells. Lutein, an oxygenated carotenoid, upregulates and activates PPAR‑γ to regulate inflammation in a renal injury model. The present study aimed to determine whether lutein activated PPAR‑γ and induced SOCS3 expression in unstimulated AR42J cells, and whether lutein inhibited activation of JAK2/STAT3 and IL‑6 expression via activation of PPAR‑γ and SOCS3 expression in cerulein‑stimulated AR42J cells. The anti‑inflammatory mechanism of lutein was determined using reverse transcription‑quantitative PCR, western blot analysis and enzyme‑linked immunosorbent assay in AR42J cells stimulated with or without cerulein. In another experiment, cells were treated with lutein and the PPAR‑γ antagonist GW9662 or the PPAR‑γ agonist troglitazone prior to cerulein stimulation to determine the involvement of PPAR‑γ activation. The results indicated that lutein increased PPAR‑γ and SOCS3 levels in unstimulated cells. Cerulein increased phospho‑specific forms of JAK2 and STAT3, and mRNA and protein expression of IL‑6, but decreased SOCS3 levels in AR42J cells. Cerulein‑induced alterations were suppressed by lutein or troglitazone. GW9662 alleviated the inhibitory effect of lutein on JAK2/STAT3 activation and IL‑6 expression in cerulein‑stimulated cells. In conclusion, lutein inhibited the activation of JAK2/STAT3 and reduced IL‑6 levels via PPAR‑γ‑mediated SOCS3 expression in pancreatic acinar cells stimulated with cerulein.

Topics: Acinar Cells; Acute Disease; Ceruletide; Humans; Interleukin-6; Lutein; Pancreatitis; PPAR gamma; STAT3 Transcription Factor; Suppressor of Cytokine Signaling 3 Protein; Suppressor of Cytokine Signaling Proteins; Troglitazone

Polo-like kinase 1 (Plk1) plays an important role in cell-cycle regulation. Recent work has suggested that Plk1 could be a biomarker of gemcitabine response in pancreatic ductal adenocarcinoma (PDAC). Although targeting Plk1 to treat PDAC has been attempted in clinical trials, the results were not promising, and the mechanisms of resistance to Plk1 inhibition is poorly understood. In addition, the role of Plk1 in PDAC progression requires further elucidation. Here, we showed that Plk1 was associated with poor outcomes in patients with PDAC. In an inducible transgenic mouse line with specific expression of Plk1 in the pancreas, Plk1 overexpression significantly inhibited caerulein-induced acute pancreatitis and delayed development of acinar-to-ductal metaplasia and pancreatic intraepithelial neoplasia. Bioinformatics analyses identified the regulatory networks in which Plk1 is involved in PDAC disease progression, including multiple inflammation-related pathways. Unexpectedly, inhibition or depletion of Plk1 resulted in upregulation of PD-L1 via activation of the NF-κB pathway. Mechanistically, Plk1-mediated phosphorylation of RB at S758 inhibited the translocation of NF-κB to nucleus, inactivating the pathway. Inhibition of Plk1 sensitized PDAC to immune checkpoint blockade therapy through activation of an antitumor immune response. Together, Plk1 suppresses PDAC progression and inhibits NF-κB activity, and targeting Plk1 can potentiate the efficacy of immunotherapy in PDAC.. Inhibition of Plk1 induces upregulation of PD-L1 expression in pancreatic ductal adenocarcinoma, stimulating antitumor immunity and sensitizing tumors to immunotherapy.

Topics: Acute Disease; Animals; B7-H1 Antigen; Carcinoma, Pancreatic Ductal; Cell Cycle Proteins; Ceruletide; Humans; Immune Checkpoint Inhibitors; Mice; NF-kappa B; Pancreatic Neoplasms; Pancreatitis; Polo-Like Kinase 1; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins

Premature intracellular trypsinogen activation has long been considered a key initiator of acute pancreatitis (AP). Cathepsin B (CTSB) activates trypsinogen, while cathepsin L (CTSL) inactivates trypsin(ogen), and both proteins play a role in the onset of AP.. AP was induced by 7 hourly intraperitoneal injections of cerulein (50 μg/kg) in wild-type and pancreas-specific conditional Ctsb knockout (Ctsb. Double deletion of Ctsb and Cstl did not affect pancreatic development or mouse growth. After 7 times cerulein injections, double Ctsb and Ctsl deficiency in mouse pancreases increased trypsin activity to the same extent as that in Ctsl-deficient mice, while Ctsb deficiency decreased trypsin activity but did not affect the severity of AP. Ctsb. Double deletion of Ctsb and Ctsl in the mouse pancreas altered intrapancreatic trypsin activity but did not affect disease severity and inflammatory response after cerulein-induced AP.

Topics: Acute Disease; Amylases; Animals; Cathepsin B; Ceruletide; Mice; Mice, Knockout; Pancreas; Pancreatitis; Trypsin; Trypsinogen

In this research, we screened out two genes upregulated in mice with acute pancreatitis (AP) by gene sequencing: microRNA (miR)-320-3p and matrix metalloprotease 8 (MMP8). This study was designed to determine whether miR-320-3p and MMP8 participate in AP development and explore the mechanisms, with a new idea for clinical diagnosis and treatment of AP. Expression of miR-320-3p, DNA methyltransferase 3a (DNMT3a), and MMP8 in mouse pancreatic tissues and AR42J cells was tested by RT-qPCR and western blot assays. Pancreatic pathological changes, serum amylase and lipase, and inflammatory factors in mouse serum and cell supernatant were measured by hematoxylin-eosin staining, automation analyzer, and enzyme-linked immunosorbent assay, respectively. Cell proliferation and apoptosis were determined by CCK-8 assay and flow cytometry. The interaction between miR-320-3p, DNMT3a, and MMP8 was verified by luciferase activity assay, ChIP-qPCR, and MSP assay. High expression of miR-320-3p and MMP8, and low expression of DNMT3a were observed in pancreatic tissues of AP mice and caerulein-induced AP cellular model. Downregulation of miR-320-3p alleviated injury of mouse pancreas, reduced the levels of serum amylase and lipase, and blocked inflammatory factor levels in AP mice. In caerulein-induced AP cellular models, inhibiting miR-320-3p facilitated proliferation and inhibited apoptosis. Upregulation of MMP8 resulted in the opposite results, which could be reversed by simultaneous inhibition of miR-320-3p. miR-320-3p targeted DNMT3a, and downregulating miR-320-3p promoted DNMT3a expression. Moreover, DNMT3a promoted DNA methylation in MMP8 promoter region, thereby inhibiting MMP8 expression in AP mouse and cellular models. This research suggests that miR-320-3p inhibits DNMT3a to reduce MMP8 methylation and increase MMP8 expression, thereby promoting AP progression.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Cell Proliferation; Ceruletide; DNA Methylation; DNA Methyltransferase 3A; Eosine Yellowish-(YS); Hematoxylin; Lipase; Luciferases; Matrix Metalloproteinase 8; Mice; MicroRNAs; Pancreatitis

Topics: Acute Disease; ADAM17 Protein; Animals; Carcinogens; Ceruletide; Cytokines; Disintegrins; Endopeptidases; Fibrosis; Interleukin-6; Ketones; Mice; Nicotine; Nitrosamines; Pancreatitis; Peptide Hydrolases; Tumor Necrosis Factor-alpha

The aim of the study was to evaluate whether a new and successful treatment opportunity can be provided in acute pancreatitis and may prevent symptomatic treatments and show its effect through etiopathogenesis. Therefore, we want to investigate the efficacy of golimumab in an experimental rat model of cerulein-induced acute pancreatitis.. A total of 35 rats, including 7 rats in each group, were distributed into 5 groups (sham, acute pancreatitis, placebo, acute pancreatitis+golimumab 5 mg/kg, and acute pancreatitis+golimumab 10 mg/kg). An experimental cerulein-induced acute pancreatitis model was accomplished by intraperitoneal cerulein injections. After sacrification, rat blood samples were collected for amylase, IL-6, and IL-1beta measurements. Histopathological analysis of the pancreas was performed with Tunel and hematoxylin and eosin staining.. Amylase, IL-6, and IL-1beta levels were found to be increased in the acute pancreatitis group. IL-1beta, amylase, IL-6 levels, and pancreatic inflammation were all significantly decreased in golimumab groups (P < .01). Moreover, in both golimumab groups, golimumab treatment significantly reduced apoptosis in pancreatic tissues (P < .05). Golimumab treatment was found to significantly reduce edema formation, inflammation, vacuolization, and fat necrosis of pancreatic tissues (P < .05).. Firstly in the literature, we investigated the efficacy of golimumab in the experimental acute pancreatitis model. In the light of our findings, it could be suggested that golimumab may be an effective and safe therapeutic option in the treatment of patients with acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Inflammation; Interleukin-6; Pancreas; Pancreatitis; Rats

Acute pancreatitis (AP) is mainly caused by acinar cells releasing various inflammatory factors, causing inflammatory storms and leading to severe pancreatitis. Detection methods and treatment targets for pancreatitis are lacking, raising the urgency of identifying diagnostic markers and therapeutic targets for AP. MicroRNAs (miRNAs) have recently been identified as molecular markers for various biological processes such as tumors, immunity, and metabolism, and the involvement of miRNAs in inflammatory responses has been increasingly studied. To explore the role of miRNAs in AP is the primary objective of this study. By using qPCR on our cerulein-induced pancreatitis cell model, it is worth noting that the change of miR-146a-5p expression in inflammation-related miRNAs in AP was predominant. Next, ELISA, CCK8, and flow cytometry were used to inspect the impact of miR-146a-5p on pancreatitis. BiBiServ bioinformatics anticipated binding ability of miR-146a-5p and 3'-untranslated region (3'UTR) of TNF receptor-associated factor 6 (TRAF6), and the dual-luciferase assay verified the combination of the two. TRAF6 knockdown verified the effect of TRAF6 on the progression of pancreatitis. Finally, rescue experiments verified the capability of miR-146a-5p and TRAF6 interaction on the Toll-like receptor 9 (TLR9)/NOD-like receptor protein 3 (NLRP3) signaling pathway and cell function. The expression of miR-146a-5p decreased in cerulein-induced AR42J pancreatic acinar cells. Functional experiments verified that miR-146a-5p facilitated the proliferation of AR42J pancreatic acinar cells and inhibited their apoptosis. Bioinformatic predictions and dual-luciferase experiments verified the actual binding efficiency between miR-146a-5p and 3'UTR of TRAF6. Our study confirmed that knockdown of TRAF6 restrained the progression of pancreatitis, and knockdown of TRAF6 rescued pancreatitis caused by miR-146a-5p downregulation by the TLR9/NLRP3 signaling pathway. Therefore, downregulation of miR-146a-5p in the induced pancreatitis cell model promotes the progression of pancreatitis via the TLR9/TRAF6/NLRP3 signaling pathway. There is potential for miR-146a-5p to serve as a diagnostic marker and therapeutic nucleic acid drug for AP.

Topics: 3' Untranslated Regions; Acute Disease; Animals; Ceruletide; Down-Regulation; MicroRNAs; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreatitis; Rats; Signal Transduction; TNF Receptor-Associated Factor 6; Toll-Like Receptor 9

Acute pancreatitis (AP) is a major, globally increasing gastrointestinal disease and a biliary origin is the most common cause. However, the effects of bile acids (BAs), given systemically, on the pancreas and on disease severity remains elusive. In this study, we have investigated the roles of different circulating BAs in animal models for AP to elucidate their impact on disease severity and the underlying pathomechanisms. BAs were incubated on isolated acini and AP was induced through repetitive injections of caerulein or L-arginine; pancreatic duct ligation (PDL); or combined biliopancreatic duct ligation (BPDL). Disease severity was assessed using biochemical and histological parameters. Serum cholecystokinin (CCK) concentrations were determined via enzyme immunoassay. The binding of the CCK1 receptor was measured using fluorescence-labeled CCK. In isolated acini, hydrophobic BAs mitigated the damaging effects of CCK. The same BAs further enhanced pancreatitis in L-arginine- and PDL-based pancreatitis, whereas they ameliorated pancreatic damage in the caerulein and BPDL models. Mechanistically, the binding affinity of the CCK1 receptor was significantly reduced by hydrophobic BAs. The hydrophobicity of BAs and the involvement of CCK seem to be relevant in the course of AP. Systemic BAs may affect the severity of AP by interfering with the CCK1 receptor.

Topics: Acute Disease; Animals; Arginine; Bile Acids and Salts; Ceruletide; Cholecystokinin; Disease Models, Animal; Hydrophobic and Hydrophilic Interactions; Mice; Pancreas; Pancreatitis

Hair follicle-derived mesenchymal stem cell (HF-MSC)-based therapies protect against acute pancreatitis (AP). However, accumulating evidence suggests that MSC-based therapy mainly involves the secretion of MSC-derived small extracellular vesicles (MSC-sEVs) through paracrine effects. Thus, the present research investigated the therapeutic effect of HF-MSC-sEVs in AP and the underlying mechanisms.. SEVs were purified from cultured HF-MSC supernatant. The effects of sEVs in vitro were analyzed on caerulein-simulated pancreatic acinar cells (PACs). The therapeutic potential of sEVs in vivo was examined in a caerulein-induced AP model. The organ distribution of sEVs in mice was determined by near-infrared fluorescence (NIRF) imaging. Serum specimens and pancreatic tissues were collected to analyze the inhibition of inflammation and pyroptosis in vivo, as well as the appropriate infusion route: intraperitoneal (i.p.) or intravenous (i.v.) injection.. HF-MSC-sEVs were taken up by PACs and improved cell viability in vitro. In vivo, sEVs were abundant in the pancreas, and the indicators of pancreatitis, including amylase, lipase, the inflammatory response, myeloperoxidase (MPO) expression and histopathology scores, in sEV-treated mice were markedly improved compared with those in the AP group, especially via tail vein injection. Furthermore, we revealed that sEVs observably downregulated the levels of crucial pyroptosis proteins in both PACs and AP tissue.. We innovatively demonstrated that HF-MSC-sEVs could alleviate inflammation and pyroptosis in PACs in AP, suggesting a refreshing cell-free remedy for AP.

Topics: Acute Disease; Animals; Ceruletide; Extracellular Vesicles; Hair Follicle; Inflammation; Mice; Pancreatitis

Acute pancreatitis (AP) is an inflammatory disease with very poor outcomes. However, the order of induction and coordinated interactions of systemic inflammatory response syndrome (SIRS) and compensatory anti-inflammatory response syndrome (CARS) and the potential mechanisms in AP are still unclear.. An integrative analysis was performed based on transcripts of blood from patients with different severity levels of AP (GSE194331), as well as impaired lung (GSE151572), liver (GSE151927) and pancreas (GSE65146) samples from an AP experimental model to identify inflammatory signals and immune response-associated susceptibility genes. An AP animal model was established in wild-type (WT) mice and Tlr2-deficient mice by repeated intraperitoneal injection of cerulein. Serum lipase and amylase, pancreas impairment and neutrophil infiltration were evaluated to assess the effects of. In summary, we discovered SIRS and CARS were stimulated in parallel, not activated consecutively. In addition, among the novel susceptibility genes,

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Ceruletide; Disease Models, Animal; Disease Progression; Lipase; Mice; Mice, Inbred C57BL; Pancreatitis; Systemic Inflammatory Response Syndrome; Toll-Like Receptor 2

Acute pancreatitis (AP) is an inflammatory disease of the pancreas. A growing number of studies have shown that long noncoding RNAs (lncRNAs) play an important role in AP progression. Here, we aimed to elucidate the role of Small Nucleolar RNA Host Gene 11(SNHG11) and its underlying molecular mechanisms behind AP progression. The in vivo and in vitro AP cell models were established by retrograde injection of sodium taurocholate and caerulein stimulation into AR42J cells and HPDE6-C7 cells, respectively. A bioinformatics website predicted the relationship between SNHG11, miR-7-5p, and Phospholipase C Beta 1(PLCB1) and validated it with a dual-luciferase reporter assay and an RNA immunoprecipitation (RIP) assay. AR42J cells and HPDE6-C7 cells were transfected with an overexpression of plasmids or shRNA to investigate the effects of the SNHG11/miR-7-5p/PLCB1 axis on cell proliferation and apoptosis, inflammatory cytokine secretion, and acute pancreatitis. Low expression of SNHG11 and PLCB1 and high expression of miR-7-5p were observed in AP pancreatic tissue and AP cell models. SNHG11 overexpression inhibited apoptosis and inflammatory responses induced by caerulein. Simultaneously, we discovered that SNHG11 regulates PLCB1 expression by sponging miR-7-5p. PLCB1 overexpression abrogated inflammatory damage exacerbated by miR-7-5p enrichment. In addition, the SNHG11/miR-7-5p/PLCB1 axis could be involved in caerulein-induced inflammatory injury by participating in the p38MAPK signaling pathway. The overexpressed SNHG11/miR-7-5p/PLCB1 axis can inhibit AP progression by participating in the p38MAPK signaling pathway, thereby providing a potential therapeutic target and therapeutic direction for AP therapy.

Topics: Acute Disease; Ceruletide; Humans; MicroRNAs; p38 Mitogen-Activated Protein Kinases; Pancreatitis; Phospholipase C beta; RNA, Long Noncoding

Acute pancreatitis is a clinical picture with a wide range of symptoms from mild inflammation to multiorgan failure and death. The aim of this study is to investigate the effects of Adalimumab (ADA) on inflammation and apoptosis in a cerulein-induced acute pancreatitis model in rats.. Experimental cerulein-induced acute pancreatitis model was created by applying 4 intraperitoneal cerulein injections at 1-h intervals. A total of 40 rats, 8 in each group, were randomly distributed into five groups. In the groups that ADA treatment was given, two different doses of ADA were administered 5 mg/kg and 20 mg/kg as low and high doses, respectively. The rats were sacrificed 12 h after the last intraperitoneal administration of ADA. Blood samples were obtained from each rat for amylase, IL-6, and IL-1β measurements. Hematoxylin and Eosin (H&E) stains were used to undertake the histopathological analysis of the pancreas. The terminal deoxynucleotidyl transferase-mediated nick-end-labeling (TUNEL) method was used to evaluate apoptosis.. : Plasma amylase, IL-6, and IL-1β levels were significantly elevated in acute pancreatitis groups (p < 0.05). It was determined that both low (5 mg/kg) and high doses (20 mg/kg) of ADA ameliorated the parameters (plasma amylase, IL-6, and IL-1β) (p < 0.05). Although significant improvements were detected in the Schoenberg scoring system and the apoptotic index from the TUNEL method after highdose ADA treatment, no significant amelioration was observed in the histopathological examinations in the low-dose ADA group.. : It has been determined that the administration of high-dose ADA effectively alleviated the symptoms of acute pancreatitis and reduced the level of apoptosis. In line with the findings of our study, we have predicted that high-dose (20 mg/kg) ADA can be used as an effective and safe drug in the treatment of patients with acute pancreatitis.

Topics: Acute Disease; Adalimumab; Amylases; Animals; Ceruletide; Disease Models, Animal; Inflammation; Interleukin-6; Pancreatitis; Rats; Rats, Wistar

Obesity is an important risk factor for severe acute pancreatitis. The necrosis of epididymal adipose tissue occurs in severe acute pancreatitis. Adipose tissue macrophages play an important role in metabolic related inflammation. Therefore, we explored the potential mechanisms between adipose tissue macrophages and obesity-related severe acute pancreatitis.. Severe acute pancreatitis mice model was induced by caerulein with lipopolysaccharide. The severity of severe acute pancreatitis was evaluated according to the morphological, general, and biochemical change. We assessed the injury of epididymal white adipose tissue, pancreas, and adipose tissue macrophages in obese mice and lean mice with severe acute pancreatitis. Outcomes of caerulein-induced severe acute pancreatitis were studied in lean and obese mice with or without lipase inhibitor orlistat.. Fat necrosis and pancreatic injury increased in the SAP groups. High levels of serum free fatty acid and triglyceride were increased significantly in the SAP group. The NLRP3-caspase1 inflammasome signal pathway in adipose tissue macrophages markedly enhanced in the SAP groups compared with control group. Free fatty acid can trigger macrophages inflammation through NLRP3-caspase1. Lipase inhibited by orlistat remarkably decreased in adipose tissue necrosis, and the levels of serum lipase, amylase, and pancreatic tissue damage decreased in the orlistat group compared with the SAP group. The NLRP3-caspase1 inflammasome pathway in adipose tissue macrophages markedly decreased in the orlistat groups compared with SAP group. The levels of serum free fatty acid and triglyceride were decreased significantly in the orlistat group.. Inflammation increases in adipose tissue macrophages of obese mice with severe acute pancreatitis. Free fatty acid generated via adipocyte lipolysis worsens inflammation in adipose tissue macrophages and the outcome of severe acute pancreatitis in obese mice through the NLRP3-caspase1 inflammasome pathway.

Topics: Acute Disease; Adipose Tissue; Animals; Caspase 1; Ceruletide; Fatty Acids, Nonesterified; Inflammasomes; Inflammation; Lipase; Macrophages; Mice; Necrosis; NLR Family, Pyrin Domain-Containing 3 Protein; Obesity; Orlistat; Pancreatitis; Triglycerides

Acute pancreatitis (AP) is an inflammatory gastrointestinal disorder affecting the pancreas. Previous study reported that tetraspanin 1 (TSPAN1) expression was significantly upregulated in the pancreas of AP patients. However, the underlying molecular mechanism of TSPAN1 in the pathogenesis of AP remains unclear. Thus, the aim of the present study was to investigate the potential role of TSPAN1 in development of AP. RT-qPCR was carried out to quantify the relative mRNA levels of TSPAN1 and anterior gradient-2 (AGR2). The CCK-8 assay was used to detect the cell viability. The TUNEL assay was performed to visualize the apoptotic cells. Western blot was performed to determine the expressions of proteins related to endoplasmic reticulum (ER) stress and apoptosis. ELISA kits were adopted to detect the concentration of inflammatory cytokines including TNF-α and IL-6. Finally, immunoprecipitation (IP) was used to verify the interaction between TSPAN1 and AGR2. TSPAN1 was upregulated in serum of AP patients and AP cell models. TSPAN1 silencing promoted the cell proliferation and inhibited inflammatory response in cerulein-induced AR42J cells. Moreover, TSPAN1 induced endoplasmic reticulum stress by binding AGR2. Interestingly, the overexpression of AGR2 abolished the effects of TSPAN1 silencing on cell proliferation and inflammatory response in cerulein-induced AR42J cells. In summary, TSPAN1 silencing protects against cerulein-induced pancreatic acinar cell injury through inhibiting ER stress-mediated by AGR2. Hence, TSPAN1 may serve as a promising therapeutic target for AP treatment.

Topics: Acinar Cells; Acute Disease; Ceruletide; Humans; Mucoproteins; Oncogene Proteins; Pancreas; Pancreatitis; Tetraspanins

Acute pancreatitis (AP), an inflammatory disorder of the pancreas, is a complicated disease without specific drug therapy. (R)-4,6-dimethoxy-3-(4-methoxy phenyl)-2,3-dihydro-1H-indanone [(R)-TML104] is a synthesized analog of the natural product resveratrol sesquiterpenes (±) -isopaucifloral F. This study aimed to investigate the effect and underlying mechanism of (R)-TML104 on AP. The experimental AP model was induced by caerulein hyperstimulation in BALB/c mice. (R)-TML104 markedly attenuated caerulein-induced AP, as evidenced by decreased pancreatic edema, serum amylase levels, serum lipase levels, and pancreatic myeloperoxidase activity. In addition, (R)-TML104 significantly inhibited the expression of pancreatic chemokines C-C motif chemokine ligand 2 and macrophage inflammatory protein-2 and the infiltration of neutrophils and macrophages. Mechanistically, (R)-TML104 activated AMP-activated protein kinase and induced sirtuin 1 (SIRT1) expression. (R)-TML104 treatment markedly induced the SIRT1-signal transducer and activator of transcription 3 (STAT3) interaction and reduced acetylation of STAT3, thus inhibiting the inflammatory response mediated by the interleukin 6-STAT3 pathway. The effect of (R)-TML104 on SIRT1-STAT3 interaction was reversed by treatment with a SIRT1 inhibitor selisistat (EX527). Together, our findings indicate that (R)-TML104 alleviates experimental pancreatitis by reducing the infiltration of inflammatory cells through modulating SIRT1.

Topics: Acute Disease; Animals; Ceruletide; Mice; Pancreas; Pancreatitis; Resveratrol; Sirtuin 1

Topics: Acute Disease; Animals; Caspase 3; Caspases; Ceruletide; Humans; Interleukin-18; Interleukin-1beta; Intracellular Signaling Peptides and Proteins; Male; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreas; Pancreatitis; Pyroptosis; Rats; TNF Receptor-Associated Factor 6

Acute pancreatitis (AP) is an acute inflammatory condition of the pancreas. Previous studies have shown that rutaecarpine (RUT), an important alkaloid component of Evodia rutaecarpa, exhibits certain protective effects against AP in rats by upregulating calcitonin gene-related peptide (CGRP). However, the molecular mechanism of RUT in AP remains unknown. This study aimed to investigate the effects of RUT on cerulein-induced AP in vivo and in vitro, and to explore the underlying molecular mechanisms. In cerulein/LPS-treated wild-type mice, but not CGRP gene knock-out mice, RUT significantly ameliorated pancreatic inflammation by alleviating histopathological changes, reducing IL-6 and TNF-α levels, and increasing in IL-10 levels. Moreover, RUT improved AP by suppressing the MAPK and NF-κB signaling pathways. These effects were mostly mediated through CGRP. Cell-based studies revealed that RUT significantly improved cell viability while suppressing the apoptosis of AR42J cells with cerulein-induced AP, downregulating IL-6 and TNF-α, stimulating IL-10 release, and inhibiting MAPK, NF-κB, and STAT3 signaling activation, all in a CGRP-dependent manner. RUT ameliorated cerulein/LPS-induced AP inflammatory responses in mice and AR42J cells in a CGRP-dependent manner and thus may represent a potential therapeutic option for AP patients. Our study provides valuable insights for AP drug development.

Topics: Acute Disease; Animals; Calcitonin; Calcitonin Gene-Related Peptide; Ceruletide; Humans; Indole Alkaloids; Mice; NF-kappa B; Pancreatitis; Quinazolines; Rats

Previous studies have provided conflicting results regarding whether the serum ghrelin concentration can reflect the severity of acute pancreatitis (AP). The present study examined the correlation between the serum ghrelin concentration and AP severity in animal models and investigated whether altered ghrelin expression in pancreatic acinar cells influences IKKβ/NF-κB signaling and pro-inflammatory cytokine production.. Mild or severe AP was induced in rats by intraperitoneal injection of cerulein or retrograde cholangiopancreatic duct injection of sodium taurocholate, respectively. After successful model induction, serum ghrelin, tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) concentrations were determined by enzyme-linked immunosorbent assay, and IKKβ/NF-κB activation was assessed by immunohistochemistry. Subsequently, stable overexpression or knockdown of ghrelin in AR42J cells was achieved by lentiviral transfection. After transfected cells and control cells were treated with cerulein for 24 h, the TNF-α and IL-1β levels in the supernatants were determined by enzyme-linked immunosorbent assay, and the expression levels of p-p65, IKKβ, and p-IKKβ were detected by Western blotting.. In rat AP models, AP severity was correlated with increased IKKβ/NF-κB activation, pro-inflammatory cytokine production, and ghrelin secretion. The levels of pro-inflammatory cytokines TNF-α and IL-1β as well as IKKβ/NF-κB signaling activity were increased upon knockdown of ghrelin in the AP acinar cell model and decreased with ghrelin overexpression.. Serum ghrelin is related to the severity of AP. Ghrelin may play a protective role in the pathogenesis of AP by inhibiting the pro-inflammatory cytokines and the activation of the IKKβ/NF-κB signaling pathway.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Cytokines; Ghrelin; I-kappa B Kinase; NF-kappa B; Pancreas; Pancreatitis; Rats; Signal Transduction; Tumor Necrosis Factor-alpha

Serum amyloid A (SAA) is an early and sensitive biomarker of inflammatory diseases, but its role in acute pancreatitis (AP) is still unclear. Here, we used a caerulein-induced mouse model to investigate the role of SAA in AP and other related inflammatory responses. In our study, we found that the expression of a specific SAA isoform, SAA3, was significantly elevated in a caerulein-induced AP animal model. In addition, SAA3-knockout (Saa3

Topics: Acute Disease; Animals; Ceruletide; Mice; Necroptosis; Pancreatitis; Serum Amyloid A Protein

Acute pancreatitis (AP) and recurring AP are serious health care problems causing excruciating pain and potentially lethal outcomes due to sepsis. The validated caerulein- (CAE) induced mouse model of acute/recurring AP produces secondary persistent hypersensitivity and anxiety-like behavioral changes for study.. To determine efficacy of acetyl-L-carnitine (ALC) to reduce pain-related behaviors and brain microglial activation along the pain circuitry in CAE-pancreatitis.. Pancreatitis was induced with 6 hly intraperitoneal (i.p.) injections of CAE (50 µg/kg), 3 d a week for 6 wk in male C57BL/6J mice. Starting in week 4, mice received either vehicle or ALC until experiment's end. Mechanical hyper-sensitivity was assessed with von Frey filaments. Heat hypersensitivity was determined with the hotplate test. Anxiety-like behavior was tested in week 6 using elevated plus maze and open field tests. Microglial activation in brain was quantified histologically by immunostaining for ionized calcium-binding adaptor molecule 1 (Iba1).. Mice with CAE-induced pancreatitis had significantly reduced mechanical withdrawal thresholds and heat response latencies, indicating ongoing pain. Treatment with ALC attenuated inflammation-induced hypersensitivity, but hypersensitivity due to abdominal wall injury caused by repeated intraperitoneal injections persisted. Animals with pancreatitis displayed spontaneous anxiety-like behavior in the elevated plus maze compared to controls. Treatment with ALC resulted in increased numbers of rearing activity events, but time spent in "safety" was not changed. After all the abdominal injections, pancreata were translucent if excised at experiment's end and opaque if excised on the subsequent day, indicative of spontaneous healing. Post mortem histopathological analysis performed on pancreas sections stained with Sirius Red and Fast Green identified wide-spread fibrosis and acinar cell atrophy in sections from mice with CAE-induced pancreatitis that was not rescued by treatment with ALC. Microglial Iba1 immunostaining was significantly increased in hippocampus, thalamus (intralaminar nuclei), hypothalamus, and amygdala of mice with CAE-induced pancreatitis compared to naïve controls but unchanged in the primary somatosensory cortex compared to naïves.. CAE-induced pancreatitis caused increased pain-related behaviors, pancreatic fibrosis, and brain microglial changes. ALC alleviated CAE-induced mechanical and heat hypersensitivity but not abdominal wall injury-induced hypersensitivity caused by the repeated injections.

Topics: Acetylcarnitine; Acute Disease; Animals; Brain; Ceruletide; Disease Models, Animal; Male; Mice; Mice, Inbred C57BL; Microglia; Pain; Pancreas; Pancreatitis

Acute pancreatitis (AP) is an inflammatory disorder of the pancreas that is associated with substantial morbidity and mortality. Chaiqin chengqi decoction (CQCQD) has been proven clinically to be an effective treatment for AP for decades in West China Hospital. Quality control for CQCQD containing many hundreds of characteristic phytochemicals poses a challenge for developing robust quality assessment metrics.. To evaluate quality consistency of CQCQD with a multi-strategy based analytical method, identify potential quality-markers (Q-markers) based on drug properties and effect characteristics, and endeavor to establish CQCQD as a globally-accepted medicine.. A typical analysis of constitutive medicinal plant materials was performed following the Chinese Pharmacopoeia. The extraction process was optimized through an orthogonal array (L. The chemical markers and other physical and chemical indices in the original materials met Chinese Pharmacopoeia standards. A total of 22 co-existing fingerprint peaks were selected and the similarity varied between 0.946 and 0.990. Batch D10 possessed the highest similarity index. HCA classified the 10 batches into 2 main groups: 7 batches represented by D10 and 3 batches represented by D1. During the initial Q-marker screen stage, 22 compounds were detected in both plant materials and decoctions, while 13 compounds were identified in plasma. Network pharmacology predicted the potential targets and pathway of AP related to the 22 compounds. All 10 batches showed reduced necrosis below 60% with the best effect achieved by D10 (~40%). The spectrum-efficacy relationship analyzed by Pearson correlation analysis indicated that emodin, rhein, aloe emodin, geniposide, hesperridin, chrysin, syringin, synephrine, geniposidic acid, magnolol, physcion, sinensetin, and baicalein showed positive correlation with pancreatic acinar cell death protection. Similar to the in vitro evaluation, batch D10 significantly reduced total histopathological scores and biochemical severity indices at a clinically-equivalent dose but batch D1 did not. The content of naringin, narirutin and baicalin in batches D1, D5 and D9 consistently exceeds the upper limit of the predicted value. Eight markers whose lower limit is predicted to be close to 0 contributed less to the material basis for AP protection.. Despite qualified materials used for CQCQD preparation, the clinical effect depends on appropriate content range of Q-markers. Emodin, rhein, aloe emodin, magnolol, hesperidin, synephrine, baicalein, and geniposide are considered as vital Q-markers in the primary screen. This study proposed a feasible platform for producing highly consistent batches of CQCQD in future study.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Mice; Necrosis; Pancreas; Pancreatitis; Quality Control

Chronic pancreatitis (CP) is an inflammatory disease of the pancreas characterized by ductal obstructions, tissue fibrosis, atrophy and exocrine and endocrine pancreatic insufficiency. However, our understanding is very limited concerning the disease's progression from a single acute inflammation, via recurrent acute pancreatitis (AP) and early CP, to the late stage CP. Poly(ADP-ribose) polymerase 1 (PARP1) is a DNA damage sensor enzyme activated mostly by oxidative DNA damage. As a co-activator of inflammatory transcription factors, PARP1 is a central mediator of the inflammatory response and it has also been implicated in acute pancreatitis. Here, we set out to investigate whether PARP1 contributed to the pathogenesis of CP. We found that the clinically used PARP inhibitor olaparib (OLA) had protective effects in a murine model of CP induced by multiple cerulein injections. OLA reduced pancreas atrophy and expression of the inflammatory mediators TNFα and interleukin-6 (IL-6), both in the pancreas and in the lungs. Moreover, there was significantly less fibrosis (Masson's trichrome staining) in the pancreatic sections of OLA-treated mice compared to the cerulein-only group. mRNA expression of the fibrosis markers TGFβ, smooth muscle actin (SMA), and collagen-1 were markedly reduced by OLA. CP was also induced in PARP1 knockout (KO) mice and their wild-type (WT) counterparts. Inflammation and fibrosis markers showed lower expression in the KO compared to the WT mice. Moreover, reduced granulocyte infiltration (tissue myeloperoxidase activity) and a lower elevation of serum amylase and lipase activity could also be detected in the KO mice. Furthermore, primary acinar cells isolated from KO mice were also protected from cerulein-induced toxicity compared to WT cells. In summary, our data suggest that PARP inhibitors may be promising candidates for repurposing to treat not only acute but chronic pancreatitis as well.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Disease Models, Animal; Fibrosis; Inflammation; Interleukin-6; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis; Pancreatitis, Chronic; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Acute pancreatitis is a common inflammatory disorder of the exocrine pancreas with no specific therapy. Intracellular nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) salvage pathway, is involved in many inflammatory disorders. In this study, we investigated the role of NAMPT in experimental acute pancreatitis.. Acute pancreatitis was induced in mice using three disparate models: (1) caerulein hyperstimulation, (2) ethanol plus palmitoleic acid, and (3) retrograde biliopancreatic ductal infusion of sodium taurocholate. The NAMPT inhibitor FK866 and NAMPT downstream product nicotinamide mononucleotide (NMN) was administered. Serum and pancreas were collected and analyzed biochemically and histologically. Bone marrow derived macrophages were isolated, cultured with cytokines or pancreatic acini, then analyzed by quantitative PCR and non-targeted metabolomics.. The levels of pancreatic NAMPT and NAD were down-regulated upon acute pancreatitis. NAMPT inhibitor FK866 suppressed M1 macrophage polarization while NMN boosted it. In co-culture of macrophages with acinar cells, inhibition of NAMPT prevented M1-like macrophage differentiation induced by injured pancreatic acini. The injured pancreatic acinar milieu induced a unique metabolic signature linked to macrophage polarization, and inhibition of NAMPT reversed these metabolites changes. Furthermore, NMN supplementation aggravated caerulein hyperstimulation pancreatitis and alcoholic pancreatitis, and inhibition of NAMPT protected against caerulein hyperstimulation, alcoholic and biliary acute pancreatitis and reducing pancreatic macrophage infiltration in vivo.. NAMPT inhibition protects against acute pancreatitis via preventing M1 macrophage polarization and restoring the metabolites related to macrophage polarization and that NAMPT could be a promising therapeutic target for acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Cytokines; Macrophages; Mice; NAD; Nicotinamide Mononucleotide; Nicotinamide Phosphoribosyltransferase; Pancreatitis; Sirtuin 1

/Objectives: The pathogenesis of hyperglycemia during acute pancreatitis (AP) remains unknown due to inaccessibility of human tissues and lack of animal models. We aimed to develop an animal model to study the mechanisms of hyperglycemia and impaired glucose tolerance in AP.. We injected ferrets with intraperitoneal cerulein (50 μg/kg, 9 hourly injections) or saline. Blood samples were collected for glucose (0, 4, 8, 12, 24h); TNF-α, IL-6 (6h); amylase, lipase, insulin, glucagon, pancreatic polypeptide (PP), glucagon-like peptide-1 (GLP-1), and gastric inhibitory polypeptide (GIP) (24h). Animals underwent oral glucose tolerance test (OGTT), mixed meal tolerance test (MMTT) at 24h or 3 months, followed by harvesting pancreas for histopathology and immunostaining.. Cerulein-injected ferrets exhibited mild pancreatic edema, neutrophil infiltration, and elevations in serum amylase, lipase, TNF-α, IL-6, consistent with AP. Plasma glucose was significantly higher in ferrets with AP at all time points. Plasma glucagon, GLP-1 and PP were significantly higher in cerulein-injected animals, while plasma insulin was significantly lower compared to controls. OGTT and MMTT showed abnormal glycemic responses with higher area under the curve. The hypoglycemic response to insulin injection was completely lost, suggestive of insulin resistance. OGTT showed low plasma insulin; MMTT confirmed low insulin and GIP; abnormal OGTT and MMTT responses returned to normal 3 months after cerulein injection.. Acute cerulein injection causes mild acute pancreatitis in ferrets and hyperglycemia related to transient islet cell dysfunction and insulin resistance. The ferret cerulein model may contribute to the understanding of hyperglycemia in acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Blood Glucose; Ceruletide; Ferrets; Gastric Inhibitory Polypeptide; Glucagon; Glucagon-Like Peptide 1; Humans; Hyperglycemia; Insulin; Insulin Resistance; Interleukin-6; Lipase; Pancreatitis; Tumor Necrosis Factor-alpha

Symptoms associated with acute pancreatitis can be debilitating, and treatment remains a challenge. This study aimed to investigate the efficacy of selectively inhibiting the soluble form of TNF (solTNF) using the biologic XPro1595 in a mouse model of acute pancreatitis.. Acute pancreatitis was induced in adult male C57Bl/6J mice by administering cerulein (8 injections of 50 µg/kg I.P., spaced an hour apart), with XPro1595 (10 mg/kg, S.C.) or vehicle being administered approximately 18 h after the last injection. Serum was collected 6 or 18 h after the last cerulein injection, pancreatic tissue was collected 2 and 7 days post-induction, and brain hippocampal tissue was collected at 7 days post-induction. The animal's pain level was assessed 3, 5 and 7 days post-induction.. The induction of acute pancreatitis promoted a strong increase in serum amylase levels, which had receded back to baseline levels by the next morning. XPro1595 treatment began after amylase levels had subsided at 18 h, and prevented pancreatic immune cell infiltration, that subsequently prevented tissue disruption and acinar cell death. These improvements in pathology were associated with a significant reduction in mechanical hypersensitivity (neuropathic pain). XPro1595 treatment also prevented an increase in hippocampal astrocyte reactivity, that may be associated with the prevention of neuropathic pain in this mouse model.. Overall, we observed that selectively inhibiting solTNF using XPro1595 improved the pathophysiological and neurological sequelae of cerulein-induced pancreatitis in mice, which provides support of its use in patients with pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Humans; Male; Mice; Pancreas; Pancreatitis; Tumor Necrosis Factor-alpha

Obesity is one of the most important risk factors for acute pancreatitis. Based on the effect of sleeve gastrectomy (SG) on improving body weight and blood lipids, we investigated whether SG is beneficial in improving pancreatitis in obese rats.. Two studies were used to evaluate the effect of SG on the first onset of pancreatitis and acute episodes of recurrent pancreatitis in obese rats. A high-fat diet (HFD) for 8 weeks resulted in obesity in rats. Study 1: Obese rats were treated with SG and sham surgery. Pancreatitis was induced by intraperitoneal injection of cerulein at 6 weeks after surgery. The severity of pancreatitis was assessed by histological examination, cytokines, and infiltration of inflammatory cells. Study 2 performed the same procedure as in study 1, except that rats were intraperitoneally injected with a small dose of cerulein three times a week for 6 weeks before surgery to induce recurrent pancreatitis.. The body weight, food intake, and blood lipids of SG rats in study 1 and study 2 were significantly lower than those of sham rats during the 6 weeks after surgery. Compared with sham rats, SG rats in both studies had fewer inflammatory cytokines, inflammatory cell infiltration, and pathological injury in the pancreas after cerulein-induced acute pancreatitis.. SG reduces the severity of the first onset of pancreatitis and the seriousness of acute episodes of recurrent pancreatitis. The improvement of lipid metabolism and body weight by SG may play an important role in this effect.

Topics: Acute Disease; Animals; Ceruletide; Gastrectomy; Obesity; Obesity, Morbid; Pancreatitis; Rats

The management of acute pancreatitis (AP) remains a challenge to clinicians worldwide for limited effective interventions. Retinoid orphan receptor gamma t (RORγt) is a therapeutic target for several diseases; however, it is unclear whether inhibiting RORγt can ameliorate AP. The relative expression of RORγt, IL-17 and IL-23 in the peripheral blood mononuclear cells of AP patients was measured by RT-PCR. An AP mouse model was induced by ceruletide, and SR1001 was injected before ceruletide administration. RORγt+ cells, T helper 17 cells (Th17), regulatory T cells (Tregs) and γδ T cells were assessed in the pancreas and spleen by flow cytometry. Higher RORγt expression in patients indicated the potential role of RORγt in AP progression. Analyses of the IL-17/IL-23 axis confirmed its role. SR1001 significantly alleviated AP histologically in the mouse model. Serum levels of amylase, IL-6, TNFalpha, IL-17 and IL-23 decreased upon SR1001 treatment. SR1001 selectively decreased the number of RORγt+, Th17, Tregs and γδ T cells in the pancreas but not the spleen. Collectively, these results showed that SR1001 exerted therapeutic effects on AP by suppressing IL-17-secreting Th17 and γδ T cells in the pancreas. Thus, SR1001 may be a promising drug for the treatment of AP in the clinic.

Topics: Acute Disease; Adult; Aged; Animals; Case-Control Studies; Ceruletide; Disease Models, Animal; Disease Progression; Female; Humans; Interleukin-17; Intraepithelial Lymphocytes; Leukocytes, Mononuclear; Male; Mice; Mice, Inbred C57BL; Middle Aged; Nuclear Receptor Subfamily 1, Group F, Member 3; Pancreatitis; Sulfonamides; Th17 Cells; Thiazoles

Acute pancreatitis (AP) is a disorder of the pancreas marked by profound inflammation and oxidative stress. Phytoconstituents presents an important toolbox of preventive strategies to combat inflammatory disorders. To this end, we selected the active constituent of Crocus sativus, crocin for evaluation against cerulein-induced AP, owing to its promising antiinflammatory activity in acute as well as chronic inflammatory conditions. The animals were randomly divided into five groups comprising of normal control, cerulein control, crocin low dose (30 mg/kg), crocin high dose (100 mg/kg), and crocin control (100 mg/kg). Various biochemical parameters and the levels of inflammatory cytokines and p65-NFκB were measured. The mechanism was investigated by histology and immunohistochemistry. We found that crocin significantly reduced the pancreatic edema, amylase, and lipase levels. It abrogated the oxidative stress incurred by cerulein challenge. We found that crocin modulated the pancreatic inflammatory cytokine levels. Crocin perturbed the nuclear translocation of p65-NFκB. Crocin reverted the pancreatic histology associated with AP. Furthermore, it upregulated the expression of Nrf-2 and downregulated the expression of IL-6, TNF-α, nitrotyrosine, and NFκB. Cumulatively, these results indicate that crocin has promising potential to prevent cerulein induced AP and regular intake of saffron can prove beneficial for the pancreatic health.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Carotenoids; Ceruletide; Crocus; Cytoprotection; Male; Mice; Oxidative Stress; Pancreas; Pancreatitis; Phytotherapy

In this study, the aim was to examine the potential protective effects of Myrtus communis subsp. communis leaf ethanol extract (MC) treatment against acute pancreatitis (AP) in rats. Thirty-two rats were grouped as the saline-pretreated control (C), MC-pretreated control (MC), saline-pretreated AP (AP), and MC-pretreated AP (MC + AP) groups. To induce AP, cerulein was administered (50 µg/kg) two times. The rats were given MC for 14 days before cerulein injection. Six hours after the final cerulein injection, the rats were sacrificed. Pancreatic damage was associated with an increase in the serum activity of lipase and amylase, the pancreatic activity of myeloperoxidase, and the pancreatic level of malondialdehyde, interleukin-1β, and interleukin-6. AP also led to a decrease in the pancreatic level of anti-inflammatory interleukin-10 and glutathione. Pretreatment with MC before the induction of AP significantly reduced the pancreatic damage observed during the histological examination as well as reversed the biochemical changes evoked by AP. PRACTICAL APPLICATIONS: Acute pancreatitis is characterized by high mortality (average about 5%; severe cases may reach about 30%). The current treatment for acute pancreatitis is mainly symptomatic. The introduction of herbal drugs may lead to the development of a new strategy in the treatment of this disease. This study revealed that MC reduced pancreatic injury by decreasing pro-inflammatory cytokines, increasing antioxidant capacity and anti-inflammatory cytokine, IL-10. To the authors' knowledge, this research is the first report showing that MC inhibits the development of AP. This observation suggests that MC may be useful in the prevention and the treatment of AP in clinical settings.

Topics: Acute Disease; Animals; Ceruletide; Myrtus; Pancreatitis; Plant Extracts; Rats

In this study, we investigated the anti-inflammatory effects of silymarin on cerulein-induced acute pancreatitis (AP) in mice.. Cerulein (50 μg/kg) was injected intraperitoneally once hourly for 6 hours to induce AP. To investigate the prophylactic effects of silymarin, dimethyl sulfoxide or silymarin (25, 50, and 100 mg/kg) was injected intraperitoneally 1 hour before cerulein injection. To investigate the therapeutic effects of silymarin, dimethyl sulfoxide or silymarin (100 mg/kg) was injected intraperitoneally 1, 3, or 5 hours after the first cerulein injection. Blood, pancreas, and lungs were harvested 6 hours after the last cerulein injection.. Pre- and posttreatment with silymarin decreased the pancreas weight/body weight ratio and serum amylase activity. Furthermore, silymarin treatment inhibited pancreas and lung injury and neutrophil infiltration during cerulein-induced AP. In addition, silymarin inhibited increased secretion of proinflammatory cytokines such as interleukin 1β, interleukin 6, and tumor necrosis factor α. Finally, mitogen-activated protein kinases (MAPKs) and nuclear factor-κB were activated by cerulein, and only p38 in MAPK was inhibited by silymarin.. These findings suggest that silymarin attenuates the severity of AP through inhibition of p38 MAPKs and that silymarin could be a potential prophylactic and therapeutic agent for the treatment of AP.

Topics: Acute Disease; Amylases; Animals; Antioxidants; Ceruletide; Cytokines; Inflammation Mediators; Mice, Inbred C57BL; Mitogen-Activated Protein Kinases; NF-kappa B; Organ Size; Pancreas; Pancreatitis; Protective Agents; Severity of Illness Index; Silymarin

The mechanisms underlying pathogenesis of acute pancreatitis (AP) are still not completely understood. An early, critical feature of AP is aberrant calcium (Ca) signaling, termed Ca overload, within pancreatic acinar cells. This study aimed to develop a model system in rats for AP induction to study the contribution of the Na-Ca exchanger 1 (NCX1) ion channel in AP pathogenesis.. To establish a rat model of AP induction, cerulein or L-arginine were intraperitoneally injected and tissue was histologically analyzed by hematoxylin and eosin staining. A cell culture-based model for AP induction was similarly created through cerulein treatment of AR42J cells. Induction of AP was also examined following exposure to the NXC1-targeted inhibitor KB-R7943. The expression of each gene was detected by Western blotting, immunofluorescence, immunohistochemistry, or quantitative reverse transcription polymerase chain reaction. Transcriptional regulation by nuclear factor (NF)-κB was detected using an NCX1 promoter-fusion dual luciferase reporter system. Cytosolic Ca was measured using a fluorescent calcium indicator.. We found that cerulein induced NCX1 expression via activation of nuclear factor NF-κB, which potentially binds to the NCX1 promoter to induce its transcription.. Our findings reveal a regulatory pathway through NF-κB/NCX1 governing Ca overload in AP development, thus providing potential targets for AP treatment.

Topics: Acinar Cells; Acute Disease; Animals; Calcium; Calcium Signaling; Cell Line, Tumor; Ceruletide; Gene Expression Regulation; Intracellular Space; Male; NF-kappa B; Pancreas; Pancreatitis; Rats, Sprague-Dawley; Sodium-Calcium Exchanger; Up-Regulation

The aim of this study was to investigate the effects of eupatilin on protein kinase D1 (PKD1) and nuclear factor kappa B (NF-κB) signaling pathways in cerulein-induced in vitro pancreatitis.. We used collagenase digestion to isolate pancreatic acinar cells from male C57BL/6 mice. In vitro acute pancreatitis was induced by treatment with a supramaximal dose of cerulein. Eupatilin was pretreated before stimulation with cerulein.. Eupatilin significantly reduced cerulein-induced amylase release in pancreatic acini. Eupatilin treatment downregulated cerulein-induced expression of interleukin (IL)-1β, IL-6, and CC chemokine ligands 2 and 5, but it upregulated expression of IL-4 and IL-10. We demonstrated that eupatilin pretreatment attenuated cerulein-induced necrosis in isolated pancreatic acinar cells. This effect of eupatilin was confirmed by lactic dehydrogenase assay, fluorescence-activated cell sorting analysis, and cytopathologic analysis. Eupatilin inhibited cerulein-induced activation of PKD1/NF-κB and the nuclear translocation of NF-κB.. Our data demonstrated that eupatilin is a potential therapeutic candidate for the treatment of pancreatitis through its ability to reduce cellular necrosis and inflammatory responses by inhibition of the PKD1/NF-κB signaling pathway.

Topics: Acinar Cells; Active Transport, Cell Nucleus; Acute Disease; Amylases; Animals; Cell Nucleus; Ceruletide; Cytokines; Flavonoids; Male; Mice, Inbred C57BL; NF-kappa B; Pancreatitis; Protein Kinase C; Signal Transduction

Disturbances in pancreatic microcirculation, beginning with vasoconstriction, are crucial in early pancreatitis and progression to necrotizing pancreatitis. Thus, vascular-targeted treatment aiming to restore a sufficient level of microcirculation through vasodilation would possibly reduce the severity of pancreatitis. Lidocaine is an anti-arrhythmic and local anesthetic drug, which also acts as a vasodilator at higher concentrations.. To evaluate the efficacy of intra-arterial infusion of lidocaine into the celiac trunk in treatment of cerulein-induced acute pancreatitis.. Wistar rats (n = 20) were randomly divided into 2 equal groups: the control group (NaCl group, n = 10) and the study group (lidocaine group, n = 10). All subjects underwent surgical intervention with intra-arterial infusion of 0.9% NaCl (control group) or 1% lidocaine hydrochloride (study group) into the celiac trunk. Blood samples were collected 5 times at regular intervals from each rat for amylase and lipase measurements. Histopathological analysis of the pancreas was performed.. A total number of 16 rats (control group n = 7, study group n = 9) were included. In the postoperative course, the study group (lidocaine group) revealed lower values of serum amylase and lipase levels compared to the control group (NaCl group), except the values at the 1st treatment point, which appeared 1 h after intraoperative drug injection. Significantly lower treatment endpoint levels of pancreatic enzymes were seen in the lidocaine group. Moreover, no differences were observed between the 1st and the last treatment point in the control group; however, these differences were significant for both enzymes in the study group. Histopathology revealed reduced pancreatitis severity in the study group compared to the controls.. Intra-arterial lidocaine infusion into the celiac trunk decreases pancreatitis severity. What is more, this study demonstrates the relevance of early vasodilation in the therapy of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Infusions, Intra-Arterial; Lidocaine; Pancreas; Pancreatitis; Random Allocation; Rats; Rats, Wistar; Treatment Outcome

Acute pancreatitis (AP) is the inflammatory response of the exocrine pancreas to various causes. Modafinil has significant anti-inflammation and anti-oxidation effects. No experiment has assessed the effects of modafinil on AP. Thus, the study aims to study the effects of modafinil on AP and its potential mechanism in vivo and vitro. 5% sodium taurocholate was retrograde injected into pancreatic duct to establish AP rat model. The severity of AP was detected by HE staining, serum amylase and lipase levels. The inflammation, oxidative stress and apoptosis were detected separately by ELISA, MDA and SOD kits, tunnel staining and Western blotting in rats. Besides, SNIP1 expression was analyzed by qPCR and Western blotting. In vivo, AR42J cells were stimulated by cerulein and lipopolysaccharide to establish AP cell model. Flow cytometry examined cell apoptosis. After the plasmids silencing SNIP1 were transfected into AP cells, the inhibitory effects of modafinil on inflammation, oxidative stress and apoptosis were significantly reversed. The results indicated that modafinil showed significant curative and therapeutic effects by regulating SNIP1 level.

Topics: Acute Disease; Animals; Cell Line; Ceruletide; Inflammation; Modafinil; Pancreatitis; Rats; RNA-Binding Proteins

Acute pancreatitis (AP) is a sudden inflammatory process of the pancreas that may also involve surrounding tissues and/or remote organs. Inflammation and parenchymal cell death are common pathological features of this condition and determinants of disease severity. Polyethylene glycols (PEGs) are non-immunogenic, non-toxic water-soluble polymers widely used in biological, chemical, clinical and pharmaceutical settings.. To evaluate the protective effect of a 35-kDa molecular weight PEG (PEG35) on the pancreatic damage associated to cerulein-induced acute pancreatitis. Wistar rats were assigned at random to a control group, a cerulein-induced AP group and a PEG35 treatment group. AP was induced by five hourly intraperitoneal injections of cerulein (50 μg/kg/bw), while the control animals received saline solution. PEG35 was administered intraperitoneally 10 minutes before each cerulein injection in a dose of 10 mg/kg. After AP induction, samples of pancreatic tissue and blood were collected for analysis. AR42J pancreatic acinar cells were treated with increasing concentrations of PEG35 prior to exposure with tumor necrosis factor α (TNFα), staurosporine or cerulein. The severity of AP was determined on the basis of plasma levels of lipase, lactate dehydrogenase activity, pancreatic edema and histological changes. To evaluate the extent of the inflammatory response, the gene expression of inflammation-associated markers was determined in the pancreas and in AR42J-treated cells. Inflammation-induced cell death was also measured in models of. Administration of PEG35 significantly improved pancreatic damage through reduction on lipase levels and tissue edema in cerulein-induced AP rats. The increased associated inflammatory response caused by cerulein administration was attenuated by a decrease in the gene expression of inflammation-related cytokines and inducible nitric oxide synthase enzyme in the pancreas. In contrast, pancreatic tissue mRNA expression of interleukin 10 was markedly increased. PEG35 treatment also protected against inflammation-induced cell death by attenuating lactate dehydrogenase activity and modulating the pancreatic levels of apoptosis regulator protein BCL-2 in cerulein hyperstimulated rats. Furthermore, the activation of pro-inflammatory markers and inflammation-induced cell death in pancreatic acinar cells treated with TNFα, cerulein or staurosporine was significantly reduced by PEG35 treatment, in a dose-dependent manner.. PEG35 ameliorates pancreatic damage in cerulein-induced AP and AR42J-treated cells through the attenuation of the inflammatory response and associated cell death. PEG35 may be a valuable option in the management of AP.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Pancreas; Pancreatitis; Polyethylene Glycols; Rats; Rats, Wistar

Astaxanthin (ATX) is a naturally occurring carotenoid and a potent antioxidant. Various anti-inflammatory effects of ATX have been examined. We aimed to investigate the protective effect of ATX and its mechanism in a cerulein-induced acute pancreatitis rat model.. The rats were randomized into 2 main groups as control (C) and acute pancreatitis group (AP). AP group was subsequently divided into subgroups as AP+vehicle (AP), AP+ATX, and ATX+peroxisome proliferator-activated receptor-alpha antagonist GW6471 (ATX+GW) groups. To induce AP, the rats were administered cerulein (50 µg/kg, intraperitonally [ip]) at 1 hour intervals, whereas the C group received saline. The AP group was treated with vehicle olive oil, ATX 40 mg/kg/orally, or GW6471 and ATX (GW1 mg/kg/ip; ATX; 40 mg/kg/peroral). Treatments were administered after the 1st cerulein injection. At the 7th hour after the final injection, the rats were killed and the pancreatic tissue was used for the determination of malondialdehyde (MDA), glutathione (GSH), and myeloperoxidase (MPO) activities and luminol-lucigenin chemiluminescence levels. Serum amylase, lipase, and histopathological analyses were performed.. Elevated serum lipase and amylase levels in the vehicle-treated AP group (p<0.01) decreased in the ATX and ATX+GW groups (p<0.05). In the AP groups, GSH was reduced and MDA, MPO, luminol, and lucigenin levels were increased (p<0.05-0.001). ATX reversed these changes (p<0.05-0.001). The vehicle-treated group revealed significant severe cytoplasmic degeneration and vacuolization, whereas ATX ameliorated these destructions. GW6471 did not abolish the positive effects of ATX biochemically or histologically.. ATX has a potent protective effect on AP via its radical scavenging and antioxidant properties. Therefore, we believe that ATX may have therapeutic potential.

Topics: Acute Disease; Animals; Antioxidants; Ceruletide; Disease Models, Animal; Oxidative Stress; Pancreas; Pancreatitis; Rats; Xanthophylls

Vagus nerve stimulation (VNS) is effective in reducing inflammation in various diseases, such as rheumatoid arthritis, colitis and acute kidney injury. The anti-inflammatory effect of vagus nerve in these diseases necessitates the interactions of neural activation and α7 nicotinic acetylcholine receptors (α7nAChRs) on splenic macrophages. In this study, we aimed to investigate the effect of VNS on severity in experimental acute pancreatitis (AP).. Two independent AP models were used, which induced in ICR mice with caerulein or pancreatic duct ligation (PDL). Thirty minutes after modeling, the left cervical carotid sheath containing the vagus nerve was electrically stimulated for 2 min. Plasma lipase and amylase activities, TNF-α levels and pancreas histologic damage were evaluated. In caerulein mice, the percentages of α7nAChR. VNS reduced plasma lipase and amylase activities, blunted the concentrations of TNF-α and protected against pancreas histologic damage in two AP models. Survival rates were improved in the PDL model after VNS. In caerulein AP mice, VNS increased the percentages of α7nAChR. VNS reduces disease severity and attenuates inflammation in AP mice. This effect is independent of spleen and is probably related to α7nAChR on macrophage.

Topics: Acute Disease; Adoptive Transfer; Animals; Biomarkers; Ceruletide; Disease Models, Animal; Evoked Potentials; Immunohistochemistry; Male; Mice; Pancreatitis; Severity of Illness Index; Survival Rate; Vagus Nerve Stimulation

During pancreatitis, autophagy is activated, but lysosomal degradation of dysfunctional organelles including mitochondria is impaired, resulting in acinar cell death. Retrospective cohort analyses demonstrated an association between simvastatin use and decreased acute pancreatitis incidence.. We examined whether simvastatin can protect cell death induced by cerulein and the mechanisms involved during acute pancreatitis. Mice were pretreated with DMSO or simvastatin (20 mg/kg) for 24 h followed by 7 hourly cerulein injections and sacrificed 1 h after last injection to harvest blood and tissue for analysis.. Pancreatic histopathology revealed that simvastatin reduced necrotic cell death, inflammatory cell infiltration and edema. We found that cerulein triggered mitophagy with autophagosome formation in acinar cells. However, autophagosome-lysosome fusion was impaired due to altered levels of LAMP-1, AMPK and ULK-1, resulting in autophagosome accumulation (incomplete autophagy). Simvastatin abrogated these effects by upregulating LAMP-1 and activating AMPK which phosphorylated ULK-1, resulting in increased formation of functional autolysosomes. In contrast, autophagosomes accumulated in control group during pancreatitis. The effects of simvastatin to promote autophagic flux were inhibited by chloroquine. Mitochondria from simvastatin-treated mice were resistant to calcium overload compared to control, suggesting that simvastatin induced mitochondrial quality control to eliminate susceptible mitochondria. Clinical specimens showed a significant increase in cell-free mtDNA in plasma during pancreatitis compared to normal controls. Furthermore, genetic deletion of parkin abrogated the benefits of simvastatin.. Our findings reveal the novel role of simvastatin in enhancing autophagic flux to prevent pancreatic cell injury and pancreatitis.

Topics: Acute Disease; Animals; Anticholesteremic Agents; Autophagy; Ceruletide; Lysosomes; Male; Membrane Fusion; Mice, Inbred C57BL; Pancreatitis; Phagosomes; Simvastatin

Acute pancreatitis (AP) is a severe inflammatory disease. Caerulin induces significant pro-inflammatory responses in macrophages, causing serve damage to pancreatic acinar cells. The potential role of Rab GTPase 21 (Rab21) in this process was tested in this study. In murine bone marrow-derived macrophages (BMDMs), caerulin induced Rab21-TRAF3-MKK3 complex association. Rab21 silencing (by targeted shRNAs) or knockout (by CRISPR/Cas9 method) largely inhibited caerulin-induced MKK3-TRAF3 association, downstream MKK3-p38 activation and production of several pro-inflammatory cytokines (IL-1β, TNF-α and IL-17). Conversely, ectopic Rab21 overexpression in BMDMs potentiated caerulin-induced MKK3-TRAF3 association and pro-inflammatory cytokines production. The cytotoxicity of caerulin-activated BMDMs to co-cultured pancreatic acinar cells was alleviated by Rab21 knockdown or knockout, but exacerbated with Rab21 overexpression. In vivo, administration of Rab21 shRNA lentivirus significantly attenuated pancreatic and systemic inflammations in caerulin-injected AP mice. Collectively, our results suggest that Rab21 mediates caerulin-induced MKK3-p38 activation and pro-inflammatory responses.

Topics: Acinar Cells; Acute Disease; Animals; Cell Death; Ceruletide; Cytokines; Enzyme Activation; Inflammation; Inflammation Mediators; Macrophages; MAP Kinase Kinase 3; Mice, Inbred C57BL; Mice, Knockout; p38 Mitogen-Activated Protein Kinases; Pancreas; Pancreatitis; rab GTP-Binding Proteins; RNA, Small Interfering; TNF Receptor-Associated Factor 3

In a continuation of previous work, Reg3γ protein was further evaluated as a biomarker of pancreatic injury using immunohistochemistry in an additional species.. Mice and rats were treated with intraperitoneal cerulein injections, creating acute pancreatic injury. Mice received 2, 4, or 6 doses, and rats received 1, 2, or 3 doses of cerulein creating low, medium, and high treatment groups. Control animals were dosed with phosphate-buffered saline at corresponding volumes and intervals. Groups of 6 animals were killed 1, 3, 6, 24, and 48 hours after final treatments. Reg3γ immunohistochemical staining and image analysis were performed on pancreatic tissue obtained 6, 24, or 48 hours after control or cerulein treatment. Staining was quantified using image analysis software to calculate area of positivity as a percentage of total tissue area.. Percent positivity of Reg3γ in both species rose by 6 hours, peaked by 24 hours across all 3 cerulein doses, and dropped significantly by 48 hours. In high-dose rats with accompanying gene expression data, Reg3γ gene expression corresponded temporally with quantitative staining data.. Reg3γ staining quantified through image analysis showed a time- and dose-response in cerulein-treated mice and rats.

Topics: Abdominal Injuries; Acute Disease; Animals; Ceruletide; Disease Models, Animal; Gene Expression Regulation; Immunohistochemistry; Injections, Intraperitoneal; Male; Mice, Inbred C57BL; Pancreatic Diseases; Pancreatitis-Associated Proteins; Rats, Sprague-Dawley; Time Factors

In the current study, we explored the role of extracellular ATP (eATP) in promoting systemic inflammation during development of acute pancreatitis (AP). Release of extracellular (e)ATP was evaluated in plasma and bronchoalveolar lavage fluid (BALF) of mice with experimental acute pancreatitis (AP). Prophylactic intervention using apyrase or suramin was used to understand the role and contribution of eATP in pancreatitis-associated systemic injury. AP of varying severity was induced in C57BL/6 mice using 1-day or 2-day caerulein, caerulein + LPS and l-arginine models. eATP was measured in plasma and BALF. Mice were treated with suramin or apyrase in the caerulein and l-arginine models of AP. Plasma cytokines, lung, and pancreatic myeloperoxidase, and morphometric analysis of pancreatic and lung histology, were used to assess the severity of pancreatitis. Plasma eATP and purinergic 2 (P2) receptors in the pancreas and lungs were significantly elevated in the experimental models of AP. Blocking the effect of eATP by suramin led to reduced levels of plasma IL-6 and TNFα as well as reduced lung, and pancreatic injury. Neutralizing eATP with apyrase reduced systemic injury but did not ameliorate local injury. The results of this study support the role of eATP and P2 receptors in promoting systemic inflammation during AP. Modulating purinergic signaling during AP can be an important therapeutic strategy in controlling systemic inflammation and, thus, systemic inflammatory response syndrome during AP.

Topics: Acute Disease; Adenosine Triphosphate; Animals; Apyrase; Arginine; Bronchoalveolar Lavage Fluid; Ceruletide; Cytokines; Inflammation; Lung; Mice; Mice, Inbred C57BL; Pancreas; Pancreatitis; Peroxidase; Receptors, Purinergic; Signal Transduction; Suramin

BACKGROUND Acute pancreatitis (AP) has a high mortality rate and often has serious complications. The Hippo-YAP signaling pathway is mainly involved in cell proliferation and stem cell self-renewal. Recent studies have reported that YAP1 plays a crucial role in pancreatic cancer initiation and acute and chronic pancreatitis (CP). However, the role of YAP1 in AP still needs to be clarified. MATERIAL AND METHODS To assess the role of YAP1 in the progression of AP, we established a cell model of AP in AR42J cells. AR42J, a rat pancreatic acinar cell line, was stimulated with caerulein to mimic AP-like acinar cell injury. Levels of interleukin (IL)-6 and tumor necrosis factor-alpha (TNF-alpha) were measured by ELISA to investigate the role of YAP1 in the progression of AP. RESULTS The results showed that YAP1 and MALAT1 were the targets of miR-194 and were upregulated in caerulein-treated AR42J cells. Overexpression of MALAT1 or YAP1 can increase the levels of IL-6 and TNF-alpha secreted by AR42J cells, while miR-194 dramatically counteracts this enhancement effect. CONCLUSIONS Our results demonstrated a regulation loop among MATAL1, miR-194, and YAP1, which dynamically regulates the progression of AP, providing a new therapeutic target for treatment of this disease.

Topics: Acute Disease; Animals; Apoptosis Regulatory Proteins; Base Sequence; Cell Line; Ceruletide; Cytoprotection; Disease Progression; Down-Regulation; Feedback, Physiological; MicroRNAs; Pancreatitis; Protein Binding; Rats; RNA, Long Noncoding; Signal Transduction; Up-Regulation; YAP-Signaling Proteins

Acute pancreatitis (AP) is a common clinical critical disease with high mortality and the exact pathogenesis is not fully elucidated. The present study aimed to uncover the function of miR-135a in the proliferation, apoptosis, and inflammatory characteristics of diseased pancreatic cells and the potential molecular mechanisms. The expression patterns of miR-135a and family with sequence similarity 129 member A (FAM129A) in patients with AP were analyzed on the basis of the GEO database. The transfection efficiency and expression level of miR-135a in AR42J cells were determined by qRT-PCR. The biological characteristics of AR42J cells treated with cerulein were detected by cell counting kit-8 (CCK-8), flow cytometry, and western blot assays. The potential interaction between miR-135a and FAM129A was confirmed by bioinformatics prediction softwares and luciferase reporter assay. MiR-135a inhibitor and pcDNA3.1-FAM129A were co-transfected to determine the regulation of miR-135a/FAM129A on inflammatory AR42J cell injury. We observed that miR-135a was highly expressed in AP samples. Depletion of miR-135a could alleviate the condition so that the AR42J cells proliferation increased, apoptosis decreased, and the expression of inflammatory cytokines enhanced. In addition, mRNA and protein expression of FAM129A were negatively regulated by miR-135a, and over-expression of FAM129A could strengthen the relief effect of miR-135a inhibitor in AP induced by cerulein. In summary, our data demonstrates that silencing miR-135a reduces AR42J cells injury and inflammatory response in AP induced by cerulein through targeting FAM129A.

Topics: Acute Disease; Animals; Apoptosis; Biomarkers, Tumor; Cell Line; Cell Line, Tumor; Cell Proliferation; Ceruletide; Cytokines; HEK293 Cells; Humans; Inflammation; MicroRNAs; Neoplasm Proteins; Pancreatitis; Rats; RNA, Messenger; Transfection

Nesfatin-1, a recently discovered peptide, was shown to have anti-inflammatory effects. Acute pancreatitis (AP) is a life-threatening condition caused by various reasons. Although the etiology of AP is well-known, its pathogenesis is not clear. The aim of this study is to investigate the possible anti-inflammatory role of nesfatin-1 and its probable protective underlying mechanisms in an acute pancreatitis model. Caerulein was applied intraperitoneally to induce acute pancreatitis in Sprague-Dawley female rats. Nesfatin-1 was administered 5 minutes before the application of caerulein to determine its potential anti-inflammatory role on AP. Five minutes before nesfatin-1 injection, in order to investigate the underlying mechanism, oxytocin receptor antagonist (atosiban), melanocortin receptor antagonist (HS024), or ghrelin receptor antagonist (cortistatin) were administered. Five minutes after nesfatin-1 administration, two doses of caerulein were applied one hour apart. The rats were sacrified 12 hours after the first caerulein dose for serum and pancreatic tissue sampling. Microscopic damage scoring, malondialdehyde and glutathione levels, myeloperoxidase activity, luminol and lucigenin chemiluminescence levels in pancreatic tissue and amylase, lipase, trypsinogen-2 levels in serum were evaluated. Oxidative damage was decreased with nesfatin-1 treatment in the acute pancreatitis model (P < 0.05 - 0.001). The administration of HS024 reversed the effect of nesfatin-1, via increasing lipase, amylase, trypsinogen-2, malondialdehyde (MDA), myeloperoxidase (MPO) and lucigenin levels (P < 0.05 - 0.01). Atosiban pre-treatment elevated MPO activity, luminol and lucigenin chemiluminescence levels (P < 0.01 - 0.001) and cortistatin increased lucigenin and luminol chemiluminescence (P < 0.05 - 0.01). Although receptor antagonists reversed the effect of nesfatin-1 on related biochemical parameters, no significant difference was found in histological scoring. Our results indicated that nesfatin-1 had an anti-inflammatory effect on acute pancreatitis via mainly effecting melanocortin receptors.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Ceruletide; Disease Models, Animal; Female; Nucleobindins; Pancreatitis; Rats; Rats, Sprague-Dawley; Receptors, Melanocortin

Pancreatitis is a disease for which there are numerous etiologies but no effective treatments. Although the expression of the pancreatitis-associated protein-1 (PAP-1) serves as a marker for the disease, its biological function is unknown. The present study was carried out to determine if PAP-1 performs a protective role against oxidative stress-induced pancreatic cell death. For this purpose, we used cerulein-stimulated pancreatic acinar AR42J cells as an experimental model of acute pancreatitis. First, we demonstrated that PAP-1 gene expression is increased by cerulein in a dose- and time-dependent manner. In parallel, the level of active nuclear factor kappaB (NF-κB) was found to be increased in cells treated with cerulein. To test whether activation of the oxidant-sensitive transcription factor NF-κB is mediated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, the primary source of reactive oxygen species, cerulein-stimulated NADPH oxidase activity was suppressed by using the NADPH oxidase inhibitor diphenyleneiodonium and, separately, by anti-sense oligonucleotides directed against NADPH oxidase subunits p22

Topics: Acinar Cells; Acute Disease; Animals; Apoptosis; Cell Survival; Cells, Cultured; Ceruletide; Gene Expression Regulation; NADPH Oxidases; NF-kappa B; Oxidative Stress; Pancreatitis; Pancreatitis-Associated Proteins; Rats; Reactive Oxygen Species

β-Arrestins (β-arrs) are regulators and mediators of G protein-coupled receptor signaling that are functionally involved in inflammation. Nuclear factor-κB p65 (NF-κBp65) activation has been observed early in the onset of pancreatitis. However, the effect of β-arrs in acute pancreatitis (AP) is unclear. The aim of this study is to investigate whether β-arrs are involved in AP through activation of NF-κBp65.. Acute pancreatitis was induced by either caerulein injection or choline-deficient supplemented with ethionine diet (CDE). β-arr1 wild-type and β-arr1 knockout mice were used in the experiment. The survival rate was calculated in the CDE model mice. Histological and western blot analyses were performed in the caerulein model. Inflammatory mediators were detected by real-time polymerase chain reaction in the caerulein-induced AP mice. Furthermore, AR42J and PANC-1 cell lines were used to further study the effects of β-arr1 in caerulein-induced pancreatic cells.. β-Arr1 but not β-arr2 is significantly downregulated in caerulein-induced AP in mice. Targeted deletion of β-arr1 notably upregulated expression of the pancreatic inflammatory mediators including tumor necrosis factor α and interleukin 1β as well as interleukin 6 and aggravated AP in caerulein-induced mice. β-Arr1 deficiency increased mortality in mice with CDE-induced AP. Further, β-arr1 deficiency enhanced caerulein-induced phosphorylation of NF-κBp65 both in vivo and in vitro.. β-Arr1 alleviates AP via repression of NF-κBp65 activation, and it is a potentially therapeutic target for AP.

Topics: Acute Disease; Animals; beta-Arrestin 1; Cell Line, Tumor; Ceruletide; Choline Deficiency; Disease Models, Animal; Down-Regulation; Ethionine; Female; Humans; Interleukin-1beta; Interleukin-6; Mice, Knockout; Pancreatitis; Phosphorylation; Survival Rate; Transcription Factor RelA; Tumor Necrosis Factor-alpha

The goal of this study was to clarify the protective role of the Wnt/β-catenin pathway agonist SKL2001 in a rat model of Caerulein-induced acute pancreatitis. AR42J cells and rats were divided into 4 groups: control, Caerulein, SKL2001 + Caerulein, and SKL2001 + control. Cell apoptosis was examined using flow cytometry. Hematoxylin-eosin staining was performed to observe pathological changes in pancreatic and small intestinal tissues. Inflammatory cytokines were detected by enzyme-linked immunosorbent assay (ELISA), while genes related to the Wnt/β-catenin pathway were quantified using quantitative real-time PCR. In vitro results showed that Caerulein promoted cell necrosis, inhibited the Wnt/β-catenin pathway, and increased the level of inflammatory cytokines. However, SKL2001 reduced cell necrosis and inflammatory cytokines and activated the Wnt/β-catenin pathway. Additionally, in vivo results demonstrated the accumulation of fluid (i.e., edema), hemorrhage, inflammation and necrosis of the pancreatic acini occurred 6 h after the final Caerulein induction, with the damage reaching a maximal level 12 h after the final Caerulein induction; meanwhile, the Wnt/β-catenin pathway was evidently inhibited with an enhanced level of inflammatory cytokines. The aforementioned damage was further aggravated 12 h later. Nevertheless, the pancreatic and small intestinal tissue damages were alleviated in Caerulein-induced rats treated with SKL2001. In conclusion, activation of the Wnt/β-catenin pathway could inhibit Caerulein-induced cell apoptosis and inflammatory cytokine release, thus improving pancreatic and intestinal damage in rats with acute pancreatitis.

Topics: Acute Disease; Animals; beta Catenin; Ceruletide; Female; Imidazoles; Isoxazoles; Male; Pancreatitis; Rats; Rats, Sprague-Dawley; Wnt Signaling Pathway

Inflammasomes promote the production of pro-inflammatory cytokines, such as interleukin (IL)-1β and IL-18, which are the representative mediators of inflammation. Abnormal activation of inflammasomes leads to the development of inflammatory diseases such as acute pancreatitis (AP). In this study, we demonstrate the inhibitory effects of a new natural compound fraxinellone on inflammasome formation and examine the role of inflammasomes in a mouse model of AP. AP was induced with hourly intraperitoneal injections of supramaximal concentrations of the stable cholecystokinin analogue cerulein (50 μg/kg) for 6 h. Mice were sacrificed 6 h after the final cerulein injection. Blood and pancreas samples were obtained for further experiments. Intraperitoneal injection of fraxinellone significantly inhibited the pancreatic activation of multiple inflammasome molecules such as NACHT, LRR and PYD domains-containing protein 3 (NLRP3), PY-CARD, caspase-1, IL-18, and IL-1β during AP. In addition, fraxinellone treatment inhibited pancreatic injury, elevation in serum amylase and lipase activities, and infiltration of inflammatory cells such as neutrophils and macrophages but had no effect on pancreatic edema. To investigate whether inflammasome activation leads to the infiltration of inflammatory cells, we used parthenolide, a well-known natural inhibitor, and IL-1 receptor antagonist mice. The inhibition of inflammasome activation by pharmacological/or genetic modification restricted the infiltration of inflammatory cells, but not edema, consistent with the results observed with fraxinellone. Taken together, our study highlights fraxinellone as a natural inhibitor of inflammasomes and that inflammasome inhibition may lead to the suppression of inflammatory cells during AP.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Benzofurans; Cell Movement; Ceruletide; Disease Models, Animal; Female; Humans; Inflammasomes; Inflammation; Macrophages; Male; Mice; Mice, Inbred C57BL; Neutrophils; Pancreatitis

Acute pancreatitis is a life-threatening disease accompanied by systemic inflammatory response. NF-κB and p38 signal pathways are activated in AP induced by cerulein. And PAKs are multifunctional effectors of Rho GTPases with kinase activity. In the present study, the function of P21-activated kinase 1 (PAK1) in AP was investigated, and found that PAK1 was up-regulated in pancreas of AP mice model, and led to NF-κB and p38 pathway activation. PAK1 inhibition by shRNA or small molecule inhibitor FRAX597 decreased NF-κB and p38 activity, also alleviated the pathological damage in the pancreas of AP mice model, including decreasing the amylase and lipase levels in serum, decreasing the levels of tumor necrosis factor-α, interleukin-6, and interleukin-1β in AP. These results suggested that PAK1 inhibition protects against AP by inhibiting NF-κB and p38 pathways, and indicated that PAK1 is a potential therapy to alleviate AP patients in clinic, and these need to be explored further.

Topics: Acute Disease; Animals; Antineoplastic Agents; Ceruletide; Cytokines; Disease Models, Animal; Humans; Mice, Inbred C57BL; NF-kappa B; p21-Activated Kinases; p38 Mitogen-Activated Protein Kinases; Pancreatitis; Pyridones; Pyrimidines; RNA Interference; Signal Transduction

Heme oxygenase-1 (HO-1) has an anti-inflammatory action in acute pancreatitis (AP). However, its mechanism of action and natural compounds/drugs to induce HO-1 in pancreas are not well understood. In this study, we investigated the regulatory mechanisms of HO-1 during AP using desoxo-narchinol-A (DN), the natural compound inducing HO-1 in the pancreas. Female C57/BL6 Mice were intraperitoneally injected with supramaximal concentrations of cerulein (50 μg/kg) hourly for 6 h to induce AP. DMSO or DN was administered intraperitoneally, then mice were sacrificed 6 h after the final cerulein injection. Administration of DN increased pancreatic HO-1 expression through activation of activating protein-1, mediated by mitogen-activated protein kinases. Furthermore, DN treatment reduced the pancreatic weight-to-body weight ratio as well as production of digestive enzymes and pro-inflammatory cytokines. Inhibition of HO-1 by tin protoporphyrin IX abolished the protective effects of DN on pancreatic damage. Additionally, DN treatment inhibited neutrophil infiltration into the pancreas via regulation of chemokine (C-X-C motif) ligand 2 (CXCL2) by HO-1. Our results suggest that DN is an effective inducer of HO-1 in the pancreas, and that HO-1 regulates neutrophil infiltration in AP via CXCL2 inhibition.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Chemokine CXCL2; Cytokines; Disease Models, Animal; Female; Heme Oxygenase-1; Humans; Inflammation Mediators; Mice; Mice, Inbred C57BL; Naphthols; Neutrophil Infiltration; Neutrophils; Pancreas; Pancreatitis

Acute pancreatitis (AP) is an exocrine dysfunction of the pancreas where oxidative stress and inflammatory cytokines play a key role in induction and progression of the disease. Studies have demonstrated that antioxidant phytochemicals have been effective in improving pancreatitis condition, but there are no clinically approved drugs till date. Our study aims to assess the preventive activity of visnagin, a novel phytochemical isolated from Ammi visnaga against cerulein induced AP. Male Swiss albino mice were divided into six groups (n = 6, each group) comprising of normal control, cerulein control, seven day pre-treatment with visnagin at three dose levels; visnagin low dose (10 mg/kg), visnagin mid dose (30 mg/kg), visnagin high dose (60 mg/kg) and visnagin control (60 mg/kg). AP was induced by six injections of cerulein (50 μg/kg, i.p.) on the 7

Topics: Acute Disease; Ammi; Amylases; Animals; Anti-Inflammatory Agents; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Khellin; Lipase; Male; Mice; Multiple Organ Failure; NF-E2-Related Factor 2; NF-kappa B; Pancreatitis; Signal Transduction

Oxidative stress plays a major role in acute pancreatitis (AP), leading to massive macrophage infiltration. Nanoyttria (NY) possesses potent free radical scavenging activity. As reactive oxygen species and inflammation play major role in AP, we hypothesized that NY may alleviate cerulein induced AP. NY ameliorated LPS induced oxidative stress in vitro. It reduced ROS, superoxide radical generation and restored the mitochondrial membrane potential in macrophages. Interestingly, NY reduced plasma amylase and lipase levels and attenuated the mitochondrial stress and inflammatory markers. NY suppressed the recruitment of inflammatory cells around the damaged pancreatic acinar cells. Furthermore, NY intervention perturbed the course of AP via reduction of endoplasmic reticulum (ER) stress markers (BiP, IRE1 and Ero1-Lα), and molecular chaperones (Hsp27 and Hsp70). We, to the best of our knowledge, report for first time that NY can attenuate experimental AP by restoration of mitochondrial and ER homeostasis through Nrf2/NFκB pathway modulation.

Topics: Acute Disease; Animals; Antioxidants; Biomarkers; Ceruletide; Endoplasmic Reticulum Stress; Inflammation; Lipid Peroxidation; Lipopolysaccharides; Macrophages; Male; Membrane Potential, Mitochondrial; Mice; Nanoparticles; Neutrophils; Nitrosation; Oxidation-Reduction; Oxidative Stress; Pancreas; Pancreatitis; RAW 264.7 Cells; Reactive Oxygen Species; Severity of Illness Index; Transcription Factor RelA; Yttrium

Acute pancreatitis (AP) is a multifactorial disease characterized by necroinflammatory changes of the pancreas. Our study is the first study which evaluated the relationship between the free radical production, enzymatic and nonenzymatic antioxidants, oxidative damage, and secretory function of the salivary glands of AP rats. Male Wistar rats were divided equally into 2 groups: control (

Topics: Acute Disease; Animals; Antioxidants; Ceruletide; Male; Oxidative Stress; Pancreatitis; Rats; Rats, Wistar; Salivary Glands

Endoplasmic reticulum (ER) stress in the pancreas is closely associated with the development of acute pancreatitis. However, the role of the protein kinase RNA-like ER kinase (PERK) in this disease is not fully understood. We investigated whether an inhibitor of the dephosphorylation of eukaryotic initiation factor 2α, salubrinal, could improve murine experimental pancreatitis through the amelioration of ER stress.. Acute pancreatitis was induced by the intraperitoneal administration of cerulein (50 μg/kg) six times at 1-h intervals followed by lipopolysaccharide (10 mg/kg). Salubrinal was administered intraperitoneally immediately after lipopolysaccharide injection and 3 h later. Mice were sacrificed 24 h after the first injection of cerulein, and serum amylase and proinflammatory cytokines were measured. The severity of pancreatitis was evaluated histologically using a scoring system. The expression levels of ER stress-related proteins were evaluated by Western blotting.. The administration of salubrinal significantly attenuated the increase in serum amylase levels and improved histologically assessed pancreatitis. The serum levels of proinflammatory cytokines were significantly suppressed in salubrinal-treated mice, as was the expression of glucose-regulated protein 78, CCAAT/enhancer-binding protein homologous protein, and cleaved caspase-3.. The amelioration of ER stress through augmentation of the PERK-signaling pathway may be a therapeutic target for the treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Ceruletide; Cinnamates; Cytokines; Endoplasmic Reticulum Stress; Eukaryotic Initiation Factor-2; Injections, Intraperitoneal; Lipopolysaccharides; Male; Mice; Mice, Inbred C57BL; Pancreatitis; Phosphorylation; Thiourea

The purpose of the present study was to investigate the effect of carbon monoxide (CO) released from CO‑releasing molecule 2 (CORM‑2) on mice with acute pancreatitis (AP). To perform the investigation, a mouse AP model was established using caerulein. The mice were treated with or without CORM‑2. The survival rate of the mice in the different groups was analyzed, and serum amylase and lipase levels were measured to assess the degree of pancreatic injury. The severity of AP was also evaluated by histological examination, and histopathological scoring of the pancreatic damage was performed. Pancreatic cell apoptosis was analyzed using a terminal deoxynucleotidyl‑transferase‑mediated dUTP nick end labelling assay. The function of the lung and liver was also assessed in the present study. Furthermore, the role of CORM‑2 on oxidative stress, intercellular adhesion molecule 1 (ICAM‑1) and vascular cell adhesion molecule 1 (VCAM‑1) expression, pro‑inflammatory cytokine production, and nuclear factor (NF)‑κB activation in the pancreas of AP mice was determined. The results demonstrated that CORM‑2 reduced the mortality, pancreatic damage, and lung and liver injury of AP mice. CORM‑2 administration also reduced systemic and localized inflammatory cell factors. Furthermore, treatment with CORM‑2 inhibited the expression of ICAM‑1 and VCAM‑1, and the activation of NF‑κB and phosphorylated inhibitor of NF‑κB subunit α, in the pancreas of AP mice. These results indicated that CO released from CORM‑2 exerted protective effects on AP mice, and the beneficial effects were likely due to inhibition of NF‑κB pathway activation.

Topics: Acute Disease; Amylases; Animals; Carbon Monoxide; Ceruletide; Cytokines; Disease Models, Animal; Intercellular Adhesion Molecule-1; Lipase; Lung; Male; Malondialdehyde; Mice; Mice, Inbred C57BL; NF-kappa B; Organometallic Compounds; Oxidative Stress; Pancreas; Pancreatitis; Peroxidase

To investigate the therapeutic effects of resolvin D1 (RvD1) on cerulein and lipopolysaccharide (LPS)-induced severe acute pancreatitis in mice.. The model of severe acute pancreatitis was induced by cerulein combined with LPS in mice. Mice were treated with RvD1 at a dose of 150 mg/kg for 4 h after the last injection of cerulein. Pathological changes and scores were assessed by HE staining, serum amylase and lipase levels were detected by ELISA, serum tumor necrosis factor-α(TNF-α) and interleukin-6 (IL-6) levels were determined by Luminex Assay.. Cerulein combined with LPS successfully induced severe acute pancreatitis model in mice. RvD1 reduced the pathological changes of pancreas in severe acute pancreatitis mice, decreased the serum levels of amylase and lipase, as well as attenuated the serum levels of TNF-α and IL-6.. RvD1 can reduce the severity of severe acute pancreatitis induced by cerulein and LPS in mice.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Docosahexaenoic Acids; Interleukin-6; Lipase; Lipopolysaccharides; Mice; Pancreas; Pancreatitis; Tumor Necrosis Factor-alpha

Acute pancreatitis (AP) is a type of sterile inflammation of the pancreas, potentially leading to systemic inflammatory response syndrome or multiple organ failure. An emerging evidence that dysfunction of miRNA expression may alter pivotal physiological functions and lead to inflammation infiltration and complication of multiple diseases, including AP. Here, the AP model was successfully replicated using cerulein in vitro and in vivo. RT-qPCR was used to detect low expression of miR-148a in AP. This study verified that IL-6 was a direct target of miR-148a. Over-expression of miR-148a decreased the mRNA and protein levels of IL-6 by RT-qPCR and Elisa. Moreover, over-expression of miR-148a improved the pathological state of AP through H&E and MPO staining and transmission electron microscopy. After over-expressing miR-148a, Western blot and immunohistochemical method were used to confirm the reduction of autophagosomes and autolysosomes, blockade of the levels of p-STAT3, LC3-II, ATG7, ATG4c, Beclin1 and the increased p62 expression in AP. The expression of LAMP-2 was not significantly different. In addition, IL-6 and AG490, the IL-6/STAT3 signaling inhibitor, were used to verify the role of IL-6/STAT3 signaling in the regulation of miR-148a on autophagy in cerulein-induced AP in vitro and in vivo. Taken together, our findings indicate that miR-148a suppresses autophagy via regulating IL-6/STAT3 signaling in cerulein-induced AP in vitro and in vivo. The miR-148a appears to be a promising candidate for the gene therapy of AP.

Topics: Acute Disease; Animals; Autophagy; Cell Line; Ceruletide; Down-Regulation; Gene Expression Regulation; Genetic Therapy; Genetic Vectors; Interleukin-6; Male; Mice; Mice, Inbred BALB C; MicroRNAs; Pancreatitis; Rats; Signal Transduction; STAT3 Transcription Factor

BACKGROUND Acute pancreatitis is an inflammatory disease of the pancreas associated with high patient morbidity. Lycium barbarum polysaccharide (LBP), a traditional Chinese medicine with an active component extracted from the goji berry, has previously been reported to have anti-inflammatory effects. This study aimed to investigate the effects of LBP in a mouse model of cerulein-induced acute pancreatitis. MATERIAL AND METHODS Acute pancreatitis was induced by intraperitoneal injection of cerulein in C57BL/6 wild-type mice or nuclear factor erythroid-2-related factor 2 (NRF2) gene knockout mice. LBP or normal saline was administrated by gavage once daily for one week before the induction of acute pancreatitis. At 12 hours after the first intraperitoneal injection of cerulein, the mice were euthanized. Blood and pancreatic tissue were sampled for histology and for the measurement of pro-inflammatory cytokines, serum amylase, and lipase. RESULTS In the untreated mouse model of cerulein-induced acute pancreatitis, amylase and lipase levels were increased, and these levels were reduced by LBP treatment when compared with vehicle treatment. In the untreated mouse model, histology of the pancreas showed edema and inflammation, which were reduced in the LBP-treated mice. In the untreated mouse model, increased levels of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) were found, which were reduced in the LBP-treated mice. NRF2 gene knockout mice with cerulein-induced acute pancreatitis showed reduced anti-inflammatory effects of LBP treatment. LBP increased the expression of NRF2 and heme oxygenase-1 (HO-1). CONCLUSIONS In a mouse model of cerulein-induced acute pancreatitis, LBP reduced inflammation by upregulating NRF2 and HO-1.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cytokines; Disease Models, Animal; Drugs, Chinese Herbal; Heme Oxygenase-1; Interleukin-6; Lipase; Medicine, Chinese Traditional; Mice; Mice, Inbred C57BL; NF-E2-Related Factor 2; NF-kappa B; Pancreas; Pancreatitis; Tumor Necrosis Factor-alpha

Acute pancreatitis (AP) is a common acute abdominal condition, frequently associated with intestinal barrier dysfunction, which aggravates AP retroactively. Butyrate exhibits anti-inflammatory effects in a variety of inflammatory diseases. However, its potential beneficial effect on AP and the underlying mechanisms have not been investigated.. Experimental AP was induced by caerulein hyperstimulation in wild-type and GPR109A. Butyrate prophylaxis attenuated AP as shown by reduced serum amylase and lipase levels, pancreatic oedema, myeloperoxidase activity, and improved pancreatic morphology. Amelioration of pancreatic damage by butyrate was associated with reduced levels of TNF-α, IL-6, and CCL2 and suppressed activation of the NLRP3 inflammasome in both pancreas and colon. Further, butyrate ameliorated pancreatic inflammation by suppressing interactions between histone deacetylase 1 (HDAC1) and AP1 and STAT1 with increased histone acetylation at H3K9, H3K14, H3K18, and H3K27 loci, resulting in suppression of NLRP3 inflammasome activation and modulation of immune cell infiltration in pancreas. Additionally, butyrate mediated STAT1/AP1-NLRP3 inflammasome suppression via HDAC1 inhibition was demonstrated in peritoneal macrophage. In colon, butyrate inhibited NLRP3 inflammasome activation via GPR109A. Accordingly, the modulatory effects of butyrate on AP, AP-associated gut dysfunction, and NLRP3 inflammasome activation were diminished in GPR109A. Our study dissected tissue-specific anti-inflammatory mechanisms of butyrate during AP, suggesting that increased colonic levels of butyrate may be a strategy to protect against AP.

Topics: Acute Disease; Administration, Oral; Animals; Anti-Inflammatory Agents, Non-Steroidal; Butyrates; Ceruletide; Female; Intestinal Diseases; Intestine, Small; Macrophages; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis

Opioids such as morphine are widely used for the management of pain associated with acute pancreatitis. Interestingly, opioids are also known to affect the immune system and modulate inflammatory pathways in non-pancreatic diseases. However, the impact of morphine on the progression of acute pancreatitis has never been evaluated. In the current study, we evaluated the impact of morphine on the progression and severity of acute pancreatitis.. Effect of morphine treatment on acute pancreatitis in caerulein, L-arginine and ethanol-palmitoleic acid models was evaluated after induction of the disease. Inflammatory response, gut permeability and bacterial translocation were compared. Experiments were repeated in mu (µ) opioid receptor knockout mice (MORKO) and in wild-type mice in the presence of opioid receptor antagonist naltrexone to evaluate the role of µ-opioid receptors in morphine's effect on acute pancreatitis. Effect of morphine treatment on pathways activated during pancreatic regeneration like sonic Hedgehog and activation of embryonic transcription factors like pdx-1 and ptf-1 were measured by immunofluorescence and quantitative PCR.. Histological data show that treatment with morphine after induction of acute pancreatitis exacerbates the disease with increased pancreatic neutrophilic infiltration and necrosis in all three models of acute pancreatitis. Morphine also exacerbated acute pancreatitis-induced gut permeabilisation and bacteraemia. These effects were antagonised in the MORKO mice or in the presence of naltrexone suggesting that morphine's effect on severity of acute pancreatitis are mediated through the µ-opioid receptors. Morphine treatment delayed macrophage infiltration, sonic Hedgehog pathway activation and expression of pdx-1 and ptf-1.. Morphine treatment worsens the severity of acute pancreatitis and delays resolution and regeneration. Considering our results, the safety of morphine for analgesia during acute pancreatitis should be re-evaluated in future human studies.

Topics: Acute Disease; Analgesics, Opioid; Animals; Arginine; Ceruletide; Disease Models, Animal; Disease Progression; Fatty Acids, Monounsaturated; Mice; Mice, Knockout; Morphine; Pancreas; Pancreatitis; Severity of Illness Index; Time Factors

Little is known about the signaling pathways that initiate and promote acute pancreatitis (AP). The pathogenesis of AP has been associated with abnormal increases in cytosolic Ca. Pancreatitis was induced in C57BL/6J mice (control) and mice deficient in peptidylprolyl isomerase D (cyclophilin D, encoded by Ppid) by administration of L-arginine (also in rats), caerulein, bile acid, or an AP-inducing diet. Parameters of pancreatitis, mitochondrial function, autophagy, ER stress, and lipid metabolism were measured in pancreatic tissue, acinar cells, and isolated mitochondria. Some mice with AP were given trehalose to enhance autophagic efficiency. Human pancreatitis tissues were analyzed by immunofluorescence.. Mitochondrial dysfunction in pancreas of mice with AP was induced by either mitochondrial Ca. In different animal models, we find a central role for mitochondrial dysfunction, and for impaired autophagy as its principal downstream effector, in development of AP. In particular, the pathway involving enhanced interaction of cyclophilin D with ATP synthase mediates L-arginine-induced pancreatitis, a model of severe AP the pathogenesis of which has remained unknown. Strategies to restore mitochondrial and/or autophagic function might be developed for treatment of AP.

Topics: Acute Disease; Animals; Arginine; Autophagy; Bile Acids and Salts; Calcium Signaling; Ceruletide; Choline Deficiency; Cyclophilins; Disease Models, Animal; Endoplasmic Reticulum Stress; Ethionine; Genetic Predisposition to Disease; Humans; Lipid Metabolism; Membrane Potential, Mitochondrial; Mice, Inbred C57BL; Mice, Knockout; Mitochondria; Mitochondrial Proton-Translocating ATPases; Pancreas; Pancreatitis; Peptidyl-Prolyl Isomerase F; Phenotype; Rats; Time Factors; Trehalose

Mitochondrial permeability transition pore inhibition is a promising approach to treat acute pancreatitis (AP). We sought to determine (i) the effects of the mitochondrial permeability transition pore inhibitor 3,5-seco-4-nor-cholestan-5-one oxime-3-ol (TRO40303) on murine and human pancreatic acinar cell (PAC) injury induced by fatty acid ethyl esters (FAEEs) or taurolithocholic acid-3-sulfate and (ii) TRO40303 pharmacokinetics and efficacy in experimental alcoholic AP (FAEE-AP).. Changes in mitochondrial membrane potential (Δψm), cytosolic Ca ([Ca]c), and cell fate were examined in freshly isolated murine or human PACs by confocal microscopy. TRO40303 pharmacokinetics were assessed in cerulein-induced AP and therapeutic efficacy in FAEE-AP induced with palmitoleic acid and ethanol. Severity of AP was assessed by standard biomarkers and blinded histopathology.. TRO40303 prevented loss of Δψm and necrosis induced by 100 μM palmitoleic acid ethyl ester or 500 μM taurolithocholic acid-3-sulfate in murine and human PACs. Pharmacokinetic analysis found TRO40303 accumulated in the pancreas. A single dose of 3 mg/kg TRO40303 significantly reduced serum amylase (P = 0.043), pancreatic trypsin (P = 0.018), and histopathology scores (P = 0.0058) in FAEE-AP.. TRO40303 protects mitochondria and prevents necrotic cell death pathway activation in murine and human PACs, ameliorates the severity of FAEE-AP, and is a candidate drug for human AP.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Esters; Fatty Acids; Humans; Membrane Potential, Mitochondrial; Mice, Inbred C57BL; Mitochondria; Necrosis; Oximes; Pancreatitis; Pancreatitis, Alcoholic; Secosteroids; Taurolithocholic Acid

A various of pharmacological effects of astaxanthin has been confirmed. However, the mechanism underlying protective effect of astaxanthin on acute pancreatitis (AP) induced by cerulein still unclear. The present study is to investigate the mechanism underlying the effect of astaxanthin on autophagy and apoptosis via the JAK/STAT3 pathway.. Intraperitoneal injection of cerulein at hourly intervals followed by lipopolysaccharide injection were used in Balb/C mice. Vehicle or astaxanthin, which intraperitoneal injected in two doses (20 mg/kg and 40 mg/kg), were injected in mice 1 h before the first cerulein injection. At 3 h after the last injection, when the pathological changes were most severe, pancreatic tissue was analyzed by pathologically scored and hematoxylin and eosin (H&E) staining. The severity of AP was assessed by histological grading, proinflammatory cytokine levels, biochemistry, myeloperoxidase (MPO) activity, and analysis of JAK/STAT3 activity.. Astaxanthin administration markedly reduced serum digestive enzyme activities, pancreatic histological scores, proinflammatory cytokine levels (tumor necrosis factor-α (TNF-α), Interleukin-1β (IL-1β), and Interleukin-6 (IL-6)), MPO and JAK/STAT3 activity.. Collectively, these results indicate that astaxanthin inhibits pancreatic injury in AP by targeting JAK/STAT3-mediated apoptosis and autophagy.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Apoptosis; Autophagy; Ceruletide; Cytokines; Disease Models, Animal; Humans; Inflammation Mediators; Janus Kinases; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Pancreas; Pancreatitis; Peroxidase; STAT3 Transcription Factor; Xanthophylls

We investigated whether intrapancreatic coagulation, with deposition of the fibrinogen-γ dimer (Fib-γD) and hypoxia, affect the severity of acute pancreatitis (AP) in mice. Pancreata of mice with AP induced by administration of cerulein or by L-arginine, or from patients with pancreatitis, had increased deposition of Fib-γD compared with control pancreata. Heparin administration protected mice from cerulein-induced AP and prevented Fib-γD formation. Cerulein administration resulted in activation and stabilization of hypoxia-inducible factor-1α (HIF1α) in pancreata of oxygen-dependent degradation domain-luciferase HIF1α reporter mice. Cerulein also led to induction of genes regulated by HIF1α, including Vegfa and Ero1a, before evidence of Fib-γD deposition or histologic features of AP. Expression of tissue factor, which is regulated by vascular endothelial growth factor, also increased following cerulein administration. Mice with acinar cell-specific disruption of Hif1a (Hif1a

Topics: Acinar Cells; Acute Disease; Animals; Arginine; Ceruletide; Disease Models, Animal; Homeostasis; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Mice; Pancreas; Pancreas, Exocrine; Pancreatitis; Vascular Endothelial Growth Factor A

Acute pancreatitis (AP) is characterized by severe inflammation and acinar cell death. Transmembrane protein 173 (TMEM173 or STING) is a DNA sensor adaptor protein on immune cells that recognizes cytosolic nucleic acids and transmits signals that activate production of interferons and the innate immune response. We investigated whether leukocyte STING signaling mediates inflammation in mice with AP.. We induced AP in C57BL/6J mice (control) and C57BL/6J-Tmem173gt/J mice (STING-knockout mice) by injection of cerulein or placement on choline-deficient DL-ethionine supplemented diet. In some mice, STING signaling was induced by administration of a pharmacologic agonist. AP was also induced in C57BL/6J mice with bone marrow transplants from control or STING-knockout mice and in mice with disruption of the cyclic GMP-AMP synthase (Cgas) gene. Pancreata were collected, analyzed by histology, and acini were isolated and analyzed by flow cytometry, quantitative polymerase chain reaction, immunoblots, and enzyme-linked immunosorbent assay. Bone-marrow-derived macrophages were collected from mice and tested for their ability to detect DNA from dying acinar cells in the presence and absence of deoxyribonuclease (DNaseI).. STING signaling was activated in pancreata from mice with AP but not mice without AP. STING-knockout mice developed less severe AP (less edema, inflammation, and markers of pancreatic injury) than control mice, whereas mice given a STING agonist developed more severe AP than controls. In immune cells collected from pancreata, STING was expressed predominantly in macrophages. Levels of cGAS were increased in mice with vs without AP, and cGAS-knockout mice had decreased edema, inflammation, and other markers of pancreatic injury upon induction of AP than control mice. Wild-type mice given bone marrow transplants from STING-knockout mice had less pancreatic injury and lower serum levels of lipase and pancreatic trypsin activity following induction of AP than mice given wild-type bone marrow. DNA from dying acinar cells activated STING signaling in macrophages, which was inhibited by addition of DNaseI.. In mice with AP, STING senses acinar cell death (by detecting DNA from dying acinar cells) and activates a signaling pathway that promotes inflammation. Macrophages express STING and activate pancreatic inflammation in AP.

Topics: Acinar Cells; Acute Disease; Animals; Cell Death; Ceruletide; Disease Models, Animal; Inflammation; Macrophages; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Nucleotides, Cyclic; Pancreas; Pancreatitis; Signal Transduction

Changes in the protein expression occurring within the initiation phase of acute pancreatitis (AP) might be vital in the development of this complex disease. However, the exact mechanisms involved in the onset of AP remains elusive and most of our knowledge about the pathobiology of AP comes from animal models. We performed in a rat pancreatitic model a high-throughput shotgun proteomic profiling of the soluble and whole membrane fractions from the pancreas during the early phase of cerulein (Cer)-induced AP. We identified 997 proteins, of which 353 were significantly different (22, 276 or 55 in both, the soluble or the membrane fractions, respectively). Gene Ontology and KEGG PATHWAY analyses revealed that these proteins were implicated in molecular mechanisms relevant to AP pathogenesis, including vesicle-mediated and protein transport, lysosomal and mitochondrial impairment or proteolysis. Numerous metabolic processes were downregulated apparently to reduce energy consumption, and a remarkable increase in inflammatory and stress responses was also highlighted. The proteomic data were verified by immunoblotting of 11 and 7 different soluble or membrane-associated proteins, either novel (VPS29 and MCTS1) or known factors in AP. Also, our first observation of the imbalance of some COP proteins during AP early phase deserves further characterization.. AP is one of the most important pathological inflammatory states of the exocrine pancreas but its pathophysiology remains incompletely understood, especially the early acinar events. Proteomic analysis of pancreatic subcellular fractions simplifies protein maps and helps in the identification of new protein alterations and biomarkers characterizing pancreatic tissue damage. Our shotgun approach has not been previously used to profile the early proteomic alterations of the disease, which are considered crucial for its development and for the founding of clinical procedures. Furthermore, our subcellular fractionation protocol allowed us to detect changes in membrane proteins so far overlooked in the proteomic study of AP. Accordingly, using TMT proteomics and bioinformatic tools, we were able to detect significant changes in protein expression related to many pathobiological pathways of acute pancreatitis as from the early phase of the disease. To our knowledge, some of these changes, such as the imbalance of some COP proteins, have never been described in this disease.

Topics: Acute Disease; Animals; Ceruletide; Lysosomes; Male; Mitochondria; Mitochondrial Proteins; Pancreatitis; Proteome; Proteomics; Rats; Rats, Wistar

Acute pancreatitis (AP) is one common clinical acute abdominal disease, for which specific pharmacological or nutritional therapies remain elusive. Lactose, a macronutrient and an inducer of host innate immune responses, possesses immune modulatory functions. The current study aimed to investigate potential modulatory effects of lactose and the interplay between the nutrient and pancreatic immunity during experimentally induced AP in mice. We found that either prophylactic or therapeutic treatment of lactose time-dependently reduced the severity of AP, as evidenced by reduced pancreatic edema, serum amylase levels, and pancreatic myeloperoxidase activities, as well as by histological examination of pancreatic damage. Overall, lactose promoted a regulatory cytokine milieu in the pancreas and reduced infiltration of inflammatory neutrophils and macrophages. On acinar cells, lactose was able to suppress caerulein-induced inflammatory signaling pathways and to suppress chemoattractant tumor necrosis factor (TNF)-α and monocyte chemotactic protein-1 production. Additionally, lactose acted on pancreas-infiltrated macrophages, increasing interleukin-10 and decreasing tumor necrosis factor alpha production. Notably, lactose treatment reversed AP-associated infiltration of activated neutrophils. Last, the effect of lactose on neutrophil infiltration was mimicked by a galectin-3 antagonist, suggesting a potential endogenous target of lactose. Together, the current study demonstrates an immune regulatory effect of lactose to alleviate AP and suggests its potential as a convenient, value-added therapeutic macronutrient to control AP, and lower the risk of its systemic complications.

Topics: Acute Disease; Animals; Ceruletide; Cytokines; Female; Immunologic Factors; Lactose; Macrophages; Mice, Inbred BALB C; Neutrophil Infiltration; Neutrophils; Pancreas; Pancreatitis; Phenotype

BACKGROUND Steroid receptor coactivator-interacting protein (SIP) inhibits the activation of nuclear factor-kappa B (NF-κB) by interacting with p65. The occurrence of acute pancreatitis (AP) is closely associated with pro-inflammatory response. The present study aimed to investigate the role of SIP on myocardial injury caused by AP. MATERIAL AND METHODS Rat pancreatic acinar tumor cell line AR42J cells were treated with caerulein to establish AP cell models. The levels of TNF-α, IL-6, cTnI, CK-MB, and LDH1 were detected by ELISA assay. The mRNA and protein expression levels of SIP, p-p65, and p65 were detected by qRT-PCR and western blot analysis, respectively. Next, the AP cell models were non-transfected or transfected with SIP plasmids or SIP siRNA. ELISA assay was also performed to test the levels of TNF-α, IL-6, cTnI, CK-MB, and LDH1. Moreover, qRT-PCR and western blot analysis were performed to measure the mRNA and protein expression levels of SIP, p-p65, and p65, respectively. RESULTS Caerulein upregulated the levels of TNF-α, IL-6, cTnI, CK-MB, and LDH1. These upregulations were reduced by SIP plasmids and promoted by SIP siRNA, respectively. Caerulein also increased the mRNA and protein expression levels of p-p65. However, the increases were attenuated by SIP plasmids and enhanced by SIP siRNA, respectively. CONCLUSIONS In conclusion, the results suggested that SIP may inhibit the inflammatory response by deactivating p65, thus reducing the myocardial damage caused by AP.

Topics: Acute Disease; Animals; Cell Line, Tumor; Ceruletide; Intracellular Signaling Peptides and Proteins; Models, Biological; Myocardium; Pancreatitis; Phosphorylation; Plasmids; Rats; RNA, Small Interfering; Transcription Factor RelA

Naringenin (Nar) is a type of flavonoid and has been shown to have anti-inflammatory and antioxidative properties. However, the effects of Nar on acute pancreatitis (AP) have not been well studied. In this study, we aimed to investigate the function of Nar in a mouse model of AP.. Mild acute pancreatitis (MAP) was induced by caerulein (Cae), and severe acute pancreatitis (SAP) was induced by L-arginine in mice. Nar was administered intraperitoneally at doses of 25, 50, or 100 mg/kg following MAP induction and at a dose of 100 mg/kg following SAP induction. The serum levels of cytokines, lipase, and amylase were determined, and pancreatic and pulmonary tissues were harvested.. The serum levels of amylase, lipase, and cytokines were significantly decreased in both MAP and SAP models after Nar treatment. The malondialdehyde (MDA) levels of the pancreatic tissue was significantly reduced in both MAP and SAP after Nar treatment. In contrast, glutathione peroxidase (GPx), glutathione reductase (GR), glutathione S-transferase (GST), total sulfhydryl (T-SH), and non-proteinsulthydryl (NP-SH) were markedly increased in both MAP and SAP after Nar treatment. The injury in pancreatic and pulmonary tissues was markedly improved as evidenced by the inhibited expression of myeloperoxidase, nod-like receptor protein 3, and interleukin 1 beta as well as the enhanced expression of nuclear factor erythroid 2-related factor 2/heme oxygenase-1 in pancreatic tissues.. Nar exerted protective effects on Cae-induced MAP and L-arginine-induced SAP in mice, suggesting that Nar may be a potential therapeutic intervention for AP.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Cytokines; Flavanones; Glutathione Peroxidase; Glutathione Reductase; Glutathione Transferase; Heme Oxygenase-1; Lipase; Male; Membrane Proteins; Mice; NF-E2-Related Factor 2; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreatitis

Acute pancreatitis (AP) is a common acute abdominal disease accompanied by systemic inflammatory response syndrome, and could even be complicated by multiple-organ damage. This study aimed to examine whether calycosin, an isoflavone isolated from Radix astragali with antioxidant and anti-inflammatory activity, could protect against AP induced by cerulein. To this end, Balb/C mice were injected with cerulein (50 μg/kg) to establish the animal model of AP. Calycosin (25 and 50 mg/kg, p.o.) was administered 1 h prior to the first cerulein injection. After the last injection of cerulein, the mice were sacrificed and blood was obtained for cytokine analysis. The pancreas was removed for morphological examination, myeloperoxidase (MPO) and malondialdehyde (MDA) analyses, immunohistochemistry, and western blot analysis. Calycosin treatment reversed the increased serum levels of amylase and lipase, alleviated the pathological damage in the pancreas, and decreased the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β in mice with AP. Additionally, calycosin significantly reduced cerulein-induced pancreatic edema, inhibited MPO activity and increased superoxide dismutase (SOD) activity, and inhibited the expression of NF-κB/p65 and phosphorylation of the inhibitor of NF-κB (IκBα) and p38 MAPK. These results suggested that calycosin protects against AP by exerting anti-inflammatory and anti-oxidative stress effects via the p38 MAPK and NF-κB signal pathways. Calycosin's benefits for AP patients need to be explored further.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Astragalus propinquus; Ceruletide; Cytokines; Disease Models, Animal; Drugs, Chinese Herbal; Isoflavones; Male; Mice, Inbred BALB C; NF-kappa B; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Pancreatitis; Signal Transduction

The objective of this study is to explore the effect of melatonin on endoplasmic reticulum stress in acute pancreatitis (AP) and the molecular mechanism.. Acute pancreatitis was induced in vivo in Sprague-Dawley rats by the retrograde injection of 5% taurocholate into the biliopancreatic duct and in vitro by treating AR42J cells with cerulein (10 nmol/L) plus lipopolysaccharide (LPS) (10 mg/L). The rats and cells were treated with melatonin (50 mg/kg in rats and 0.5, 1, and 2 mmol/L in AR42J cells) 30 minutes before AP was induced. After 9 hours, the cells and rat pancreas tissue were collected for Western blot, reverse transcription polymerase chain reaction, histological examination, immunohistochemistry, and immunofluorescence analysis.. Inositol-requiring 1α (IRE1α)-mediated Jun N-terminal kinase (JNK)/nuclear factor-kappa B (NF-κB) pathway were activated early in AR42J cells and rat AP models. Melatonin significantly inhibited the expression of proinflammatory cytokines. Western blot and immunohistochemical results all indicated that melatonin regulated apoptosis-related protein expression. In addition, melatonin treatment resulted in significantly reduced pancreatic tissue injury, as revealed by histological changes and pathological scores. Furthermore, melatonin treatment significantly reduced the activation of IRE1α-mediated JNK/NF-κB pathway-related proteins.. These findings suggest that melatonin protects AR42J cells and Sprague-Dawley rats against AP-associated injury, probably through downregulation of IRE1α-mediated JNK/NF-κB pathways.

Topics: Acute Disease; Animals; Antioxidants; Apoptosis; Cell Line; Ceruletide; Cytokines; Endoplasmic Reticulum Stress; Gene Expression; Lipopolysaccharides; Male; Melatonin; Pancreas; Pancreatitis; Rats, Sprague-Dawley; Signal Transduction; Taurocholic Acid

Acute pancreatitis (AP) is a common and severe gastrointestinal inflammatory disease with poorly understood pathogenesis. We adopted cerulein-induced pancreatitis, a well-established rat model shearing similarities with human AP, to determine the disease background. Special interest was placed on sphingolipids, because their signaling pathways are involved in many pathological states including hepatic steatosis, heart infarction, or pancreatic origin type 1 diabetes.. Sphingolipid levels in the blood and pancreas were determined by the means of chromatography (thin-layer and high-performance liquid chromatography).. We found that AP leads to activation of ceramide de novo synthesis pathway, as evidenced by a significant increment in sphinganine, that is, ceramide synthesis precursor, content (+3.8-fold). Surprisingly, despite the reported growth in sphinganine concentration, we observed a reduced (-38%) ceramide level in the pancreas of rats with AP. The results could be explained by subsequent hydrolysis of ceramide to other secondary messengers, that is, sphingosine (+4-fold) or sphingosine-1-phosphate (+3-fold).. Because it is known that sphingosine-1-phosphate and some of its analogs could have a protective role against AP complications, our findings may contribute to elaboration of new therapeutic strategies in the management of this severe medical condition.

Topics: Acute Disease; Animals; Ceramides; Ceruletide; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Disease Models, Animal; Humans; Pancreas; Pancreatitis; Rats, Wistar; Signal Transduction; Sphingomyelins; Sphingosine

Inflammatory response is a determinant in the pathological progression of acute pancreatitis (AP). Previous studies have shown that the activation of hedgehog (Hh) signaling is a remarkable change in caerulein-induced AP. However, the relationship between Hh signaling and inflammation is largely ambiguous.. The AP mouse model was induced by injection of cerulein, and histological staining and serum enzymology assays were used to evaluate the establishment of AP. Western blot assay was used to determine the protein levels, cleavage of apoptotic proteins, and activation of the NF-κB signaling pathway. Cytokine array was used to screen inflammatory cytokines, and target cytokines' transcriptional expression and serum levels were examined by real-time PCR and enzyme-linked immunosorbent assay, respectively.. The key transcriptional factor in Hh signaling, Gli2, was upregulated in the pancreas and other tissues during the process of AP, and it seems to be a characteristic feature of local inflammation in pancreatic tissue and systemic inflammatory response in multiple organs. The inflammatory NF-κB pathway is required for the activation of Hh signaling, as blockade of the NF-κB pathway by pyrrolidine dithiocarbamate impaired the Gli2 upregulation. Manipulation of Gli2 expression altered the activation of the NF-κB pathway correspondingly, as well as the cell apoptosis in cerulein-induced AP. Moreover, Gli2 upregulation changed the cytokine expression profile in mouse pancreatic acinar cells, mainly decreasing the pro-inflammatory cytokines interleukin (IL)-6, interferon-γ, and FasL. The anti-inflammatory cytokine IL-10 was upregulated by Gli2 overexpression. Interdiction of Gli2 by the Gli-specific inhibitor GANT61 exacerbated AP in mice and altered the balance of inflammatory cytokines.. This study indicates that Hh activation during AP development is a negative feedback of the inflammatory response, restricting inflammatory injury to the pancreas and other tissues. Thus, manipulation of Hh signaling should shed light on limiting inflammation and alleviating AP damage.

Topics: Acinar Cells; Acute Disease; Animals; Cell Line; Ceruletide; Cytokines; Disease Models, Animal; Hedgehog Proteins; Male; Mice; Mice, Inbred C57BL; NF-kappa B; Pancreatitis; Pyrrolidines; Signal Transduction; Thiocarbamates; Up-Regulation; Zinc Finger Protein Gli2

Acute pancreatitis (AP) is a serious inflammatory disorder of the pancreas with considerable mortality. The clinical therapy is hampered due to lack of any approved drug for AP. In this study, we developed curcumin (cur)-loaded poly (lactic-co-glycolic acid) cur microparticles (CuMPs) for sustained release. CuMPs were prepared by emulsion solvent evaporation method and characterized for shape, size, compatibility, and entrapment efficiency. The in vitro drug release and in vivo pharmacokinetic studies confirmed sustained release pattern of cur from CuMPs. The pharmacodynamic study was conducted in cerulein induced AP model. Prophylactic treatment was planned with single dose of CuMPs (equivalent to 7.5 mg/kg of cur) and compared with free cur given orally (100 mg/kg) and intraperitoneally (7.5 mg/kg) daily for 7 days. Interestingly, the effects of CuMPs were superior compared to the free drug administered either orally or intraperitoneally through repeated administrations. CuMPs showed significant decrease of serum amylase and lipase levels, oxidative and nitrosative stress was also significantly decreased. Moreover, CuMPs impressively decreased inflammatory cytokines. Our results may pave a way to propose similar strategy for many of promising natural products to combat several oxidative stress-mediated disorders via sustained release microparticle approaches.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Ceruletide; Curcumin; Cytokines; Delayed-Action Preparations; Drug Liberation; Male; Mice; Oxidative Stress; Pancreas; Pancreatitis; Polylactic Acid-Polyglycolic Acid Copolymer; Rats, Sprague-Dawley

Acute pancreatitis (AP) is a common inflammatory disease in gastrointestinal tract. Our previous study has shown that caerulin induces TNF receptor-associated factor 3 (TRAF3)-p38 signaling activation and pro-inflammatory response in macrophages, causing damage to co-cultured pancreatic acinar cells. Dihydromyricetin (DHM) is a flavonoid extracted from Ampelopsis grossedentata, which has displayed anti-inflammation and anti-oxidant functions. Our results here show that DHM potently inhibited caerulin-induced expression and productions of multiple pro-inflammatory cytokines (IL-1β, TNF-α and IL-17) in murine bone marrow-derived macrophages (BMDMs). DHM significantly inhibited caerulin-induced TRAF3 protein stabilization, TRAF3-mitogen-activated protein kinase kinase 3 (MKK3) association and following MKK3-p38 activation in BMDMs. Significantly, DHM was ineffective against caerulin in TRAF3-silenced BMDMs. Importantly, DHM supplement attenuated the cytotoxicity of caerulin-activated BMDMs to co-cultured pancreatic acinar cells, resulting in significantly decreased acinar cell death and apoptosis. In vivo, DHM co-administration largely attenuated pancreatic and systemic inflammation in caerulin-injected AP mice. Together, DHM inhibits caerulin-induced TRAF3-p38 signaling activation and AP response. DHM could be further studied as a potential anti-AP agent.

Topics: Acute Disease; Animals; Cell Death; Cell Survival; Cells, Cultured; Ceruletide; Cytokines; Flavonols; Macrophages; Mice; Mice, Inbred C57BL; p38 Mitogen-Activated Protein Kinases; Pancreatitis; Signal Transduction; TNF Receptor-Associated Factor 3

To date, there still is a lack of specific acute pancreatitis markers and specifically an early marker that can reliably predict disease severity. The inflammatory response in acute pancreatitis is mediated in part through oxidative stress and calcineurin-NFAT (Nuclear Factor of Activated T-cells) signaling, which is inducing its own negative regulator, regulator of calcineurin 1 (RCAN1). Caerulein induction is a commonly used in vivo model of experimental acute pancreatitis. Caerulein induces CN-NFAT signaling, reactive oxygen species and inflammation.. To screen for potential markers of acute pancreatitis, we used the caerulein model of experimental acute pancreatitis (AP) in C57Bl/6 J mice. Pancreata from treated and control mice were used for expression profiling. Promising gene candidates were validated in cell culture experiments using primary murine acinar cells and rat AR42J cells. These candidates were then further tested for their usefulness as biomarkers in mouse and human plasma.. We identified a number of novel genes, including Regulator of calcineurin 1 (Rcan1) and Sestrin 2 (Sesn2) and demonstrated that they are induced by oxidative stress, by stimulation with H. We demonstrated that Rcan1 is regulated by oxidative stress and identified RCAN1 as a potential diagnostic marker of AP.

Topics: Acute Disease; Animals; Biomarkers; Calcium-Binding Proteins; Ceruletide; Gene Expression Profiling; Gene Expression Regulation; Intracellular Signaling Peptides and Proteins; Mice; Mice, Inbred C57BL; Muscle Proteins; Oxidative Stress; Pancreatitis; RNA, Messenger

Severe acute pancreatitis is a highly lethal disease caused by systemic inflammatory response syndrome, leading to multiple organ failure. We recently showed that histidine-rich glycoprotein (HRG) supplemental therapy ameliorated septic acute respiratory distress syndrome due to unnecessary neutrophil activation and immunothrombosis formation. Here, we evaluated the effect of HRG on lung inflammation followed by pancreatitis in a severe acute pancreatitis mouse model.. Mice received intraperitoneal injections of cerulein 7 times (100 μg/kg each) at 1-hour intervals to induce acute pancreatitis. Immediately after the first cerulein injection, phosphate-buffered saline, human serum albumin (20 mg/kg), or HRG (20 mg/kg) was intravenously injected. One hour after the last cerulein injection, phosphate-buffered saline or lipopolysaccharide (5 mg/kg) was intravenously injected into the tail vein. We evaluated lung inflammatory level after pancreatitis.. We observed significantly decreased plasma HRG levels in an acute pancreatitis mouse model. Histidine-rich glycoprotein treatment inhibited lung edema and the accumulation of neutrophil in severe acute pancreatitis, but HRG did not directly affect pancreatitis. Moreover, HRG suppressed tumor necrosis factor α, inducible nitric oxide synthase, interleukin 6, and neutrophil elastase mRNA expression and myeloperoxidase activity in the lung.. These data suggested that HRG ameliorated lung inflammation secondary to pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Edema; Gene Expression; Humans; Interleukin-6; Leukocyte Elastase; Lung; Neutrophils; Nitric Oxide Synthase Type II; Pancreatitis; Proteins; Severity of Illness Index; Tumor Necrosis Factor-alpha

The proteasome is involved in the activation of NF-κB and can regulate the progression of inflammatory diseases. However, the role of proteasome in acute pancreatitis (AP) has not been demonstrated. In this study, we first observed that the protein level and activity of proteasome 20S were increased significantly in pancreatic injury tissues after caerulein-induced mild acute pancreatitis (MAP) induction, which was in consistent with the expression of the NF-κB nucleoprotein and positively correlated with the severity of AP. Then, bortezomib, a classical proteasome inhibitor, was used to intervene the progression of MAP in mice. The results showed that bortezomib administration reduced the serum amylase and lipase levels and mitigated histopathological manifestation of pancreatic injury in mice. Meanwhile, bortezomib decreased the expression of NF-κB p65 nucleoprotein as well as total proteasome 20S protein, and inhibited the activity of 20S in pancreatic tissues. In addition, we found that bortezomib could protect pancreatic acinar cell against necrosis and mitigate the severity of AP in a severe acute pancreatitis model induced by sodium taurocholate hydrate. Taken together, our study for the first time confirmed that the proteasome participated in the pathogenesis of AP and its inhibitor bortezomib could protect against AP in mice.

Topics: Acinar Cells; Acute Disease; Animals; Bortezomib; Ceruletide; Disease Progression; Male; Mice, Inbred ICR; Necrosis; Pancreas; Pancreatitis; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Protective Agents; Taurocholic Acid; Transcription Factor RelA

Acute pancreatitis is an inflammatory disorder of the pancreas that may precipitate due to various reasons such as chronic alcoholism, gall stone obstruction, and life style. Current treatment options offer limited efficacy, as they provide only symptomatic relief. This study is an attempt to study the effects of Withaferin A (WFA) against Cerulein-induced acute pancreatitis in mice. Animals were pretreated with WFA via intraperitoneal route, for 7 days. Plasma amylase and lipase, tissue malondialdehyde (MDA), and glutathione were evaluated for all groups. Western blot analysis; haematoxylin and eosin staining of the liver, lung, and pancreas; immunohistochemistry for nitrotyrosine; and myeloperoxidase activity were performed. Haematoxylin and eosin stained sections significantly revealed the altered architecture and thereby damage in the pancreas, lungs, and liver that has been low in treatment groups. Increased myeloperoxidase and nitrotyrosine have also been reduced upon treatment with WFA. Increased levels of MDA, NO, and expression of myeloperoxidase and nitrotyrosine in the parameters estimated add evidence to the role of oxidative stress and inflammation in acute pancreatitis. WFA evidently altered these conditions upon pretreatment. Our study shows that this novel steroidal compound has potent anti-inflammatory property. Natural compounds can therefore be good remedies against many diseases if incorporated in routine diet as dietary supplement.

Topics: Acute Disease; Animals; Ceruletide; Inflammation; Male; Mice; Oxidative Stress; Panax; Pancreatitis; Withania; Withanolides

Acute pancreatitis (AP), a common abdominal inflammatory disorder, is characterized by premature intracellular activation of digestive proteases within pancreatic acini and a consecutive systemic inflammatory response. Although the mechanism remains to be fully understood, inflammation is the main cause of pancreatic damage in AP. A novel compound [4-(2-acetoxy-3-((R)-3-(benzylthio)-1-methoxy-1-oxopropan-2-ylamino)-3-oxopropyl)-1,2-phenylene diacetate (DSC)], derived from danshensu, exhibits anti-inflammatory and anti-apoptotic properties

Topics: Acute Disease; Animals; Antioxidants; Biological Products; Cells, Cultured; Ceruletide; Disease Models, Animal; Female; Heme Oxygenase-1; Inflammasomes; Inflammation; Lactates; Macrophages; Mice; Mice, Inbred BALB C; Neutrophils; NF-E2-Related Factor 2; NF-kappa B; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreas; Pancreatitis; Phenylacetates; Signal Transduction; STAT3 Transcription Factor

Our previous study demonstrated that a deficiency of microRNA 21 (miR-21) protects mice from acute pancreatitis, yet the underlying molecular networks associated with miR-21 in pancreatitis and pancreatitis-associated lung injury remain unexplored.. We used next generation sequencing to analyze gene expression profiles of pancreatic tissues from wild-type (WT) and miR-21 knockout (KO) mice treated with caerulein by using a 1-day treatment protocol. The Database for Annotation, Visualization, and Integrated Discovery gene annotation tool and Ingenuity Pathway Analysis were used to analyze the molecular pathways, while quantitative real-time PCR, western blotting, and immunohistochemistry were used to explore the molecular mechanisms.. We identified 152 differentially expressed genes (DEGs) in pancreata between WT and KO mice treated with caerulein. Cellular biogenesis and metabolism were the major pathways affected between WT and KO mice, whereas cell death and inflammatory response discriminated between WT and KO mice under acute pancreatitis. We validated 16 DEGs, consisting of 6 upregulated genes and 10 downregulated genes, involved in pancreatic injury. In particular, the upregulation of Pias3 and downregulation of Hmgb1 in KO pancreata coincided with a reduced severity of pancreatitis. In addition, we found Hmgb1 stimulation resulted in the overexpression of miR-21 in peripheral blood mononuclear cells, and deletion of miR-21 led to a reduction of caerulein-induced acute pancreatitis-associated lung injury by repressing Hmgb1 expression.. Our data support the hypothesis that miR-21 modulates the inflammatory response during acute pancreatitis through the upregulation of Pias3 and downregulation of Hmgb1. Our findings further underscore a role for miR-21 in the promotion of acute pancreatitis.

Topics: Acute Disease; Animals; Cells, Cultured; Ceruletide; Female; Gene Deletion; Gene Expression Regulation; HMGB1 Protein; Mice; Mice, Inbred C57BL; Mice, Knockout; MicroRNAs; Molecular Sequence Annotation; Pancreatitis; Protein Inhibitors of Activated STAT

Pancreatic acinar cell necrosis and inflammatory responses are two key pathologic processes in acute pancreatitis (AP), which determines the severity and outcome of the disease. Recent studies suggest that necroptosis, a programed form of necrosis, is involved in the pathogenesis of AP, but the underlying mechanisms remain unknown. We investigated the expression of necrosome components, including receptor-interacting protein (RIP) 1, RIP3, and mixed lineage kinase domain-like (MLKL), and the molecular mechanisms in pancreatitis-associated necroptosis. We found that RIP3 and phosphorylated MLKL expression was positively related to the degree of necrosis, whereas RIP1 expression was negatively related to the degree of necrosis. Pharmacologic inhibition of RIP1 kinase activity exerted no protection against caerulein/cholecystokinin-8-induced AP, but knockdown of RIP1 with siRNA increased acinar cell necrosis and inhibition of NF-κB activation. RIP1 inhibition led to enhanced RIP3 expression. RIP3 and MLKL inhibition decreased acinar cell necrosis, in which the inhibition of RIP3 reduced the phosphorylation level of MLKL. RIP3 inhibition had no effect on trypsinogen activation but partly inhibited inflammasome activation. Our study strongly suggests that the imbalance between RIP1 and RIP3 shifts the cell death to necrosis, which unravels a new molecular pathogenesis of mechanism of AP and may provide insight into the development of novel therapeutic agent for other necrosis-related diseases.

Topics: Acinar Cells; Acute Disease; Animals; Apoptosis; Ceruletide; Cholecystokinin; Irritants; Male; Mice, Inbred BALB C; Mice, Inbred C57BL; Necrosis; Pancreatitis; Peptide Fragments; Phosphorylation; Protein Kinase Inhibitors; Rats, Sprague-Dawley; Receptor-Interacting Protein Serine-Threonine Kinases

Acute pancreatitis (AP) is one of the most common diseases in gastroenterology. However, neither the etiology nor the pathophysiology of the disease is fully understood and no specific or effective treatment has been developed. Heparanase is an endoglycosidase that cleaves heparan sulfate (HS) side chains of HS sulfate proteoglycans into shorter oligosaccharides, activity that is highly implicated in cellular invasion associated with cancer metastasis and inflammation. Given that AP involves a strong inflammatory aspect, we examined whether heparanase plays a role in AP. Here, we provide evidence that pancreatic heparanase expression and activity are significantly increased following cerulein treatment. Moreover, pancreas edema and inflammation, as well as the induction of cytokines and signaling molecules following cerulein treatment were attenuated markedly by heparanase inhibitors, implying that heparanase plays a significant role in AP. Notably, all the above features appear even more pronounced in transgenic mice over expressing heparanase, suggesting that these mice can be utilized as a sensitive model system to reveal the molecular mechanism by which heparanase functions in AP. Heparanase, therefore, emerges as a potential new target in AP, and heparanase inhibitors, now in phase I/II clinical trials in cancer patients, are hoped to prove beneficial also in AP.

Topics: Acute Disease; Animals; Cathepsin L; Ceruletide; Disease Models, Animal; Disease Susceptibility; Enzyme Activation; Gene Expression; Glucuronidase; Humans; Immunohistochemistry; Mice; Mice, Transgenic; Neutrophil Infiltration; Neutrophils; NF-kappa B; Pancreas; Pancreatitis; STAT3 Transcription Factor

Acute pancreatitis (AP) causes morbidity and mortality. The aim of the present study was to investigate the protective effect of tropisetron against AP induced by cerulein. Cerulein (50μg/kg, 5 doses) was used to induce AP in mice. Six hours after final cerulein injection, animals were decapitated. Hepatic/pancreatic enzymes in the serum, pancreatic content of malondialdehyde (MDA), pro-inflammatory cytokines and myeloperoxidase (MPO) activity were measured. Tropisetron significantly attenuated pancreatic injury markers and decreased the amount of elevated serum amylase, lipase, alanine aminotransferase (ALT), aspartate aminotransferase (AST), MPO activities and pro-inflammatory cytokines levels caused by AP in mice. Tropisetron didn't affect the pancreatic levels of MDA. Our results suggest that tropisetron could attenuate cerulein-induced AP by combating inflammatory signaling. Further clinical studies are needed to confirm its efficacy in patients with AP.

Topics: Acute Disease; Animals; Ceruletide; Cytokines; Disease Models, Animal; Indoles; Lipase; Male; Malondialdehyde; Mice; NF-kappa B; Pancreas; Pancreatitis; Peroxidase; Protective Agents; Signal Transduction; Tropisetron

Oxidative stress is an important regulator in the pathogenesis of acute pancreatitis (AP). Reactive oxygen species induce activation of inflammatory cascades, inflammatory cell recruitment, and tissue damage. NF-κB regulates inflammatory cytokine gene expression, which induces an acute, edematous form of pancreatitis. Protein kinase C δ (PKCδ) activates NF-κB as shown in a mouse model of cerulein-induced AP. Docosahexaenoic acid (DHA), an ω-3 fatty acid, exerts anti-inflammatory and antioxidant effects in various cells and tissues. This study investigated whether DHA inhibits cerulein-induced AP in rats by assessing pancreatic edema, myeloperoxidase activity, levels of lipid peroxide and IL-6, activation of NF-κB and PKCδ, and by histologic observation. AP was induced by intraperitoneal injection (i.p.) of cerulein (50 μg/kg) every hour for 7 h. DHA (13 mg/kg) was administered i.p. for three days before AP induction. Pretreatment with DHA reduced cerulein-induced activation of NF-κB, PKCδ, and IL-6 in pancreatic tissues of rats. DHA suppressed pancreatic edema and decreased the abundance of lipid peroxide, myeloperoxidase activity, and inflammatory cell infiltration into the pancreatic tissues of cerulein-stimulated rats. Therefore, DHA may help prevent the development of pancreatitis by suppressing the activation of NF-κB and PKCδ, expression of IL-6, and oxidative damage to the pancreas.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Disease Models, Animal; Docosahexaenoic Acids; Inflammation; Interleukin-6; Lipid Peroxides; Male; NF-kappa B; Pancreas; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species

Acute pancreatitis (AP) is a potentially life-threatening gastrointestinal disease involving intracellular activation of digestive enzymes and pancreatic acinar cell injury. The present study was performed to investigate whether methane-rich saline (MS) was involved in the regulation of AP.. MS (16ml/kg) was administered at different dosing frequencies on mice with cerulein-induced AP. Serum amylase, lipase and histopathological changes in the pancreas tissue were measured. Serum cytokine TNFα, IL-6, IFNγ and IL-10 were detected by ELISA. The mRNA levels of these inflammatory cytokines in the pancreas were detected by real time-PCR. Myeloperoxidase (MPO) and superoxide dismutase (SOD) were determined using commercial kits. Apoptosis was assessed by immunohistochemistry and Western blot.. MS treatment reversed the increased serum level of amylase and lipase, alleviated the pathological damage in the pancreas, and decreased the expression of TNFα, IL-6, IFNγ and IL-10 in cerulean-induced AP mice. In addition, MPO was down-regulated and SOD was up-regulated in the MS treated pancreas, indicating that MS had an anti-oxidant effect against AP. Furthermore, MS protected pancreatic cells against cerulean-induced apoptosis and abolished cleaved caspase-3.. MS exerted anti-inflammatory, anti-oxidant and anti-apoptotic effects on cerulein-induced AP in mice and may proved to be a promising therapeutic agent for the clinical treatment of pancreatitis.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Antioxidants; Apoptosis; Ceruletide; Cytokines; Humans; Inflammation Mediators; Lipase; Male; Methane; Mice; Mice, Inbred C57BL; Oxidative Stress; Pancreatitis; Saline Waters

To create acute pancreatitis condition experimentally in rats using cerulein, and to reveal histopathological effects in pancreatic tissue with erdosteine.. An experimental study.. Department of General Surgery, Duzce University, Turkey, from June to October 2014.. Thirty male Wistar albino rats were divided into three groups. No procedures were applied to Group 1. The rats in Group 2 and Group 3 were injected cerulein, to establish an experimental pancreatitis model and the blood amylase and lipase values were examined. The rats in Group 3 were given 10 mg/kg erdosteine. This treatment was continued for another 2 days and the rats were sacrificed. The pancreatic tissues were examined histopathologically for edema, inflammation, acinar necrosis, fat necrosis, and vacuolization.. The lipase and amylase values and the histopathological examination of pancreatic tissues evidenced that the experimental acute pancreatitis model was established and edema, inflammation, acinar necrosis, fat necrosis, and vacuolization were observed in the pancreatic tissues. The statistical results suggest that erdosteine can decrease the edema, inflammation, acinar necrosis, fat necrosis and vacuolization scores in the tissues.. The severity of acute pancreatitis, induced by cerulein in rats, is reduced with the use of erdosteine.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Edema; Expectorants; Lipase; Male; Necrosis; Pancreas; Pancreatitis; Rats; Rats, Wistar; Thioglycolates; Thiophenes; Treatment Outcome

Acute pancreatitis (AP) is a disease that can cause local and systemic complications that may have high morbidity and mortality. Currently, there is not any specific treatment for AP. In this study, we created an experimental model of AP in rats, and we aimed to demonstrate the histological effectiveness of tocilizumab treatment that antagonizes interleukin-6 (IL-6), one of the key cytokines in the development of AP.. Forty-eight rats were divided into six groups for this study. AP model was created by subcutaneous injections of cerulein (20 μg/kg) four times at 1-h intervals. Tocilizumab 4 mg/kg was administered to one of the treatment groups and 8 mg/kg to the other treatment group intraperitoneally. The effects of tocilizumab were revealed by examining pancreatic tissue of the rats histopathologically according to the Schonberg scoring system.. A comparison between tocilizumab treatment group and AP control group provides statistically significant improvement in AP (p<0.0001). Furthermore, the dose of 8 mg/kg is shown to be more effective than 4 mg/kg (p=0.004).. Our study points out that tocilizumab may be an effective agent for pancreatitis treatment.

Topics: Acute Disease; Animals; Antibodies, Monoclonal, Humanized; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Gastrointestinal Agents; Pancreas; Pancreatitis; Rats

BACKGROUND Aberrant regulation of nuclear factor-κB (NF-κB) and the signaling pathways that regulate its activity have been found to be involved in various pathologies, particularly cancers, as well as inflammatory and autoimmune diseases. Acute pancreatitis (AP) is a complex pathological process, depending on autodigestion caused by premature activation of zymogens. This study aimed to investigate the effect of high expression of TNIP2 gene on AP and AP-induced myocardial injury. MATERIAL AND METHODS To investigate the effect of TNIP2 on AP and AP-induced myocardial injury, we established an AP cell model and rat model. HE staining was applied for histological examination. ELISA was used to determine the level of pro-inflammatory cytokines (TNF-α and IL-6) and myocardial injury markers (LDH and CK-MB). QRT-PCR and Western blot analysis were performed to determine the mRNA and protein level of related genes, respectively. RESULTS We found that the protein level of TNIP2 was relatively higher in the normal AR42J cells. At 4 h after stimulating with cerulein, the protein level of TNIP2 decreased, reached a minimum at 8 h, and then gradually increased. We also found that TNIP2 was correlated with the activation of NF-κB in cerulein-stimulated AR42J cells, and TNIP2 over-expression inhibited the inflammatory response caused by cerulein. Moreover, our results suggest that TNIP2 over-expression relieved the cerulein-triggered inflammatory response and AP-induced myocardial injury in mice. CONCLUSIONS TNIP2 was shown to exert a protective effect on AP and AP-induced myocardial injury.

Topics: Acute Disease; Adaptor Proteins, Signal Transducing; Animals; Ceruletide; Cytokines; Disease Models, Animal; Female; Mice; Mice, Inbred ICR; Myocardial Infarction; NF-kappa B; Pancreatitis; RNA, Messenger; Signal Transduction; Tumor Necrosis Factor-alpha

Studies have implied the positive association of JAK2/STAT3 signaling with the onset and severity of acute pancreatitis (AP). However, definitive functional study of JAK2/STAT3 signaling in the pathogenesis of acute pancreatitis in vivo is missing and its potential as a therapeutic target and the underlying mechanisms remain to be determined.. The aim of this study was to explore the role of JAK2/STAT3 signaling in the pathogenesis of hyperlipidemia-intensified caerulin-induced AP and its potential as a therapeutic target.. Using the caerulin-induced acute pancreatitis rat model, we showed that JAK2/STAT3 signaling was activated in pancreas and systemic inflammation was increased during AP. Pharmacological suppression of JAK2 by its inhibitor AG490 robustly protected against tissue damage, attenuated JAK2/STAT3 signaling and inflammatory responses. Local pancreatic tissue damage and phosphor- JAK2 in the pancreatic tissue were enhanced in animals fed with high fat diet compared to chow-diet fed animals. Interestingly, JAK2 inhibitor AG490 significantly inhibited pancreas necrosis and systemic inflammation in animals fed with high fat or chow-diet, but did not affect STAT3 signaling.. These results establish that JAK2 activation plays a significant role in the pathogenesis of caerulin-induced AP in animals on both chow and high-fat diets by regulating necrosis and systemic inflammation. Thus, our results not only clarify novel signaling mechanisms in AP but also suggest that JAK2 might constitute a target in the management of hyperlipidemia-intensified caerulin-induced AP.

Topics: Acute Disease; Animals; Ceruletide; Dietary Fats; Hyperlipidemias; Janus Kinase 2; Male; Pancreatitis; Rats; Rats, Sprague-Dawley; Signal Transduction; STAT3 Transcription Factor; Tyrphostins

Inhibitory κB kinase (IKK)/nuclear factor κB (NF-κB) signalling has been implicated in the pathogenesis of pancreatitis, but its precise function has remained controversial. Here, we analyse the contribution of IKK/NF-κB signalling in epithelial cells to the pathogenesis of pancreatitis by targeting the IKK subunit NF-κB essential modulator (NEMO) (IKKγ), which is essential for canonical NF-κB activation.. Mice with a targeted deletion of NEMO in the pancreas were subjected to caerulein pancreatitis. Pancreata were examined at several time points and analysed for inflammation, fibrosis, cell death, cell proliferation, as well as cellular differentiation. Human samples were used to corroborate findings established in mice.. In acute pancreatitis, NEMO deletion in the pancreatic parenchyma resulted in minor changes during the early phase but led to the persistence of inflammatory and fibrotic foci in the recovery phase. In chronic pancreatitis, NEMO deletion aggravated inflammation and fibrosis, inhibited compensatory acinar cell proliferation, and enhanced acinar atrophy and acinar-ductal metaplasia. Gene expression analysis revealed sustained activation of profibrogenic genes and the CXCL12/CXCR4 axis in the absence of epithelial NEMO. In human chronic pancreatitis samples, the CXCL12/CXCR4 axis was activated as well, with CXCR4 expression correlating with the degree of fibrosis. The aggravating effects of NEMO deletion were attenuated by the administration of the CXCR4 antagonist AMD3100.. Our results suggest that NEMO in epithelial cells exerts a protective effect during pancreatitis by limiting inflammation and fibrosis and improving acinar cell regeneration. The CXCL12/CXCR4 axis is an important mediator of that effect and may also be of importance in human chronic pancreatitis.

Topics: Acute Disease; Animals; Biomarkers; Ceruletide; Chemokine CXCL12; Chronic Disease; Disease Progression; Epithelial Cells; Fibrosis; Humans; Intracellular Signaling Peptides and Proteins; Mice, Inbred C57BL; Mice, Knockout; NF-kappa B; Pancreas; Pancreatitis; Receptors, CXCR4; Regeneration

Reseda odorata L. has long been used in traditional Asian medicine for the treatment of diseases associated with oxidative injury and acute inflammation, such as endotoxemia, acute lung injury, acute myocardial infarction and hepatitis. Luteolin, the main component of Reseda odorata L., which is also widely found in many natural herbs and vege-tables, has been shown to induce heme oxygenase-1 (HO-1) expression to exert anti-inflammatory and antioxidant effects. In this study, we aimed to examine the effects of luteolin on mice with severe acute pancreatitis (SAP), and to explore the underlying mechanisms. Cerulein and lipopolysaccharide were used to induce SAP in male Institute of Cancer Research (ICR) mice in the SAP group. The SAP group was divided into 4 subgroups, as follows: the vehicle, luteolin, zinc protoporphyrin (ZnPP) only, and luteolin (Lut) + ZnPP (luteolin plus zinc protoporphyrin treatment) groups. The wet/dry weight ratios, hematoxylin and eosin staining and pathological scores of pancreatic tissues were assessed and compared to those of the control mice. Amylase, lipase, nuclear factor-κB (NF-κB) and myeloperoxidase activities, and malondialdehyde, tumor necrosis factor α (TNFα), interleukin (IL)-6, IL-10 and HO-1 levels, as well as the expression of HO-1 were determined in serum and/or pancreatic tissue samples. SAP was successfully induced in male mice compared to normal control mice. The wet/dry weight ratios, pathological scores, and amylase and lipase activity, as well as the levels of TNFα and IL-6 were significantly reduced in the pancreatic tissues of the mice in the Lut group compared with those of the mice in the vehicle group. The Lut group exhibited a significant increase in HO-1 expression in the pancreas and enhanced serum HO-1 and IL-10 levels compared with the vehicle group. The suppression of HO-1 activity in the ZnPP group significantly abolished the protective effects of luteolin. NF-κB expression in the pancreatic tissues from the mice in the Lut + ZnPP group was significantly increased following the suppression of HO-1 activity. On the whole, our findings demonstrate that luteolin protects mice from SAP by inducing HO-1-mediated anti-inflammatory and antioxidant activities, in association with the suppression of the activation of the NF-κB pathway.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Chromatography, High Pressure Liquid; Cytokines; Heme Oxygenase-1; Inflammation Mediators; Lipopolysaccharides; Luteolin; Male; Mice, Inbred ICR; Models, Biological; Necrosis; NF-kappa B; Pancreas; Pancreatitis; Protective Agents; Protoporphyrins

Detecting and monitoring exocrine pancreatic damage during nonclinical and clinical testing is challenging because classical biomarkers amylase and lipase have limited sensitivity and specificity. Novel biomarkers for drug-induced pancreatic injury are needed to improve safety assessment and reduce late-stage attrition rates. In a series of studies, miR-216a and miR-217 were evaluated as potential biomarkers of acute exocrine pancreatic toxicity in rats. Our results revealed that miR-216a and miR-217 were almost exclusively expressed in rat pancreas and that circulating miR-216a and miR-217 were significantly increased in rats following administration of established exocrine pancreatic toxicants caerulein (CL) and 1-cyano-2-hydroxy-3-butene (CHB) as well as in rats administered a proprietary molecule known to primarily affect the exocrine pancreas. Conversely, neither microRNA was increased in rats administered a proprietary molecule known to cause a lesion at the pancreatic endocrine-exocrine interface (EEI) or in rats administered an established renal toxicant. Compared with amylase and lipase, increases in miR-216a and miR-217 were of greater magnitude, persisted longer, and/or correlated better with microscopic findings within the exocrine pancreas. Our findings demonstrate that in rats, miR-216a and miR-217 are sensitive and specific biomarkers of acute exocrine pancreatic toxicity that may add value to the measurement of classical pancreatic biomarkers.

Topics: Acute Disease; Alkenes; Animals; Biomarkers; Ceruletide; Exocrine Pancreatic Insufficiency; Humans; Male; Mice, Inbred C57BL; MicroRNAs; Nitriles; Organ Specificity; Pancreas, Exocrine; Rats, Sprague-Dawley; Rats, Wistar; Sensitivity and Specificity

This study was conducted to assess the preventive/therapeutic effects of combined administration of resveratrol and guggulsterone on cerulein-induced acute pancreatitis in mice.. Acute pancreatitis was induced by intraperitoneal injection of cerulein in mice. Serum amylase assay and histology were performed to measure the severity of pancreatitis. Western blotting and multiplex cytokine/chemokine analysis were conducted to understand the action mechanisms of the reagents.. Serum amylase assay and histology revealed that the severity of acute pancreatitis was reduced by the combinatory treatment with resveratrol and guggulsterone, but the ratio of the band intensity implied that reduced nuclear factor-κB activation is primarily responsible for the effect. The reduced amounts of keratinocyte chemoattractant (chemokine [C-X-C motif] ligand 1), interferon gamma-induced protein 10 (C-X-C motif chemokine 10) and interleukin 6 expression in the sera could be involved in attenuated immune cell migration and reduced inflammation by these reagents.. Combinatory treatment with resveratrol and guggulsterone marginally reduced cerulein-induced mild acute pancreatitis in mice.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents, Non-Steroidal; Ceruletide; Chemokines; Cytokines; Female; Mice, Inbred C57BL; NF-kappa B; Pancreatitis; Pregnenediones; Resveratrol; STAT3 Transcription Factor; Stilbenes

Topics: Acute Disease; Ceruletide; Humans; Pancreas; Pancreatitis; S100A12 Protein

To detect the expression ofsignal transducer and activator of transcription 3 (STAT3) in pancreatic tissue of the mouse model of pancreatitis, and to explore its role in the evolution of acute pancreatitis.. Forty-eight healthy male balb/c mice were randomly divided into 3 groups (. Compared with control group, serum amylase activity, pancreatic wet weight ratio and lung MPO activity were significantlyincreased (. The expression of p-STAT3 in MAP and SAP groups are significantly different from that in control group, which indicates that STAT3 isclosely related in acute pancreatitis. Inhibition of STAT3 activity is a potential target to alleviate acute pancreatitis progression.

Topics: Acute Disease; Animals; Ceruletide; Male; Mice; Mice, Inbred BALB C; Pancreas; Pancreatitis; Signal Transduction; STAT3 Transcription Factor

Animal models are essential to understand the pathogenesis of acute pancreatitis (AP) and to develop new therapeutic strategies. Although it has been shown that cerulein-induced AP is associated with pain in experimental animals, most experiments are carried out without any pain-relieving treatment because researchers are apprehensive of an interference of the analgetic agent with AP-associated inflammation. In light of the growing ethical concerns and the legal tightening regarding animal welfare during experiments, this attitude should be changed.. Acute pancreatitis was induced by cerulein in the C57BL/6J and FVB/N mouse inbred strains. One group received vehicle only, and the other was treated with metamizol as analgetic agent. Pain sensation and parameters of AP were analyzed as well as the effect of metamizol in the pancreas and its actions in the brain.. We report that oral administration of metamizol protects cerulein-treated mice from abdominal pain without influencing the clinical and histopathological course of the disease. In addition, it could be shown that metamizol reduces the central pain response.. This study reveals that oral administered metamizol has no influence on the cerulein-induced AP and can be given as an analgesic to increase animal welfare in experiments with induced AP.

Topics: Abdominal Pain; Acute Disease; Administration, Oral; Animals; Anti-Inflammatory Agents, Non-Steroidal; Blotting, Western; Brain; Ceruletide; Cyclooxygenase 2; Dinoprostone; Dipyrone; Disease Models, Animal; Humans; Hypothalamus; Male; Mice, Inbred C57BL; Mice, Inbred Strains; Pancreatitis; Proto-Oncogene Proteins c-fos; Thalamus

The objective of this study was to determine the mechanism by which activation of peroxisome proliferator-activated receptor-γ promotes apoptosis of acinar cells in pancreatitis.. AR42j cells pretreated with the peroxisome proliferator-activated receptor-γ agonist pioglitazone were activated by cerulein as an in vitro model of acute pancreatitis. Inflammatory cytokines and amylase were detected by enzyme-linked immunosorbent assay. Cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell apoptosis was measured by flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining. Activity of caspases was determined. Bax and Bcl-2 levels were assayed by Western blot.. Cytokines, amylase, and cellular proliferation decreased in pioglitazone-pretreated cells. Pioglitazone increased the activity of caspases 3, 8, and 9 in cerulein-activated AR42j cells as well as in the pancreas of rats 3 hours after induction of severe acute pancreatitis. Acinar cell apoptosis was induced by reducing the mitochondrial membrane potential in the pioglitazone group. Pioglitazone increased expression of proapoptotic Bax proteins and decreased antiapoptotic Bcl-2 in cerulein-induced AR42j cells and decreased Bcl-2 levels in pancreatic tissue of severe acute pancreatitis rats 1 and 3 hours after induction.. Pioglitazone may promote apoptosis of acinar cells through both intrinsic and extrinsic apoptotic pathways in acute pancreatitis.

Topics: Acinar Cells; Acute Disease; Amylases; Anilides; Animals; Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Caspases; Cell Line, Tumor; Cell Survival; Ceruletide; Cytokines; Enzyme-Linked Immunosorbent Assay; Humans; Male; Membrane Potential, Mitochondrial; Pancreatitis; Pioglitazone; PPAR gamma; Proto-Oncogene Proteins c-bcl-2; Rats, Sprague-Dawley; Severity of Illness Index; Thiazolidinediones

Activation of the transcription factor NF-κB and expression of pro-inflammatory mediators have been considered as major events of acute pancreatitis (AP). Karyopherin alpha 2 (KPNA2), a member of the importin α family, reportedly modulates p65 subcellular localization.. This study aimed to investigate the expression and possible functions of KPNA2 in the AP cell and animal model, focusing on its association with NF-κB activation.. An AP cell model was established with the cerulein-stimulated AR42J and isolated rat pancreatic acinar cells. The AP rat model was induced by the intraperitoneal injection of cerulein. The secretion of TNF-α, IL-6, and LDH was detected by ELISA kits and the production of NO using nitric oxide kit. Expression of KPNA2 was measured by RT-PCR and Western blot. Expression levels of IKKα, phosphorylation of p65, and total p65 were detected by Western blot. Co-localization of KPNA2 with p65 was observed by immunofluorescence assay. To determine the biological functions of KPNA2 in cerulein-induced inflammatory response, RNA interference was employed to knockdown KPNA2 expression in AR42J and isolated pancreatic acini cells.. Cerulein stimulated KPNA2 expression and IL-6, TNF-α, NO, and LDH production in rat pancreatic acinar cells. Cerulein triggered the phosphorylation and nuclear translocation of NF-κB p65 subunit, indicating the NF-κB activation. The co-localization and nuclear accumulation of KPNA2 and p65 were detected in cerulein-treated cells. Knocking down KPNA2 hindered cerulein-induced nuclear transportation of p65 and alleviated the subsequent inflammatory response in rat pancreatic acinar cells. Additionally, KPNA2 expression was significantly up-regulated in cerulein-induced AP rat model.. KPNA2-facilitated p65 nuclear translocation promotes NF-κB activation and inflammation in acute pancreatitis.

Topics: Acinar Cells; Acute Disease; alpha Karyopherins; Animals; Blotting, Western; Cell Line; Ceruletide; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Fluorescent Antibody Technique; Gene Expression Regulation; Gene Knockdown Techniques; I-kappa B Kinase; Inflammation; Interleukin-6; Lactate Dehydrogenases; Male; NF-kappa B; Nitric Oxide; Pancreas; Pancreatitis; Phosphorylation; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription Factor RelA; Tumor Necrosis Factor-alpha; Up-Regulation

Under conditions of inflammation in the absence of micro-organisms (sterile inflammation), necrotic cells release damage-associated molecular patterns that bind to Toll-like receptors on immune cells to activate a signaling pathway that involves activation of IκB kinase and nuclear factor κB (NF-κB). Little is known about the mechanisms that control NF-κB activity during sterile inflammation. We analyzed the contribution of B-cell CLL/lymphoma 3 (BCL3), a transcription factor that associates with NF-κB, in control of sterile inflammation in the pancreas and biliary system of mice.. Acute pancreatitis (AP) was induced in C57BL/6 (control) and Bcl3(-/-) mice by intraperitoneal injection of cerulein or pancreatic infusion of sodium taurocholate. We also studied Mdr2(-/-) mice, which develop spontaneous biliary inflammation, as well as Bcl3(-/-)Mdr2(-/-) mice. We performed immunohistochemical analyses of inflamed and noninflamed regions of pancreatic tissue from patients with AP or primary sclerosing cholangitis (PSC), as well as from mice. Immune cells were characterized by fluorescence-activated cell sorting analysis. Control or Bcl3(-/-) mice were irradiated, injected with bone marrow from Bcl3(-/-) or control mice, and AP was induced.. Pancreatic or biliary tissues from patients with AP or PSC had higher levels of BCL3 and phosphorylated RelA and IκBα in inflamed vs noninflamed regions. Levels of BCL3 were higher in pancreata from control mice given cerulein than from mice without AP, and were higher in biliary tissues from Mdr2(-/-) mice than from control mice. Bcl3(-/-) mice developed more severe AP after administration of cerulein or sodium taurocholate than control mice; pancreata from the Bcl3(-/-) mice with AP had greater numbers of macrophages, myeloid-derived suppressor cells, dendritic cells, and granulocytes than control mice with AP. Activation of NF-κB was significantly prolonged in Bcl3(-/-) mice with AP, compared with control mice with AP. Bcl3(-/-)Mdr2(-/-) mice developed more severe cholestasis and had increased markers of liver injury and increased proliferation of biliary epithelial cells and hepatocytes than Mdr2(-/-) mice. In experiments with bone marrow chimeras, expression of BCL3 by acinar cells, but not myeloid cells, was required for reduction of inflammation during development of AP. BCL3 inhibited ubiquitination and proteasome-mediated degradation of p50 homodimers, which prolonged binding of NF-κB heterodimers to DNA.. BCL3 is up-regulated in inflamed pancreatic or biliary tissues from mice and patients with AP or cholangitis. Its production appears to reduce the inflammatory response in these tissues via blocking ubiquitination and proteasome-mediated degradation of p50 homodimers.

Topics: Acute Disease; Animals; ATP Binding Cassette Transporter, Subfamily B; ATP-Binding Cassette Sub-Family B Member 4; B-Cell Lymphoma 3 Protein; Bile Ducts; Bone Marrow Transplantation; Ceruletide; Cholangitis, Sclerosing; Humans; I-kappa B Proteins; Mice, Inbred C57BL; Mice, Knockout; NF-kappa B p50 Subunit; NF-KappaB Inhibitor alpha; Pancreas; Pancreatitis; Phosphorylation; Proteasome Endopeptidase Complex; Protein Multimerization; Proteolysis; Proto-Oncogene Proteins; Signal Transduction; Taurocholic Acid; Time Factors; Transcription Factor RelA; Transcription Factors; Ubiquitination

To explore the effect of B7-H3 monoclonal antibody (mAb) on cerulein-induced acute pancreatitis (AP).. Mice were randomly divided into three groups: control group, AP group and B7-H3 mAb treatment group. AP was induced in mice by intraperitoneal injections of cerulein. B7-H3 mAb was administered to the mice by subcutaneous injection 1 hour before the injections of cerulein. The blood, pancreas and lung tissues of the mice were collected 6, 12 and 24 hours after cerulein induction. Expression of B7-H3 protein was detected in the pancreas tissues of the control and AP groups by Western blotting and immunohistochemistry. Serum activities of amylase and lipase were tested by VITROS 5600 Integrated System. The pancreas wet-dry mass ratio was used to value the edema of pancreas. Pathological changes of pancreas and lung tissues were evaluated by HE staining. Serum levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-1β were detected by ELISA in all groups.. The level of B7-H3 protein increased in the pancreas tissues of the AP group after successful induction of cerulein, and reached the peak at 12 hours. Serum activities of amylase and lipase in the AP group were significantly higher than those in the control group, while decreased obviously after the intervention of B7-H3 mAb. H&E staining showed that evident inflammation appeared in pancreas and lung tissues of the AP group, and the inflammation and wet-dry mass ratio were markedly reduced in the treatment group. The levels of proinflammatory factors TNF-α, IL-6 and IL-1β in the AP group showed a time-dependent increase, and peaked at 12 hours, while in the treatment group were relatively lower.. B7-H3 is over-expressed in cerulein-induced AP. Anti-B7-H3 mAb can attenuate the inflammation and alleviate the injury of pancreas and lung tissues.

Topics: Acute Disease; Amylases; Animals; Antibodies, Monoclonal; B7 Antigens; Blotting, Western; Ceruletide; Enzyme-Linked Immunosorbent Assay; Immunohistochemistry; Interleukin-1beta; Interleukin-6; Lipase; Lung; Male; Mice; Pancreas; Pancreatitis; Time Factors; Tumor Necrosis Factor-alpha

Trans-resveratrol is a natural stilbenoid possessing multifarious pharmacological benefits; however, when orally consumed, it is rapidly metabolised by colonic microflora and converted to dihydro-resveratrol. Thus, this microbial metabolite is of great therapeutic relevance. In the present study, upon the oral administration of dihydro-resveratrol (10-50 mg/kg), the severity of acute pancreatitis in the cerulein-treated rats was significantly ameliorated as evidenced by decreased α-amylase activities in the plasma and lessened oedema formation in the pancreatic parenchyma. In addition, the generation of intracellular reactive oxidative products, including malondialdehyde and protein carbonyls, was accordingly reduced, so as the production of pro-inflammatory cytokines. While inhibiting the activities of NADPH oxidase and myeloperoxidase, the depletion of glutathione was considerably restored. Importantly, the attenuation of pancreatic oxidative damage by dihydro-resveratrol was associated with a down-regulation of the nuclear factor-kappaB and phosphatidylinositol 3'-kinase-serine/threonine kinase signalling pathways. Furthermore, we demonstrated that the solubility of dihydro-resveratrol was at least 5 times higher than trans-resveratrol whilst exhibiting a much lower cytotoxicity. Collectively, the current findings accentuate new mechanistic insight of dihydro-resveratrol in pancreatic oxidative damage, and advocate its therapeutic potential for the management of acute pancreatitis, particularly for patients unresponsive to trans-resveratrol due to the lack of proper microbial strains.

Topics: Acute Disease; alpha-Amylases; Animals; Antioxidants; Blotting, Western; Ceruletide; Cytokines; Inflammation Mediators; Malondialdehyde; Microscopy, Fluorescence; Molecular Structure; NF-kappa B; Oxidative Stress; Pancreas; Pancreatitis; Phosphatidylinositol 3-Kinases; Protein Carbonylation; Proto-Oncogene Proteins c-akt; Rats, Sprague-Dawley; Resveratrol; Signal Transduction; Stilbenes

Systemic damage in acute pancreatitis (AP) can be characterized by oxidative stress and the release of pro-inflammatory cytokines. Roflumilast has been shown to be a potent anti-inflammatory and antioxidant agent. In the present study, we aimed to investigate the effect of roflumilast in cerulein-induced AP.. Thirty-two male rats were divided into four groups: group 1 (sham), group 2 (Roflumilast), group 3 (AP), and group 4 (AP + Roflumilast). AP was induced by injecting 4 × 75 μg/kg of body weight at an interval of 1 h. Rats were killed after 12 h following the last cerulein administration. AP was confirmed by measuring the serum amylase level and inflammatory features.. Morphological changes were observed in the pancreas. Amylase levels were higher in the AP and AP + Roflumilast groups than the sham and Roflumilast groups. The serum levels of TNF-α, IL-1β, and IL-6 increased in the AP group, whereas they decreased in the Roflumilast group. The total oxidant activity (TOA) was higher and the total antioxidant capacity (TAC) was lower in the AP group. The administration of roflumilast decreased the TOA and increased the TAC in comparison with the AP group (p < 0.05 for both).. Roflumilast significantly decreases oxidative stress and inflammatory mediators in the plasma, pancreas, and lung in cerulein-induced AP rats.

Topics: Acute Disease; Aminopyridines; Amylases; Animals; Benzamides; Ceruletide; Cyclopropanes; Disease Models, Animal; Inflammation Mediators; Interleukin-1beta; Interleukin-6; Lung; Male; Oxidative Stress; Pancreas; Pancreatitis; Rats, Wistar; Tumor Necrosis Factor-alpha

Autophagy promotes tumor progression downstream of oncogenic KRAS, yet also restrains inflammation and dysplasia through mechanisms that remain incompletely characterized. Understanding the basis of this paradox has important implications for the optimal targeting of autophagy in cancer. Using a mouse model of cerulein-induced pancreatitis, we found that loss of autophagy by deletion of Atg5 enhanced activation of the IκB kinase (IKK)-related kinase TBK1 in vivo, associated with increased neutrophil and T-cell infiltration and PD-L1 upregulation. Consistent with this observation, pharmacologic or genetic inhibition of autophagy in pancreatic ductal adenocarcinoma cells, including suppression of the autophagy receptors NDP52 or p62, prolonged TBK1 activation and increased expression of CCL5, IL6, and several other T-cell and neutrophil chemotactic cytokines in vitro Defective autophagy also promoted PD-L1 upregulation, which is particularly pronounced downstream of IFNγ signaling and involves JAK pathway activation. Treatment with the TBK1/IKKε/JAK inhibitor CYT387 (also known as momelotinib) not only inhibits autophagy, but also suppresses this feedback inflammation and reduces PD-L1 expression, limiting KRAS-driven pancreatic dysplasia. These findings could contribute to the dual role of autophagy in oncogenesis and have important consequences for its therapeutic targeting. Cancer Immunol Res; 4(6); 520-30. ©2016 AACR.

Topics: Acute Disease; Adenocarcinoma; Animals; Autophagy; Autophagy-Related Protein 5; B7-H1 Antigen; Benzamides; Cell Transformation, Neoplastic; Ceruletide; Chemokine CCL5; Cytokines; Enzyme Activation; Gene Deletion; Mice; Pancreatic Neoplasms; Pancreatitis; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins p21(ras); Pyrimidines; Signal Transduction; Tumor Cells, Cultured

To evaluate the effects of certolizumab, a pegylated monoclonal antibody to tumor necrosis factor α (TNF-α), on experimentally induced acute pancreatitis.. Healthy Wistar Albino male rats (n = 36) were randomly divided into 4 groups (9 rats in each group): group 1, control group; group 2, certolizumab group; group 3, cerulein group; and group 4, cerulein + certolizumab group. Acute edematous pancreatitis was induced via intraperitoneal injection of 80-μg/kg cerulein (20 μg/kg, 4 times at 1-hour intervals) in groups 3 and 4. Certolizumab (10 μg) was intraperitoneally administered in groups 2 and 4. Serum levels of amylase, lipase, TNF-α, and lactate dehydrogenase were evaluated. Histopathology and immunohistochemistry of the pancreatic tissue for assessing the activities of malondialdehyde, myeloperoxidase, TNF-α, and caspase-3 were also performed after 72 hours.. Certolizumab treatment significantly decreased the serum levels of amylase, lipase, and lactate dehydrogenase. Histopathological edema, hemorrhage, parenchymal necrosis, and infiltration scores were also decreased, along with a decrease in malondialdehyde, myeloperoxidase, TNF-α, and caspase-3 activities.. This study suggests that certolizumab is a beneficial treatment mode for reducing the severity of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Caspase 3; Ceruletide; Male; Pancreatitis; Rats; Rats, Wistar; Tumor Necrosis Factor-alpha

Chronic pancreatitis is a significant risk factor for pancreatic cancer. Previously, we demonstrated that the pancreatic cancer cells show enhanced expression of collapsin response mediator protein 4 (CRMP4) that strongly correlates with severe venous invasion, liver metastasis, and poor prognosis. However, involvement of CRMP4 in acute or chronic pancreatitis remains unknown.. Acute and chronic pancreatitis mice models were developed by periodic injection of caerulein. The expression levels of CRMP4 and its phosphorylation were examined.. Elevated CRMP4 levels were observed in the infiltrated lymphocytes as well as in the pancreas parenchyma of both acute and chronic pancreatitis. The expression pattern of phosphorylated CRMP4 was similar to that of CRMP4. Cdk5 partially co-localized with the phosphorylated CRMP4.. Pancreatitis induces CRMP4 expression in the pancreas parenchyma and in the infiltrated lymphocytes. Overlapping expression of CRMP4 and Cdk5 may suggest that the Cdk5 is at least, in part, responsible for the phosphorylation of CRMP4. The results suggest that CRMP4 is involved in the inflammatory response in pancreatitis. Understanding the mechanisms of CRMP4 would help us to develop novel therapeutic strategies against acute or chronic pancreatitis, and pancreatic cancer.

Topics: Acute Disease; Animals; Biopsy, Needle; Cell Transformation, Neoplastic; Ceruletide; Chronic Disease; Cyclin-Dependent Kinase 5; Disease Models, Animal; Gene Expression Regulation; Humans; Immunohistochemistry; Mice; Nerve Tissue Proteins; Pancreatic Neoplasms; Pancreatitis; Phosphorylation; Precancerous Conditions; RNA, Small Interfering

Background. Chronic pancreatitis is one of the main risk factors for pancreatic cancer. In acute and chronic pancreatitis, oxidative stress is thought to play a key role. In this respect, the recently described mitochondria-targeted antioxidant SkQ1 effectively scavenges reactive oxygen species at nanomolar concentrations. Therefore, we aimed to characterize the influence of SkQ1 on tissue injury and pain in acute and chronic pancreatitis. Methods. Both acute and chronic pancreatitis were induced in C57BL/6 mice by intraperitoneal cerulein injections and treatment with SkQ1 was carried out by peroral applications. Hyperalgesia was assessed by behavioral observation and measurement of abdominal mechanical sensitivity. Blood serum and pancreatic tissue were harvested for analysis of lipase and histology. Results. SkQ1 did not influence pain, serological, or histological parameters of tissue injury in acute pancreatitis. In chronic pancreatitis, a highly significant reduction of pain-related behavior (p < 0.0001) was evident, but histological grading revealed increased tissue injury in SkQ1-treated animals (p = 0.03). Conclusion. After SkQ1 treatment, tissue injury is not ameliorated in acute pancreatitis and increased in chronic pancreatitis. However, we show an analgesic effect in chronic pancreatitis. Further studies will need to elucidate the risks and benefits of mitochondria-targeted antioxidants as an analgesic.

Topics: Acute Disease; Analgesics; Animals; Antioxidants; Behavior, Animal; Biomarkers; Ceruletide; Disease Models, Animal; Female; Hyperalgesia; Lipase; Mice, Inbred C57BL; Mitochondria; Motor Activity; Pain Perception; Pancreas; Pancreatitis; Pancreatitis, Chronic; Plastoquinone; Risk Factors

Pancreatic injury in rats is primarily detected through histopathological changes and conventional serum biomarkers such as amylase and lipase. However, amylase and lipase have a short half-life and are markers of acinar, not islet cell injury. We investigated whether circulating microRNA (miR) levels that are enriched in acinar cells (miR-217, miR-216a/b) or islet cells (miR-375) could serve as markers of pancreatic injury. Rats were treated with a single dose of either vehicle, streptozotocin (STZ), caerulein, or acetaminophen (APAP), and necropsied at 4, 24, and 48h. Pancreas, liver, heart, kidney and skeletal muscle were analyzed for histopathology. Blood was collected at necropsy and processed to serum for amylase/lipase enzymatic determinations and miR qPCR analysis. Caerulein induced degeneration/necrosis of acinar cells at 4h that persisted for 48h. Caerulein-induced injury was associated with increases in serum amylase/lipase (4h), miR-216a/b (4, 24h). In contrast, serum miR-217 was detected at all time points examined. STZ did not induce increases in either amylase or lipase but did induce increases in miR-375 levels at 4 and 24h. No increases in miR-375 were observed in caerulein-treated rats, and no increases were observed in miR-217 and miR-216a/b in STZ-treated rats. APAP induced centrilobular necrosis in the liver 24h after treatment, but did not induce pancreatic injury or increases in miR-217 or miR-375. Our results suggest that miR-217 and miR-375 represent promising biomarkers of pancreatic injury in rats.

Topics: Acetaminophen; Acinar Cells; Acute Disease; Amylases; Animals; Biomarkers; Ceruletide; Disease Models, Animal; Half-Life; Islets of Langerhans; Kidney; Lipase; Liver; Male; MicroRNAs; Muscle, Skeletal; Necrosis; Pancreas; Rats; Rats, Sprague-Dawley; Streptozocin

Acinar cells represent the primary target in necroinflammatory diseases of the pancreas, including pancreatitis. The signaling pathways guiding acinar cell repair and regeneration following injury remain poorly understood. The purpose of this study was to determine the importance of Hepatocyte Growth Factor Receptor/MET signaling as an intrinsic repair mechanism for acinar cells following acute damage and chronic alcohol-associated injury. Here, we generated mice with targeted deletion of MET in adult acinar cells (MET-/-). Acute and repetitive pancreatic injury was induced in MET-/- and control mice with cerulein, and chronic injury by feeding mice Lieber-DeCarli diets containing alcohol with or without enhancement of repetitive pancreatic injury. We examined the exocrine pancreas of these mice histologically for acinar death, edema, inflammation and collagen deposition and changes in the transcriptional program. We show that MET expression is relatively low in normal adult pancreas. However, MET levels were elevated in ductal and acinar cells in human pancreatitis specimens, consistent with a role for MET in an adaptive repair mechanism. We report that genetic deletion of MET in adult murine acinar cells was linked to increased acinar cell death, chronic inflammation and delayed recovery (regeneration) of pancreatic exocrine tissue. Notably, increased pancreatic collagen deposition was detected in MET knockout mice following repetitive injury as well alcohol-associated injury. Finally, we identified specific alterations of the pancreatic transcriptome associated with MET signaling during injury, involved in tissue repair, inflammation and endoplasmic reticulum stress. Together, these data demonstrate the importance of MET signaling for acinar repair and regeneration, a novel finding that could attenuate the symptomology of pancreatic injury.

Topics: Acinar Cells; Acute Disease; Alcohol Drinking; Animals; Ceruletide; Chronic Disease; Collagen; Disease Models, Animal; Gene Deletion; Humans; Inflammation; Macrophages; Mice, Inbred C57BL; Pancreas; Pancreatitis, Chronic; Proto-Oncogene Proteins c-met; Regeneration; Wound Healing

Pancreatic cancer is one of the most aggressive cancers and has an extremely poor prognosis. Despite recent progress in both basic and clinical research, most pancreatic cancers are detected at an incurable stage owing to the absence of disease-specific symptoms. Thus, developing novel approaches for detecting pancreatic cancer at an early stage is imperative. Our in silico and immunohistochemical analyses showed that KIAA1199 is specifically expressed in human pancreatic cancer cells and pancreatic intraepithelial neoplasia, the early lesion of pancreatic cancer, in a genetically engineered mouse model and in human patient samples. We also detected secreted KIAA1199 protein in blood samples obtained from pancreatic cancer mouse models, but not in normal mice. Furthermore, we found that assessing KIAA1199 autoantibody increased the sensitivity of detecting pancreatic cancer. These results indicate the potential benefits of using KIAA1199 as a biomarker for early-stage pancreatic cancer.

Topics: Acute Disease; Animals; Autoantibodies; Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Ceruletide; Databases, Genetic; Disease Models, Animal; Early Diagnosis; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Hyaluronoglucosaminidase; Male; Mice; Mice, Inbred C57BL; Pancreatic Neoplasms; Pancreatitis; Proteins; Tissue Array Analysis

Acute pancreatitis is basically considered as activation of inactive proenzymes in the pancreas and digestion of the gland itself. This study was performed to determine if prolidase enzyme, which plays a role in collagen metabolism, could be used as a parameter to assess the severity of pancreatitis in experimentally induced mild and severe pancreatitis.. To create experimentally induced acute pancreatitis 0.1 ml of normal saline solution (NSS) was given five times with an interval of one hour to rats in the first group; 50 µg/kg of cerulein five times with an interval of one hour in the second group; 80 µg/kg of cerulein five times with an interval of one hour in the third group, in the form of intraperitoneal injection.. When the serum prolidase values at beginning, 1st, 5th and 24th hours in group II and III were compared among themselves, there was a statistically significant increase(p < 0.05). The evaluation between groups revealed a statistically significant increase in the value of serum prolidase in group II and group III compared with the control group (p < 0.05). In comparisons performed with tissue values, a statistically significant increase determined in the value of serum prolidase in group II and group III compared with the control group was observed (p < 0.05).. The findings obtained in our study showed that prolidase activity increases directly proportionaly with the severity of pancreatitis. This allows us to postulate that prolidase enzyme activities provide guidance about the metabolism of collagen in patients with acute pancreatitis, serious damage occurring in collagen protein and metabolic control is further distorted depending on the duration and intensity of damage but to be able to speak more precisely, there is a need for further, more detailed and extensive researchs (Tab. 8, Fig. 2, Ref. 30).

Topics: Acute Disease; Amylases; Animals; Ceruletide; Collagen; Dipeptidases; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Severity of Illness Index

We aimed to evaluate the anti-inflammatory and inhibitory effects of Lithospermum erythrorhizon (LE) on cerulein-induced acute pancreatitis (AP) in a mouse model.. Acute pancreatitis was induced via intraperitoneal injection of cerulein (50 μg/kg) every hour for 6 times. In the LE, water extract (100, 250, or 500 mg/kg) was administered intraperitoneally 1 hour before the first injection of cerulein. Six hours after AP, blood, the pancreas, and the lung were harvested for further examination. In addition, pancreatic acinar cells were isolated using a collagenase method, and then, we investigated the acinar cell viability and cytokine productions.. Treatment with LE reduced pancreatic damage and AP-associated lung injury and attenuated the severity of AP, as evidenced by the reduction in neutrophil infiltration, serum amylase and lipase levels, trypsin activity, and proinflammatory cytokine expression. In addition, treatment with LE inhibited high mobility group box 1 expression in the pancreas during AP. In accordance with in vivo data, LE inhibited the cerulein-induced acinar cell death, cytokine productions, and high-mobility group box 1 expression. Furthermore, LE also inhibited the activation of p38 mitogen-activated protein kinases.. These results suggest that LE plays a protective role during the development of AP by inhibiting the activation of p38.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Biomarkers; Cell Survival; Cells, Cultured; Ceruletide; Cytokines; Cytoprotection; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Activation; Female; HMGB1 Protein; Inflammation Mediators; Lithospermum; Mice, Inbred C57BL; Neutrophil Infiltration; p38 Mitogen-Activated Protein Kinases; Pancreas; Pancreatitis; Phytotherapy; Plant Extracts; Plants, Medicinal; Severity of Illness Index; Signal Transduction; Time Factors

In this study, we identified the protein kinases that play the most distinct roles in the occurrence of acute pancreatitis (AP).. Gene expression profile data were downloaded from Gene Expression Omnibus database (GSE3644). The sample was from caerulein-induced AP mice. The intersection of the differentially expressed genes in AP mice taken from a protein kinase database was obtained for screening of the protein kinase encoded genes that were differentially expressed. Database for annotation, visualization, and integrated discovery was used for the functional enrichment analysis. Kinase inhibitors that regulated these kinases were retrieved from PubMed through text mining.. Twenty-nine differentially expressed kinase encoded genes were identified through screening. The functional enrichment analysis demonstrated that the functions of these genes were primarily enriched in "mitogen-activated protein kinase signaling pathway," followed by "extracellular regulated protein kinases pathway," "neurotrophin signaling pathway," "adherens junction," and "gap junction." SRC and epidermal growth factor receptor (EGFR) were related to extracellular regulated protein kinases pathway and also related to adherens junction as well as gap junction. On the basis of the regulated kinases, the kinase inhibitors reported in the literature were classified into multiple groups.. EGFR and SRC may be coexpressed in AP. The kinase inhibitors working together in SRC and EGFR may play better efficacy in the treatment of AP.

Topics: Acute Disease; Animals; Ceruletide; Databases, Genetic; Disease Models, Animal; ErbB Receptors; Gene Expression Profiling; Gene Expression Regulation, Enzymologic; Gene Regulatory Networks; Oligonucleotide Array Sequence Analysis; Pancreatitis; Protein Interaction Maps; Protein Kinase Inhibitors; RNA, Messenger; Signal Transduction; src-Family Kinases

Pancreatic acinar cell necrosis and subsequent inflammatory response aggravate acute pancreatitis (AP). Tetraspanin CD9 has been reported to mediate inflammatory signaling by regulating molecular organization at the cell surface. This study aimed to investigate the role of CD9 in caerulein-induced AP (CIP) in mice.. The expression of CD9 was detected in CIP in mice in vivo and cholecystokinin (CCK)/recombinant mouse tumor necrosis factor (rmTNF)-α induced pancreatic acinar cell death in vitro by quantitative real-time polymerase chain reaction, Western blot and immunofluorescence. The roles of CD9 in pancreatic acinar cell death and inflammatory response were further studied through the deletion of CD9 expression using small interfering RNA (siRNA).. CD9 was markedly upregulated in pancreatic tissues in mice during the early onset of CIP and was located mainly at the pancreatic acinar cell surface, which was associated with pancreatic damage. Additionally, incubation with CCK or rmTNF-α directly increased the expression of CD9 in isolated mice pancreatic acinar cells in vitro. The deletion of CD9 expression partially reversed both pancreatic acinar cell death induced by CCK and mRNA levels of proinflammatory cytokines produced by damaged acinar cells.. These results indicate that increased CD9 expression may be involved in pancreatic injury, possibly via the promotion of cytokine expressions in CIP in mice.

Topics: Acinar Cells; Acute Disease; Animals; Ceruletide; Cholecystokinin; Cytokines; Female; Gene Expression; Mice; Mice, Inbred BALB C; Pancreas; Pancreatitis; Random Allocation; RNA; RNA, Small Interfering; Signal Transduction; Tetraspanin 29; Tumor Necrosis Factor-alpha; Up-Regulation

The onset of acute pancreatitis (AP) is characterized by early protease activation followed by inflammation and organ damage, but the mechanisms are poorly understood.. We hypothesized that histone deacetylase (HDAC) inhibition might exert protective effects on AP and investigated the role of HDAC in trypsin activation, inflammation, and tissue damage in severe AP.. Male C57Bl/6 mice were treated i.p. with the HDAC inhibitor trichostatin A (2 mg/kg) prior to retrograde infusion of taurocholic acid (5 %) into the pancreatic duct. Serum levels of amylase and interleukin (IL)-6, pancreatic levels of macrophage inflammatory protein-2 (MIP-2) as well as tissue morphology and myeloperoxidase activity in the pancreas and lung were determined 24 h after taurocholate challenge. Trypsin activation was analyzed in isolated acinar cells. Quantitative RT-PCR was used to examine the expression of pro-inflammatory mediators in the pancreas.. Pretreatment with trichostatin A decreased amylase levels by 70 % and protected against tissue injury in the pancreas. Moreover, HDAC inhibition reduced systemic IL-6 by more than 95 % and pulmonary myeloperoxidase activity by 75 %. Notably, inhibition of HDAC abolished taurocholate-induced gene expression of cyclooxygenase-2, MIP-2, monocyte chemotactic protein-1, IL-6, and IL-1β in the pancreas. In addition, HDAC inhibition reduced cerulein-induced trypsinogen activation in isolated acinar cells.. Our findings show that HDAC regulates trypsin activation, inflammation, and tissue damage in AP. Thus, targeting HDAC could serve as novel therapeutic approach in the management of severe AP.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Ceruletide; Chemokine CXCL2; Cytoprotection; Disease Models, Animal; Enzyme Activation; Histone Deacetylase Inhibitors; Histone Deacetylases; Hydroxamic Acids; Inflammation Mediators; Injections, Intraperitoneal; Interleukin-6; Lung; Male; Mice, Inbred C57BL; Pancreas; Pancreatitis; Peroxidase; Signal Transduction; Taurocholic Acid; Trypsin

The mitogen-activated protein kinases (MAPKs) signaling pathway is involved in inflammatory process. However, the mechanism is not clear. The present study was to investigate the role of p38 MAPK in acute pancreatitis in mice.. Mice were divided into 4 groups: saline control; acute pancreatitis induced with repeated injections of cerulein; control plus p38 MAPK inhibitor SB203580; and acute pancreatitis plus SB203580. The pancreatic histology, pancreatic enzymes, cytokines, myeloperoxidase activity, p38 MAPK and heat shock protein (HSP) 60 and 70 were evaluated.. Repeated injections of cerulein resulted in acute pancreatitis in mice, accompanying with the activation of p38 MAPK and overexpression of HSP60 and HSP70 in the pancreatic tissues. Treatment with SB203580 significantly inhibited the activation of p38 MAPK, and furthermore, inhibited the expression of HSP60 and HSP70 in the pancreas, the inflammatory cytokines in the serum, and myeloperoxidase activity in the lung.. The p38 MAPK signaling pathway is involved in the regulation of inflammatory response and the expression of HSP60 and HSP70 in acute pancreatitis.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Biomarkers; Ceruletide; Chaperonin 60; Disease Models, Animal; HSP70 Heat-Shock Proteins; Imidazoles; Inflammation Mediators; Lung; Mice, Inbred C57BL; Mitochondrial Proteins; p38 Mitogen-Activated Protein Kinases; Pancreas; Pancreatitis; Peroxidase; Phosphorylation; Protein Kinase Inhibitors; Pyridines; Signal Transduction

Guggulsterone (GS), a plant steroid and a compound found at high levels in Commiphora myrrha, exhibits anti-inflammatory, anti-cancer, and cholesterol-lowering effects. However, the potential of GS to ameliorate acute pancreatitis (AP) is unknown. The aim of this study was to evaluate the effects of GS on cerulein-induced AP. AP was induced by intraperitoneally injecting supramaximal concentrations of the stable cholecystokinin analog cerulein (50 μg/kg) hourly for 6 h. In the GS-treated group, GS was administered intraperitoneally (10, 25, or 50mg/kg) 1 h before the first cerulein injection. Mice were sacrificed 6 h after the final cerulein injection. Blood samples were collected to measure serum lipase levels and evaluate cytokine production. The pancreas and lung were rapidly removed for morphologic and histological examinations, flow cytometry analysis, myeloperoxidase (MPO) assay, and real-time reverse transcription-polymerase chain reaction analysis. Pre-treatment with GS attenuated cerulein-induced histological damage, reduced pancreas weight/body weight ratio, decreased serum lipase levels, inhibited infiltrations of macrophages and neutrophils, and suppressed cytokine production. Additionally, GS treatment suppressed the activation of extracellular signal-regulated protein kinase (ERK) and c-Jun N-terminal kinase (JNK) in the pancreas in cerulein-induced pancreatitis. In conclusion, our results suggest that GS attenuates AP via deactivation of ERK and JNK.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Blotting, Western; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Extracellular Signal-Regulated MAP Kinases; Female; Injections, Intraperitoneal; JNK Mitogen-Activated Protein Kinases; Lipase; Mice, Inbred C57BL; Pancreas; Pancreatitis; Pregnenediones

Although oxidative stress has been strongly implicated in the development of acute pancreatitis (AP), antioxidant therapy in patients has so far been discouraging. The aim of this study was to assess potential protective effects of a mitochondria-targeted antioxidant, MitoQ, in experimental AP using in vitro and in vivo approaches. MitoQ blocked H2O2-induced intracellular ROS responses in murine pancreatic acinar cells, an action not shared by the control analogue dTPP. MitoQ did not reduce mitochondrial depolarisation induced by either cholecystokinin (CCK) or bile acid TLCS, and at 10 µM caused depolarisation per se. Both MitoQ and dTPP increased basal and CCK-induced cell death in a plate-reader assay. In a TLCS-induced AP model MitoQ treatment was not protective. In AP induced by caerulein hyperstimulation (CER-AP), MitoQ exerted mixed effects. Thus, partial amelioration of histopathology scores was observed, actions shared by dTPP, but without reduction of the biochemical markers pancreatic trypsin or serum amylase. Interestingly, lung myeloperoxidase and interleukin-6 were concurrently increased by MitoQ in CER-AP. MitoQ caused biphasic effects on ROS production in isolated polymorphonuclear leukocytes, inhibiting an acute increase but elevating later levels. Our results suggest that MitoQ would be inappropriate for AP therapy, consistent with prior antioxidant evaluations in this disease.

Topics: Acinar Cells; Acute Disease; Animals; Antioxidants; Apoptosis; Ceruletide; Cholecystokinin; Disease Models, Animal; Inflammation; Male; Membrane Potential, Mitochondrial; Mice; Mitochondria; Necrosis; Organophosphorus Compounds; Oxidative Stress; Pancreas; Pancreatitis; Reactive Oxygen Species; Taurolithocholic Acid; Ubiquinone

Hypertriglyceridemic pancreatitis (HTGP) is often encountered clinically as a common form of recurrent acute pancreatitis (AP). It is important to evaluate the management of severe hypertriglyceridemia (HTG) or anti-inflammation in the prophylaxis of HTGP in the clinic. FTY720 (2-amino-2[2-(4-octylphenyl) ethyl]-1, 3-propanediol) is a new anti-inflammatory agent with low toxicity and reported to ameliorate lung injury with pancreatitis in rat. We evaluated its protective affection on AP induced by seven hourly intraperitoneal injection of cerulein in apolipoprotein CIII transgenic mice with severe HTG. FTY720 at 1.5 mg/kg was administered by gastric lavage daily for 3 days before induction of AP. The effects of FTY720 to protect against HTGP were assessed by serum amylase, pancreatic pathological scores, immunostaining, and the expression of inflammatory cytokine genes. As a result, injection of cerulein resulted in more severe pathological changes of AP and higher monocyte chemoattractant protein 1 expression in the pancreas in transgenic than in nontransgenic mice. FTY720 pretreatment improved the pathological severity of AP and decreased the expression of monocyte chemoattractant protein 1 in the pancreas significantly, especially near fourfold reduction in transgenic mice. However, FTY720 did not affect plasma triglyceride levels, and other inflammatory factors and plasma amylase were not correlated with the extent of pancreatic damage in AP with or without FTY720 administration. In summary, our study in a new model, apolipoprotein CIII transgenic mice, demonstrated that HTG mice are susceptible to induction of AP. Prophylactic treatment of FTY720 can significantly attenuate cerulein-induced AP and hence warrant further investigation of sphingosine-1-phosphate receptors agonist for potential clinical application in recurrent attacks of HTGP.

Topics: Acute Disease; Amylases; Animals; Apolipoprotein C-III; Ceruletide; Chemokine CCL2; Drug Evaluation, Preclinical; Female; Fingolimod Hydrochloride; Hypertriglyceridemia; Immunosuppressive Agents; Lymphocyte Count; Mice, Transgenic; Pancreas; Pancreatitis

Pancreatitis is a significant clinical problem and the lack of effective therapeutic options means that treatment is often palliative rather than curative. A deeper understanding of the pathogenesis of both acute and chronic pancreatitis is necessary to develop new therapies. Pathological changes in pancreatitis are dependent on innate immune cell recruitment to the site of initial tissue damage, and on the coordination of downstream inflammatory pathways. The chemokine receptor CXCR2 drives neutrophil recruitment during inflammation, and to investigate its role in pancreatic inflammation, we induced acute and chronic pancreatitis in wild-type and Cxcr2(-/-) mice. Strikingly, Cxcr2(-/-) mice were strongly protected from tissue damage in models of acute pancreatitis, and this could be recapitulated by neutrophil depletion or by the specific deletion of Cxcr2 from myeloid cells. The pancreata of Cxcr2(-/-) mice were also substantially protected from damage during chronic pancreatitis. Neutrophil depletion was less effective in this model, suggesting that CXCR2 on non-neutrophils contributes to the development of chronic pancreatitis. Importantly, pharmacological inhibition of CXCR2 in wild-type mice replicated the protection seen in Cxcr2(-/-) mice in acute and chronic models of pancreatitis. Moreover, acute pancreatic inflammation was reversible by inhibition of CXCR2. Thus, CXCR2 is critically involved in the development of acute and chronic pancreatitis in mice, and its inhibition or loss protects against pancreatic damage. CXCR2 may therefore be a viable therapeutic target in the treatment of pancreatitis.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Ceruletide; Cytoprotection; Disease Models, Animal; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Neutrophil Infiltration; Neutrophils; Pancreas; Pancreatitis; Pancreatitis, Chronic; Peptides; Receptors, Interleukin-8B; Signal Transduction; Time Factors

We describe the first mouse model of pancreatic intraepithelial neoplasia (PanIN) lesions induced by alcohol in the presence and absence of chronic pancreatitis.. Pdx1-Cre;LSL-K-ras mice were exposed to Lieber-DeCarli alcohol diet for 6 weeks with cerulein injections. The PanIN lesions and markers of fibrosis, inflammation, histone deacetylation, epithelial-to-mesenchymal transition (EMT), and cancer stemness were measured by immunohistochemistry and Western.. Exposure of Pdx1-Cre;LSL-K-ras mice to an alcohol diet significantly stimulated fibrosis and slightly but not significantly increased the level of PanIN lesions associated with an increase in tumor-promoting M2 macrophages. Importantly, the alcohol diet did not increase activation of stellate cells. Alcohol diet and cerulein injections resulted in synergistic and additive effects on PanIN lesion and M2 macrophage phenotype induction, respectively. Cerulein pancreatitis caused stellate cell activation, EMT, and cancer stemness in the pancreas. Pancreatitis caused histone deacetylation, which was promoted by the alcohol diet. Pancreatitis increased EMT and cancer stemness markers, which were not further affected by the alcohol diet.. The results suggest that alcohol has independent effects on promotion of PDAC associated with fibrosis formed through a stellate cell-independent mechanism and that it further promotes early PDAC and M2 macrophage induction in the context of chronic pancreatitis.

Topics: Acetylation; Acute Disease; Animals; Carcinoma in Situ; Cell Transformation, Neoplastic; Ceruletide; Disease Models, Animal; Epithelial-Mesenchymal Transition; Ethanol; Fibrosis; Histones; Macrophages; Mice, Transgenic; Neoplastic Stem Cells; Pancreas; Pancreatic Neoplasms; Pancreatic Stellate Cells; Pancreatitis; Pancreatitis, Alcoholic; Pancreatitis, Chronic; Time Factors

Lupeol is a triterpenoid commonly found in fruits and vegetables and is known to exhibit a wide range of biological activities, including antiinflammatory and anti-cancer effects. However, the effects of lupeol on acute pancreatitis specifically have not been well characterized. Here, we investigated the effects of lupeol on cerulein-induced acute pancreatitis in mice. Acute pancreatitis was induced via an intraperitoneal injection of cerulein (50 µg/kg). In the lupeol treatment group, lupeol was administered intraperitoneally (10, 25, or 50 mg/kg) 1 h before the first cerulein injection. Blood samples were taken to determine serum cytokine and amylase levels. The pancreas was rapidly removed for morphological examination and used in the myeloperoxidase assay, trypsin activity assay, and real-time reverse transcription polymerase chain reaction. In addition, we isolated pancreatic acinar cells using a collagenase method to examine the acinar cell viability. Lupeol administration significantly attenuated the severity of pancreatitis, as was shown by reduced pancreatic edema, and neutrophil infiltration. In addition, lupeol inhibited elevation of digestive enzymes and cytokine levels, such as tumor necrosis factor (TNF)-α, interleukin (IL)-1, and interleukin (IL)-6. Furthermore, lupeol inhibited the cerulein-induced acinar cell death. In conclusion, these results suggest that lupeol exhibits protective effects on cerulein-induced acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Cell Survival; Ceruletide; Cytokines; Injections, Intraperitoneal; Lipase; Male; Mice; Mice, Inbred C57BL; Neutrophil Infiltration; Pancreas; Pancreatitis; Pentacyclic Triterpenes; Peroxidase; Plant Extracts; Protective Agents; Tumor Necrosis Factor-alpha

Extracellular histones are rapidly cleared by the liver and rarely detectable in the circulation unless there is extensive cell death, as in severe trauma and sepsis. This study investigated whether circulating histones are elevated in experimental acute pancreatitis models and correlate to disease severity.. Acute pancreatitis was induced in mice by: (1) 4 or (2) 12 intraperitoneal injections of cerulein (50 μg/kg) at 1 hour apart; (3) retrograde infusion of 3.5% sodium taurocholate into the biliopancreatic duct. Mice were sacrificed at various time points to collect blood and tissues. Severity of pancreatitis was assessed by biochemical markers and histopathology. Circulating histones were determined by Western blotting.. Four cerulein injections induced edematous pancreatitis, whereas 12 cerulein injections and ductal taurocholate infusion caused necrotizing pancreatitis. Circulating histones were barely detectable in the blood of animals with edematous pancreatitis but significantly increased in necrotizing pancreatitis. The levels of circulating histones were strongly correlated to histopathological scores of necrosis of the pancreas.. Circulating histones increased significantly in necrotizing pancreatitis due to extensive pancreatic acinar cell death. Levels of circulating histones may have translational potential as a biomarker of disease severity.

Topics: Acute Disease; Analysis of Variance; Animals; Biomarkers; Blotting, Western; Ceruletide; Disease Models, Animal; Histones; Humans; Male; Mice, Inbred C57BL; Pancreatitis; Pancreatitis, Acute Necrotizing; Severity of Illness Index; Taurocholic Acid

Activation of the transcription factor κB (NF-κB) and secretion of pro-inflammatory mediators are major events in acute pancreatitis (AP). Recently, O-linked-N-acetylglucosamine (O-GlcNAc) modification, one type of posttranslational modifications, reportedly attunes NF-κB function. However, the expression of O-GlcNAc transferase (OGT), the enzyme responsible for O-GlcNAcylation of proteins, in AP, and the possible contribution of OGT-mediated O-GlcNAcylation to the NF-κB inflammatory activation in pancreatic acinar cells and to the AP progression have not been understood. This study focused on the effects and mechanisms of OGT-mediated O-GlcNAcylation during AP.. An AP cell model was established with the caerulein-stimulated AR42 J rat pancreatic acinar cells. The secretion of pro-inflammatory cytokines TNF-α was detected by ELISA kits, and the production of NO was determined using the colorimetric Griess reaction. Expression of OGT was measured by RT-PCR and Western blot. Expression levels of RL2, phosphorylation of p65, total p65, IKKα were detected by Western blot. The NF-κB activity was evaluated by luciferase reporter gene assay. To determine the biological functions of OGT in caerulein-induced inflammatory response, RNA interference and PUGNAc, the inhibitor of O-GlcNAcase (OGA) was employed to regulate OGT expression in AR42 J cells.. Caerulein significantly up-regulated the expression of OGT, and increased the global protein O-GlcNAcylation level in AR42 J cells. Reduction of OGT by small interfering RNA (siRNA) inhibited caerulein-triggered inflammation, assessed by the production of pro-inflammatory mediators (TNF-α and NO). We also demonstrated that O-GlcNAcylation directly modified the NF-κB p65 subunit and its upstream activating kinases IKKα in AR42 J cells. Lowering O-GlcNAcylation by OGT knockdown attenuated p65 activating phosphorylation, nuclear translocation, NF-κB transcriptional activity and levels of NF-κB transcriptional targets TNF-α and NO; on the contrary, elevating O-GlcNAc through PUGNAc increased IKKα and p65 O-GlcNAcylation accompanied by increased p65 phosphorylation, activity and levels of TNF-α and NO in caerulein-treated cells.. Our results demonstrate for the first time that OGT-mediated O-GlcNAcylation promotes NF-κB signaling activation and inflammation in pancreatic acinar cells, which might promote the progression of AP.

Topics: Acinar Cells; Acute Disease; Acylation; Animals; Cell Line; Ceruletide; Cytokines; Immunoprecipitation; Inflammation; N-Acetylglucosaminyltransferases; NF-kappa B; Nitric Oxide; Pancreatitis; Rats; RNA Interference; RNA, Small Interfering; Tumor Necrosis Factor-alpha

The objective of this study was to investigate the potential protective effects of fucoidan, an L- and P-selectin modulator, in 2 murine models of acute pancreatitis.. Acute pancreatitis was induced in mice either by the retrograde infusion of taurolithocholic acid sulfate into the pancreatic duct or by intraperitoneal injections of cerulein (50 μg/kg per hour). The experimental groups received fucoidan (25 mg/kg, intravenously) before pancreatitis induction, whereas control groups received only saline. After 24 hours, serum amylase, lipase, interleukin 1β (IL-1β), tumor necrosis factor α (TNF-α), and nitrite were measured. In addition, myeloperoxidase (MPO) activity (lung and pancreas) and histological assessment (pancreas) were determined.. Serum amylase, lipase, nitrite, TNF-α, and IL-1β, and pancreatic and lung MPO were increased in both taurolithocholic acid sulfate and cerulein acute pancreatitis compared with the respective control groups. Fucoidan significantly decreased the augmented levels of amylase, lipase, pancreatic and lung MPO, TNF-α, IL-1β, and nitrite in both models. Pancreas histological changes observed in both acute pancreatitis models were significantly attenuated by fucoidan.. Fucoidan reduced the severity of acute pancreatitis in mice by decreasing neutrophil infiltration and systemic inflammation, suggesting that modulation of selectins may constitute a promising therapeutic approach.

Topics: Acute Disease; Amylases; Analysis of Variance; Animals; Anti-Ulcer Agents; Ceruletide; Interleukin-1beta; L-Selectin; Lipase; Lung; Male; Mice; Neutrophil Infiltration; Nitrites; P-Selectin; Pancreas; Pancreatitis; Peroxidase; Polysaccharides; Taurolithocholic Acid; Tumor Necrosis Factor-alpha

The protein tyrosine phosphatases (PTPs) SHP-1, SHP-2 and PTP1B are overexpressed early on during the development of cerulein -induced acute pancreatitis (AP) in rats, and their levels can be modulated by some species of mitogen-activated protein kinases (MAPKs), the intracellular levels of cAMP and by general leukocyte infiltration, the latter at least for SHP-2 and PTP1B. In this study we show that cerulein treatment activates extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) but not p38 MAPK during the early phase of cerulein-induced AP (2h after the first injection of cerulein). Therefore, by using the MAPK inhibitors SP600125 (a specific JNK inhibitor) and PD98059 (a specific ERK inhibitor), we have unmasked the particular MAPK that underlies the modulation of the expression levels of these PTPs. JNK would act by preventing SHP-1 protein expression from increasing beyond a certain level. ERK 1/2 was the main MAPK involved in the increase in SHP-2 protein expression due to cerulein. JNK negatively modulated the SH2-domain containing PTPs. Both MAPKs played a role in the increase in PTP1B protein expression due to cerulein. Finally, by using the white blood cell inhibitors vinblastine sulfate, gadolinium chloride and FK506 (tacrolimus), we show that the macrophage activity or T-lymphocytes does not modulate the expression of any of the PTPs, although neutrophil infiltration was found to be a regulator of SHP-2 and PTP1B protein expression due to cerulein.

Topics: Acute Disease; Animals; Anthracenes; Ceruletide; Cyclic AMP; Flavonoids; Immunoblotting; Immunohistochemistry; JNK Mitogen-Activated Protein Kinases; Male; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Neutrophil Infiltration; Pancreas; Pancreatitis; Protein Tyrosine Phosphatase, Non-Receptor Type 1; Protein Tyrosine Phosphatase, Non-Receptor Type 11; Protein Tyrosine Phosphatase, Non-Receptor Type 6; Rats; Rats, Wistar; Time Factors

In the past decades, a greater understanding of acute pancreatitis has led to improvement in mortality rates. Nevertheless, this disease continues to be a health care system problem due to its economical costs. Future strategies such as antioxidant supplementation could be very promising, regarding to beginning and progression of the disease. For this reason, this study was aimed at assessing the effect of exogenous administration of resveratrol during the induction process of acute pancreatitis caused by the cholecystokinin analog cerulein in rats. Resveratrol pretreatment reduced histological damage induced by cerulein treatment, as well as hyperamylasemia and hyperlipidemia. Altered levels of corticosterone, total antioxidant status, and glutathione peroxidase were significantly reverted to control levels by the administration of resveratrol. Lipid peroxidation was also counteracted; nevertheless, superoxide dismutase enzyme was overexpressed due to resveratrol pretreatment. Related to immune response, resveratrol pretreatment reduced pro-inflammatory cytokine IL-1β levels and increased anti-inflammatory cytokine IL-10 levels. In addition, pretreatment with resveratrol in cerulein-induced pancreatitis rats was able to reverse, at least partially, the abnormal calcium signal induced by treatment with cerulein. In conclusion, this study confirms antioxidant and immunomodulatory properties of resveratrol as chemopreventive in cerulein-induced acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Antioxidants; Ceruletide; Corticosterone; Female; Glutathione Peroxidase; Interleukin-10; Interleukin-1beta; Lipase; Lipid Peroxidation; Male; Malondialdehyde; Oxidative Stress; Pancreas; Pancreatitis; Rats; Rats, Wistar; Resveratrol; Stilbenes; Superoxide Dismutase

High calcium concentrations are an established risk factor for pancreatitis. We have investigated whether increasing magnesium concentrations affect pathological calcium signals and premature protease activation in pancreatic acini, and whether dietary or intraperitoneal magnesium administration affects the onset and course of experimental pancreatitis.. Pancreatic acini were incubated with up to 10 mM magnesium; [Ca(2+)](i) (fura-2AM) and intracellular protease activation (fluorogenic substrates) were determined over 60 min. Wistar rats received chow either supplemented or depleted for magnesium (<300 ppm to 30 000 ppm) over two weeks before pancreatitis induction (intravenous caerulein 10 µg/kg/h/4 h); controls received 1 µg/kg/h caerulein or saline. C57BL6/J mice received four intraperitoneal doses of magnesium (NaCl, Mg(2+) 55 192 or 384 mg/kg bodyweight) over 72 h, then pancreatitis was induced by up to eight hourly supramaximal caerulein applications. Pancreatic enzyme activities, protease activation, morphological changes and the immune response were investigated.. Increasing extracellular Mg(2+) concentration significantly reduced [Ca(2+)](i) peaks and frequency of [Ca(2+)](i) oscillations as well as intracellular trypsin and elastase activity. Magnesium administration reduced pancreatic enzyme activities, oedema, tissue necrosis and inflammation and somewhat increased Foxp3-positiv T-cells during experimental pancreatitis. Protease activation was found in animals fed magnesium-deficient chow-even with low caerulein concentrations that normally cause no damage.. Magnesium supplementation significantly reduces premature protease activation and the severity of pancreatitis, and antagonises pathological [Ca(2+)](i) signals. Nutritional magnesium deficiency increases the susceptibility of the pancreas towards pathological stimuli. These data have prompted two clinical trials on the use of magnesium in patients at risk for pancreatitis.

Topics: Acute Disease; Animals; Biomarkers; Calcium; Ceruletide; Dietary Supplements; Disease Progression; Hydrolases; Magnesium; Magnesium Deficiency; Male; Mice; Pancreatitis; Peptide Hydrolases; Rats; Rats, Wistar; Severity of Illness Index; Treatment Outcome

Consistent, sensitive biomarkers of exocrine pancreatic injury (EPIJ) in animal models and humans have historically represented a poorly met need for investigators and clinicians.. Sprague-Dawley CD/International Genetic Standard system (IGS) rats were administered cerulein or cyanohydroxybutene (CHB) to induce EPIJ. Serum samples were taken at time points between 1- and 168-hr postinjection (PI), and rats were sacrificed between 24- and 168-hr PI.. We investigated a series of serum-based biomarkers including amylase, lipase, pancreas-enriched microRNAs (miRs) and inflammation biomarkers compared with concurrent hematology and pancreatic histology.. Microscopic EPIJ was not associated with consistent changes in hematology or inflammation biomarkers. Increased severity scores for EPIJ correlated with increased amylase and lipase values, although severity of EPIJ did not always correlate with the magnitude of enzyme increases. Microscopic EPIJ was most severe at 24 to 48 hr; increases in miR-216a (32-fold) and miR-375 (23-fold) were present at 24 hr and, along with enzymes, were normalized by 48 hr in the cerulein study. MiRs-216a and 375 were increased by ∼800- and 500-fold, respectively, at 24 hr while miR-375 remained elevated until 72 hr in the CHB study. Impact statement: Pancreas-enriched miRs hold promise as novel serum-based biomarkers for EPIJ.

Topics: Acute Disease; Alkenes; Amylases; Animals; Biomarkers; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Exocrine Pancreatic Insufficiency; Lipase; Male; MicroRNAs; Nitriles; Pancreas; Rats; Rats, Sprague-Dawley

The aim of this study was to evaluate the effects of Opuntia humifusa (OH) on cerulein-induced acute pancreatitis (AP).. Acute pancreatitis was induced via intraperitoneal injection of cholecystokinin analog cerulein (50 μg/kg). In the OH pretreatment group, OH was administered intraperitoneally (100, 250, or 500 mg/kg) 1 hour before first cerulein injection. In the posttreatment group, OH was administered intraperitoneally (500 mg/kg) 1 hour after the first cerulein injection. Furthermore, we isolated the pancreatic acinar cells using collagenase method, then investigated the acinar cell viability, cytokine productions, and the regulating mechanisms.. The both pretreatment and posttreatment of OH treatment attenuated the severity of AP, as shown by the histology of the pancreas and lung, and inhibited neutrophil infiltration; serum amylase and lipase activities; proinflammatory cytokine expression such as interleukin 1, interleukin 6, and tumor necrosis factor α; and cell death including apoptosis and necrosis. Furthermore, OH inhibited the activation of c-Jun N-terminal kinases.. These results suggest that OH reduces the severity of AP by inhibiting acinar cell death through c-Jun N-terminal kinases.

Topics: Acinar Cells; Acute Disease; Amylases; Animals; Apoptosis; Blotting, Western; Cell Survival; Cells, Cultured; Ceruletide; Cytokines; Dose-Response Relationship, Drug; Female; Gene Expression; HMGB1 Protein; Injections, Intraperitoneal; Lipase; Mice; Mice, Inbred C57BL; Mitogen-Activated Protein Kinases; Opuntia; Pancreas; Pancreatitis; Plant Extracts; Reverse Transcriptase Polymerase Chain Reaction; Time Factors

High mobility group box 1 (HMGB1) is an abundant protein that regulates chromosome architecture and also functions as a damage-associated molecular pattern molecule. Little is known about its intracellular roles in response to tissue injury or during subsequent local and systemic inflammatory responses. We investigated the function of Hmgb1 in mice after induction of acute pancreatitis.. We utilized a Cre/LoxP system to create mice with pancreas-specific disruption in Hmbg1 (Pdx1-Cre; HMGB1(flox/flox) mice). Acute pancreatitis was induced in these mice (HMGB1(flox/flox) mice served as controls) after injection of l-arginine or cerulein. Pancreatic tissues and acinar cells were collected and analyzed by histologic, immunoblot, and immunohistochemical analyses.. After injection of l-arginine or cerulein, Pdx1-Cre; HMGB1(flox/flox) mice developed acute pancreatitis more rapidly than controls, with increased mortality. Pancreatic tissues of these mice also had higher levels of serum amylase, acinar cell death, leukocyte infiltration, and interstitial edema than controls. Pancreatic tissues and acinar cells collected from the Pdx1-Cre; HMGB1(flox/flox) mice after l-arginine or cerulein injection demonstrated nuclear catastrophe with greater nucleosome release when compared with controls, along with increased phosphorylation/activation of RELA nuclear factor κB, degradation of inhibitor of κB, and phosphorylation of mitogen-activated protein kinase. Inhibitors of reactive oxygen species (N-acetyl-l-cysteine) blocked l-arginine-induced DNA damage, necrosis, apoptosis, release of nucleosomes, and activation of nuclear factor κB in pancreatic tissues and acinar cells from Pdx1-Cre; HMGB1(flox/flox) and control mice. Exogenous genomic DNA and recombinant histone H3 proteins significantly induced release of HMGB1 from mouse macrophages; administration of antibodies against H3 to mice reduced serum levels of HMGB1 and increased survival after l-arginine injection.. In 2 mouse models of acute pancreatitis, intracellular HMGB1 appeared to prevent nuclear catastrophe and release of inflammatory nucleosomes to block inflammation. These findings indicate a role for the innate immune response in tissue damage.

Topics: Acute Disease; Animals; Arginine; Cell Death; Ceruletide; Disease Models, Animal; DNA Damage; High Mobility Group Proteins; Histones; Immunity, Innate; Mice; Mice, Inbred C57BL; Mice, Knockout; Nucleosomes; Oxidative Stress; Pancreas; Pancreatitis; Reactive Oxygen Species; Repressor Proteins; Signal Transduction; Time Factors

Angiotensin-converting enzyme (ACE) and its effector peptide angiotensin II (Ang II) have been implicated in the pathogenesis of pancreatitis. Angiotensin-converting enzyme 2 (ACE2) degrades Ang II to angiotensin-(1-7) [Ang-(1-7)] and has recently been described to have an antagonistic effect on ACE signalling. However, the specific underlying role of ACE2 in the pathogenesis of severe acute pancreatitis (SAP) is unclear. In the present study, the local imbalance of ACE and ACE2, as well as Ang II and Ang-(1-7) expression, was compared in wild-type (WT) and ACE2 knock-out (KO) or ACE2 transgenic (TG) mice subjected to cerulein-induced SAP. Serum amylase, tumour necrosis factor-α, interleukin (IL)-1β, IL-6 and IL-10 levels and histological morphometry were used to determine the severity of pancreatitis. In WT mice, pancreatic ACE and Ang II and serum Ang II expression increased (P < 0.05), while pancreatic ACE2 and Ang-(1-7) and serum Ang-(1-7) levels were also significantly elevated (P < 0.05) from 2 to 72 h after the onset of SAP. However, the ratio of pancreatic ACE2 to ACE expression was significantly reduced (from 1.46 ± 0.09 to 0.27 ± 0.05, P < 0.001) and paralleled the severity of pancreatitis. The Ace2 KO mice exhibited increased levels of tumour necrosis factor-α, IL-1β, IL-6, multifocal coagulative necrosis and inflammatory infiltrate, and lower levels of serum IL-10 and pancreatic Ang-(1-7) (4.70 ± 2.13 versus 10.87 ± 2.51, P < 0.001) compared with cerulein-treated WT mice at the same time point. Conversely, Ace2 TG mice with normal ACE expression were more resistant to SAP challenge as evidenced by a decreased inflammatory response, attenuated pathological changes and increased survival rates. These data suggest that the ACE2-ACE imbalance plays an important role in the pathogenesis of SAP and that pancreatic ACE2 is an important factor in determining the severity of SAP.

Topics: Acute Disease; Amylases; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Biomarkers; Ceruletide; Disease Models, Animal; Genotype; Inflammation Mediators; Male; Mice, Inbred C57BL; Mice, Knockout; Necrosis; Pancreas; Pancreatitis; Peptide Fragments; Peptidyl-Dipeptidase A; Phenotype; Severity of Illness Index; Time Factors

The aim of this study was to investigate whether BML-111 can exert protective effects on cerulein-induced acute pancreatitis-associated lung injury (APALI) via activation of nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant responsive element (ARE) signaling pathway. Severe acute pancreatitis (SAP) was established by intraperitoneal injection of cerulein (50 μg/kg) seven times at hourly intervals and Escherichia coli lipopolysaccharide (10 mg/kg) once after the last dose of cerulein immediately. BML-111 (1 mg/kg) was administered 1 h before the first injection of cerulein. Samples were taken at 3, 6, 12, and 24 h after the last injection. Pathologic lesions of the pancreas and lung tissues as well as the levels of serum amylase were analyzed; Myeloperoxidase (MPO), malondialdehyde (MDA), superoxide dismutase (SOD), Nrf2, heme oxygenase-1 (HO-1), and. quinone oxidoreductase-1 (NQO1) of lung tissue were determined. The findings revealed that the injuries of pancreas and lung were typically induced by cerulein. The administration of BML-111 reduced the levels of serum amylase, lung MPO, lung MDA, the wet-to-dry weight ratio, and the pathology injury scores of the lung and pancreas, which increased in the SAP group. The expressions of Nrf2, HO-1, NQO1, and activity of SOD in lung tissue increased in the BML-111 group compared with those in the SAP group. This study indicates that BML-111 may play a critical protective role in APALI induced by cerulein. The underlying mechanisms of protective role may be attributable to its antioxidant effects through the activation of Nrf2/ARE pathway.

Topics: Acute Disease; Acute Lung Injury; Animals; Ceruletide; Disease Models, Animal; Heme Oxygenase-1; Heptanoic Acids; Lipopolysaccharides; Lung; Male; Malondialdehyde; Membrane Proteins; Mice; Mice, Inbred BALB C; NAD(P)H Dehydrogenase (Quinone); NF-E2-Related Factor 2; Pancreatitis; Peroxidase; Response Elements; Signal Transduction; Superoxide Dismutase

Ulinastatin is a drug used effectively to alleviate symptoms and improve the pathophysiology of various types of pancreatitis. However, the molecular mechanism responsible for its action remains unknown. Therefore, we further explore the therapeutic effects of ulinastatin and investigate possible molecular pathways modulated by this drug in the development of severe acute pancreatitis (SAP).. SAP mouse model was created by administering intraperitoneal injections of cerulein and lipopolysaccharide. Pancreatic injury was assessed by performing biochemical and histological assays and by measuring the inflammatory response of the pancreas. Specifically, we examined changes in the expression of components of the rennin-angiotensin system (RAS), including angiotensin-converting enzyme (ACE)-angiotensin II (Ang II)-angiotensin type 1 receptor (AT-1R), and ACE2-Ang-(1-7)-Mas receptor.. When SAP mouse models were treated with ulinastatin at a dosage of 50,000 U/kg body weight, we found, through biochemical and histopathological analyses, that the pancreatic injury was significantly ameliorated. Administration of ulinastatin to SAP mice led to increased expression of ACE2, Ang-(1-7), and Mas receptor, decreased expression of serum Ang II and pancreatic AT-1R, and no alterations in the expression of pancreatic ACE and Ang II when compared to cerulein-treated control group that did not receive ulinastatin.. This study shows that ulinastatin has differential effects on the two axes of the RAS during SAP. Our results further suggest that upregulation of components of the ACE2-Ang-(1-7)-Mas pathway might be an important mechanism contributing to the therapeutic role of ulinastatin in alleviating pancreatitis-associated symptoms.

Topics: Acute Disease; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Ceruletide; Disease Models, Animal; Gene Expression; Glycoproteins; Lipopolysaccharides; Mice, Inbred C57BL; Molecular Targeted Therapy; Pancreatitis; Peptide Fragments; Peptidyl-Dipeptidase A; Prospective Studies; Receptor, Angiotensin, Type 1; Renin-Angiotensin System; Severity of Illness Index

Pancreatic acinar cell necrosis is indicative of severe pancreatitis and the degree of necrosis is an index of its outcome. We studied whether the dose and duration of injury correlates with severity, particularly in terms of necrosis, in caerulein-induced acute pancreatitis (AP) in Swiss albino mice. In addition to control group 1 (G1), groups 2 and 3 received four injections of caerulein every hour but were sacrificed at five hours (G2) and nine hours (G3) respectively, and group 4 received eight injections and was sacrificed at nine hours (G4). The severity of pancreatitis was assessed histopathologically and biochemically. The histopathological scores of pancreatitis in groups 3 and 4 were significantly higher than in groups 1 and 2 (4 vs. 1, 4 vs. 2, 3 vs. 1, 3 vs. 2; P < 0.05). TUNEL-positive apoptotic cells were significantly higher in groups 2 and 3 compared with groups 1 and 4 (P < 0.05). Necrosis was significantly more in group 4 than other groups (37.49% (4.68) vs. 19.97% (1.60) in G2; 20.36% (1.56) in G3; P = 0.006 for G 2 vs. 4 and P = 0.019 for G 3 vs. 4). Electron microscopy revealed numerous autophagosomes in groups 2 and 3 and mitochondrial damage and necrosis in group 4. The pancreatic and pulmonary myeloperoxidase activity in group 4 was significantly higher than that in the other groups (P < 0.01). Hence, severity of pancreatitis is a function of the dose of injurious agent, while inflammation is both dose and duration dependent, which may also explain the wide spectrum of severity of AP seen in clinical practice.

Topics: Acute Disease; Animals; Apoptosis; Behavior, Animal; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Humans; Kidney; Lung; Male; Mice; Necrosis; Pancreas; Pancreatitis; Peroxidase; Severity of Illness Index; Time Factors

Severe acute pancreatitis is a life-threatening disease with a high mortality; so far, no causal treatment is known. The aim of this study was to evaluate the therapeutic potential of hydroxyethyl starch (HES) and cell-free hemoglobin in an experimental model.. Thirty-nine pigs were randomly assigned into 3 groups. Severe acute pancreatitis was induced by intraductal injection of glycodeoxycholic acid in combination with intravenous administration of cerulein. All animals were kept in isovolemic conditions by application of Ringer solution, 10% HES, or cell-free hemoglobin. The pancreatic microcirculation was evaluated over 8 hours. Thereafter, the animals were observed for 6 days followed by killing of the animals and histopathologic examination.. The administration of HES and cell-free hemoglobin led to improved microcirculation and tissue oxygenation compared with the Ringer's group. Consequently, the histopathologic damage was reduced (5.5 [3-8.5] vs 9.5 [7.5-11]; P < 0.001). In addition, the mean survival was significantly longer at 121 hours (95% confidence interval, 102-139) versus the Ringer group's 57 hours (95% confidence interval, 32-82; P < 0.001).. The administration of HES and cell-free hemoglobin can improve microcirculation in severe acute porcine pancreatitis, with consequent reduction in histopathologic damage and mortality. Therefore, this might represent an interesting therapeutic option in the treatment of severe acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Glycodeoxycholic Acid; Hemoglobins; Hydroxyethyl Starch Derivatives; Isotonic Solutions; Microcirculation; Oxygen Consumption; Pancreas; Pancreatitis; Random Allocation; Ringer's Solution; Severity of Illness Index; Survival Analysis; Swine; Treatment Outcome

Acute pancreatitis (AP) is a complicated disease which is largely undiscovered. Fisetin, a natural flavonoid from fruits and vegetables, has been shown to have anti-inflammatory, antioxidant, and anti-cancer activities in various disease models. However, the effects of fisetin on AP have not been determined. Pre- and post- treatment of mice with fisetin reduced the severity of AP and pancreatitis-associated lung injury and inhibited several biochemical parameters (pancreatic weight to body weight ratio, amylase, lipase, and myeloperoxidase activity) and production of inflammatory cytokines. In pancreatic acinar cells, fisetin also inhibited cell death and production of inflammatory cytokines. In addition, fisetin inhibited activation of c-Jun NH2-terminal kinase (JNK) and nuclear factor (NF)-κB in vivo and in vitro. In conclusion, these results suggest that fisetin exhibits anti-inflammatory effect on AP and could be a beneficial agent in the treatment of AP and its pulmonary complications.

Topics: Acinar Cells; Acute Disease; Administration, Oral; Animals; Ceruletide; Down-Regulation; Enzyme Activation; Female; Flavonoids; Flavonols; Injections, Intraperitoneal; JNK Mitogen-Activated Protein Kinases; Lung; Mice; Mice, Inbred C57BL; NF-kappa B; Pancreas; Pancreatitis; Signal Transduction

Acute pancreatitis (AP) is defined as an inflammatory disease of the pancreas. The purpose of this study was to examine the effectiveness of Anakinra on cerulein-induced experimental pancreatitis rat model by using the results of biochemical and histopathological findings.. Cerulein was administered to induce AP in rats. Group 1 was the sham group. Subcutancerulein was injected to the rats in group 2 for experimental pancreatitis group. In groups 3 and 4, 100 and 50 mg/kg intraperitoneal Anakinra were injected after the induction of experimental pancreatitis by subcutaneous cerulein in rats, respectively. Lastly, in group 5, rats were injected with intraperitoneal saline and subcutan cerulean for placebo group. The following parameters were evaluated: histopathological score of pancreatitis, apoptotic index, amylase, lipase, TNF-α levels, IL-1β and the leukocyte count.. When the results of serum amylase, lipase, TNF-α and IL-1β levels, the leukocyte count, histopathologic scores and apoptotic indices of control group compared to the results of other groups, the differences exhibited statistical significance (all p < 0.05). On the other hand, when the results of fourth group compared with the results of third group, the data demonstrated statistical insignificance (p > 0.05). However, no any significant differences were found between the results of fourth and fifth groups (p > 0.05).. In the light of these results, cerulein is an appropriate agent for experimental AP rat model and Anakinra has a favorable therapeutic effect on acute experimental pancreatitis model. Moreover, Anakinra significantly decreases cerulein-related pancreatic tissue injury and pancreatic apoptosis.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Ceruletide; Disease Models, Animal; Interleukin 1 Receptor Antagonist Protein; Interleukin-1beta; Leukocyte Count; Lipase; Male; Pancreatitis; Rats; Rats, Wistar; Tumor Necrosis Factor-alpha

Diosmetin (3', 5, 7-trihydroxy-4'-methoxyflavone), the aglycone part of the flavonoid glycosides diosmin occurs naturally in citrus fruit, was considered to exhibit anti-inflammatory and antioxidant properties. Our study aimed to investigate the effect of diosmetin in a murine model of cerulein-induced acute pancreatitis (AP). Experimental AP was induced in mice by seven intraperitoneal injection of cerulein (50 ug/kg) at hourly intervals. Diosmetin (100 mg/kg) or vehicle was pretreated 2 h before the first cerulein injection. After 6 h, 9 h, 12 h of the first cerulein injection, the severity of acute pancreatitis was evaluated biochemically and morphologically. Pretreatment with diosmetin significantly reduced serum levels of amylase and lipase; the histological injury; the secretion of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6; myeloperoxidase (MPO) activity, trypsinogen activation peptide (TAP) level, the expression of inducible nitric oxide synthase (iNOS); and the nuclear factor (NF)-κB activation in cerulein-induced AP. This study showed that administration of diosmetin demonstrated a beneficial effect on the course of cerulein-induced AP in mice. Therefore, diosmetin may become a new therapeutic agent in future clinical trials for treatment of AP.

Topics: Active Transport, Cell Nucleus; Acute Disease; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Cytoprotection; Disease Models, Animal; Flavonoids; Inflammation Mediators; Male; Mice, Inbred C57BL; NF-kappa B; Oxidative Stress; Pancreas; Pancreatitis; Severity of Illness Index; Signal Transduction; Time Factors

To investigate the migratory path of stem cells in pancreatic tissues damaged by pancreatitis and to preliminarily identify stem cells that efficiently contribute to the repair of damaged pancreatic tissues.. An animal model of acute pancreatitis was established, in which rats in the experimental group were given intraperitoneal (IP) injections of caerulein. Before the rats were sacrificed, 5-bromo-2'-deoxyuridine (BrdU) was administered by IP injection to label proliferating pancreatic cells. The localization and distribution of the stem cell-specific marker proteins nestin and c-kit in pancreatic tissues were examined using an immunohistochemical approach, and proliferation-specific BrdU incorporation was also analyzed.. (1) The nestin-positive cells first appeared in the pancreatic interlobar vessels, and then, were observed in the pancreatic acinar and islet tissues. (2) C-kit-positive cells were located only in the pancreatic islets. (3) BrdU-positive cells first appeared in the area surrounding the interlobular region, and then were diffusely distributed and filled the pancreatic lobules.. (1) The stem cells, participated in the repair of damaged pancreatic tissue, appear firstly in the pancreatic interlobar vessels, then migrate toward the pancreatic lobules by using the interlobar vessels as channels and penetrate through the vascular endothelium into the pancreatic acinar tissues. A portion of the stem cells eventually penetrate into the islet tissue. (2) Exogenous stem cells, rather than the tissue-resident stem cells, efficiently contribute to the repair of damaged pancreatic tissues.

Topics: Acute Disease; Animals; Biomarkers; Cell Movement; Cell Proliferation; Ceruletide; Disease Models, Animal; Female; Immunohistochemistry; Male; Nestin; Pancreas; Pancreatitis; Proto-Oncogene Proteins c-kit; Rats, Sprague-Dawley; Stem Cells; Time Factors; Wound Healing

The gene p8 was initially described in pancreatic tissue during acute experimental pancreatitis, a disease that is characterized by a systemic immune response. Although early reports suggested that p8 affects leukocyte migration during acute pancreatitis (AP), no studies revealing its immune-modulatory effects have been performed.. We investigated the composition of the cellular immune system in naive p8 knockout (p8(−/−)) mice and compared with matched wild-type mice during pancreatitis.. In young mice, there were no relevant differences in the composition of peripheral and splenic CD3(+), CD3(+)CD4(+), CD3(+)CD8(+), CD11b(+)Gr-1(-), and Gr-1 cells. In mature p8(−/−) mice, increased splenic CD4CD25FoxP3 cells, spleen siderosis, and increased marginal zones in the splenic white pulp were found. During AP, peripheral and splenic CD3(+) and CD3CD4 declined stronger in the p8(−/−) mice. The spleen of the p8(−/−) mice showed severe hypoplasia of the white pulp and mild hyperplasia of the red pulp. This was associated with a significantly increased rate of apoptosis.. We conclude that p8 has no impact on the cellular composition of the adaptive and innate immune systems in noninflammatory conditions. However, it may limit apoptosis and maintain homeostasis of the immune reaction during AP.

Topics: Acute Disease; Animals; Apoptosis; Cell Count; Ceruletide; DNA-Binding Proteins; Female; Hemosiderosis; Lymphocyte Count; Lymphocyte Subsets; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myeloid Cells; Neoplasm Proteins; Organ Specificity; Pancreatitis; Splenic Diseases; Splenomegaly

Oxidative stress is recognized as a pivotal effector of several pathogenic processes, including acute pancreatitis. Reactive oxygen species not just cause damage on the main cellular components, but also influence the expression of antioxidant system genes. Antioxidant molecules, such as melatonin, could be good candidates for the treatment of this multidimensional disease. The present study was to evaluate the chemopreventive effect of melatonin in a rat model of cerulein-induced acute pancreatitis.. Four subcutaneous injections of cerulein (20 μg/kg body weight) were given to Wistar rats at two hours intervals; melatonin was injected intraperitoneally (25 mg/kg body weight) 30 minutes before each injection of cerulein. Lipid peroxidation, protein oxidation (carbonyl groups), total antioxidant status, and glutathione peroxidase activity were determined in pancreatic tissue using commercial kits.. The chemopreventive administration of melatonin caused a reduction in lipid peroxidation and protein oxidation due to injections of cerulein. Additionally, melatonin treatment was also able to revert glutathione peroxidase activity and total antioxidant status near to control levels, suggesting that melatonin could prevent from oxidative phenomena in the pancreas, such as lipid peroxidation and protein oxidation, and could stimulate, directly or indirectly, the expression of antioxidant enzymes.. Melatonin, a polyvalent antioxidant, protected the pancreatic damage via the decrease of oxidative stress and increase of the activities of antioxidant enzymes in cerulein-induced acute pancreatitis.

Topics: Acute Disease; Animals; Antioxidants; Ceruletide; Cytoprotection; Disease Models, Animal; Female; Glutathione Peroxidase; Lipid Peroxidation; Male; Melatonin; Oxidative Stress; Pancreas; Pancreatitis; Protein Carbonylation; Rats, Wistar

Acute pancreatitis (AP) is a frequent gastrointestinal disorder that causes significant morbidity, and its incidence has been progressively increasing. AP starts as a local inflammation in the pancreas that often leads to systemic inflammatory response and complications. Soluble epoxide hydrolase (sEH) is a cytosolic enzyme whose inhibition in murine models has beneficial effects in inflammatory diseases, but its significance in AP remains unexplored.. To investigate whether sEH may have a causal role in AP we utilized Ephx2 knockout (KO) mice to determine the effects of sEH deficiency on cerulein- and arginine-induced AP. sEH expression increased at the protein and messenger RNA levels, as well as enzymatic activity in the early phase of cerulein- and arginine-induced AP in mice. In addition, amylase and lipase levels were lower in cerulein-treated Ephx2 KO mice compared with controls. Moreover, pancreatic mRNA and serum concentrations of the inflammatory cytokines IL-1B and IL-6 were lower in cerulein-treated Ephx2 KO mice compared with controls. Further, Ephx2 KO mice exhibited decreased cerulein- and arginine-induced NF-κB inflammatory response, MAPKs activation and decreased cell death. Conclusions -These findings demonstrate a novel role for sEH in the progression of cerulein- and arginine-induced AP.

Topics: Acute Disease; Animals; Cell Death; Ceruletide; Disease Models, Animal; Epoxide Hydrolases; Gene Expression; Male; MAP Kinase Signaling System; Mice; Mice, Knockout; NF-kappa B; Pancreatitis

Severe acute pancreatitis (SAP) is a serious systemic disease with a sustained high mortality rate. Extensive evidence has shown that gut barrier dysfunction plays a critical role in the pathophysiology of SAP.. Investigating the role of intestinal mucosa oxidative stress in gut barrier dysfunction of SAP.. Twenty-four BALB/c mice were randomly divided into two groups with twelve mice each group. The SAP group mice received six intraperitoneal injections of cerulein (50 µg/kg) at 1-hour intervals, then given one intraperitoneal injection of 10 mg/kg lipopolysaccharide (LPS from E. coli) for inducing SAP. Normal saline was given to the mice of control group. The animals of each group were averaged to two batches. Four and eight hours after the final injection, respectively, mice were anesthetized and blood and tissue samples were harvested for examination. The pathological changes of pancreas and gut were observed and scored. The serum levels of diamine oxidase (DAO), amylase and tumor necrosis factor-alpha (TNF-α) were measured. The contents of malondialdehyde (MDA) and reduced glutathione (GSH) and activity of superoxide dismutase (SOD) and xanthine oxidase (XO) in gut mucosa were detected. In gut mucosa, the caspase-3 activity was measured and the cell apoptosis and apoptosis index (AI) were determined by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. The data were analyzed by ANOVA and t-test.. At four and eight hours after SAP induction, the SAP group mice had significantly higher pancreatic and gut pathological scores (p < 0.01) and increased serum levels of amylase (p < 0.05), DAO and TNF-α (p < 0.01) and increased MDA contents and XO activity of gut mucosa (p < 0.01) compared with those of control mice. There were significantly lower GSH contents (p < 0.05) and SOD activity (p < 0.01) of gut mucosa in the SAP mice. It was also observed that the gut mucosa cells of SAP mice had significantly higher caspase-3 activity and apoptosis index (p < 0.01).. In SAP, waterfall-style release of inflammatory factors such as TNF-α led to ischemia-reperfusion injury of gut mucosa which resulted in serious oxidative stress and activation of caspase-3 pathway and severe apoptosis of gut mucosa. Therefore, intestinal mucosal oxidative stress may play an important role in the mechanism of gut barrier dysfunction.

Topics: Acute Disease; Analysis of Variance; Animals; Apoptosis; Caspase 3; Ceruletide; Disease Models, Animal; In Situ Nick-End Labeling; Inflammation Mediators; Intestinal Mucosa; Male; Mice; Mice, Inbred BALB C; Oxidative Stress; Pancreatitis; Random Allocation; Reperfusion Injury; Severity of Illness Index; Tumor Necrosis Factor-alpha

Glucagon-like peptide-1 (GLP-1) promotes insulin release; however, the relationship between the GLP-1 signal and chronic pancreatitis is not well understood. Here we focus on chemokine (C-C motif) ligand 2 (CCL2) and its receptor (CCR2) axis, which regulates various immune cells, including macrophages, to clarify the mechanism of GLP-1-mediated insulin secretion in chronic pancreatitis in mice. One and multiple series of repetitive cerulein administrations were used to induce acute and chronic cerulein pancreatitis, respectively. Acute cerulein-administered CCR2-knockout (KO) mice showed suppressed infiltration of CD11b(+)Gr-1(low) macrophages and pancreatic inflammation and significantly upregulated insulin secretion compared with paired wild-type (WT) mice. However, chronic cerulein-administered CCR2-KO mice showed significantly increased infiltration of CD11b(+)/Gr-1(-) and CD11b(+)/Gr-1(high) cells, but not CD11b(+)/Gr-1(low) cells, in pancreas with severe inflammation and significantly decreased insulin secretion compared with their WT counterparts. Furthermore, although serum GLP-1 levels in chronic cerulein-administered WT and CCR2-KO mice were comparably upregulated after cerulein administrations, GLP-1 receptor levels in pancreases of chronic cerulein-administered CCR2-KO mice were significantly lower than in paired WT mice. Nevertheless, a significantly higher hyperglycemia level in chronic cerulein-administered CCR2-KO mice was markedly restored by treatment with a GLP-1 analog to a level comparable to the paired WT mice. Collectively, the CCR2/CCL2 axis-mediated CD11b(+)-cell migration to the pancreas is critically involved in chronic pancreatitis-mediated hyperglycemia through the modulation of GLP-1 receptor expression and insulin secretion.

Topics: Acute Disease; Animals; CD11b Antigen; Ceruletide; Chronic Disease; Disease Models, Animal; Female; Glucagon-Like Peptide-1 Receptor; Glucose Intolerance; Hyperglycemia; Insulin; Insulin Secretion; Islets of Langerhans; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreatitis, Chronic; Receptors, CCR2; Receptors, Glucagon

To evaluate the inhibitory effects of Scolopendra subspinipes mutilans (SSM) on cerulein-induced acute pancreatitis (AP) in a mouse model.. SSM water extract (0.1, 0.5, or 1 g/kg) was administrated intraperitoneally 1 h prior to the first injection of cerulein. Once AP developed, the stable cholecystokinin analogue, cerulein was injected hourly, over a 6 h period. Blood samples were taken 6 h later to determine serum amylase, lipase, and cytokine levels. The pancreas and lungs were rapidly removed for morphological examination, myeloperoxidase assay, and real-time reverse transcription polymerase chain reaction. To specify the role of SSM in pancreatitis, the pancreatic acinar cells were isolated using collagenase method. Then the cells were pre-treated with SSM, then stimulated with cerulein. The cell viability, cytokine productions and high-mobility group box protein-1 (HMGB-1) were measured. Furthermore, the regulating mechanisms of SSM action were evaluated.. The administration of SSM significantly attenuated the severity of pancreatitis and pancreatitis associated lung injury, as was shown by the reduction in pancreatic edema, neutrophil infiltration, vacuolization and necrosis. SSM treatment also reduced pancreatic weight/body weight ratio, serum amylase, lipase and cytokine levels, and mRNA expression of multiple inflammatory mediators such as tumor necrosis factor-α and interleukin-1β. In addition, treatment with SSM inhibited HMGB-1 expression in the pancreas during AP. In accordance with in vivo data, SSM inhibited the cerulein-induced acinar cell death, cytokine, and HMGB-1 release. SSM also inhibited the activation of c-Jun NH2-terminal kinase, p38 and nuclear factor (NF)-κB.. These results suggest that SSM plays a protective role during the development of AP and pancreatitis associated lung injury via deactivating c-Jun NH2-terminal kinase, p38 and NF-κB.

Topics: Acute Disease; Acute Lung Injury; Amylases; Animals; Anti-Inflammatory Agents; Arthropod Venoms; Cell Survival; Cells, Cultured; Ceruletide; Cytokines; Disease Models, Animal; Enzyme Activation; HMGB1 Protein; Inflammation Mediators; JNK Mitogen-Activated Protein Kinases; Lipase; Mice; Mice, Inbred C57BL; NF-kappa B; p38 Mitogen-Activated Protein Kinases; Pancreas; Pancreatitis; Signal Transduction; Time Factors

We have previously reported that bee venom (BV) has a protective role against acute pancreatitis (AP). However, the effects of apamin, the major compound of BV, on AP have not been determined. The aim of this study was to evaluate the effects of apamin on cerulein-induced AP.. AP was induced via intraperitoneal injection of supramaximal concentrations of the stable cholecystokinin analogue cerulein (50 μg/kg) every hour for 6 times. In the apamin treatment group, apamin was administered subcutaneously (10, 50, or 100 μg/kg) at both 18 and 1 h before the first cerulein injection. The mice were sacrificed at 6 h after the final cerulein injection. Blood samples were obtained to determine serum amylase and lipase levels, as well as cytokine production. The pancreas and lung were rapidly removed for morphologic and histological examination, myeloperoxidase (MPO) assay, and real-time reverse transcription-polymerase chain reaction. Furthermore, we isolated the pancreatic acinar cells to specify the role of apamin in AP.. Pre-treatment with apamin inhibited histological damage, pancreatic weight/body weight ratio, serum level of amylase and lipase, MPO activity, and cytokine production. In addition, apamin treatment significantly inhibited cerulein-induced pancreatic acinar cell death. Furthermore, apamin treatment inhibited the cerulein-induced activation of c-Jun NH2-terminal kinases (JNK).. These results could suggest that apamin could protect against AP by inhibition of JNK activation.

Topics: Acute Disease; Animals; Apamin; Ceruletide; Cholecystokinin; Cytokines; Disease Models, Animal; Injections, Intraperitoneal; Injections, Subcutaneous; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mitogen-Activated Protein Kinase Kinases; NF-kappa B; Pancreas; Pancreatitis

To investigate the pathophysiological role of C/EBP homologous protein (CHOP) in severe acute pancreatitis and associated lung injury.. A severe acute pancreatitis model was induced with 6 injections of cerulein (Cn, 50 μg/kg) at 1-h intervals, then intraperitoneal injection of lipopolysaccharide (LPS, 7.5 mg/kg) in CHOP-deficient (Chop(-/-)) mice and wild-type (WT) mice. Animals were sacrificed under anesthesia, 3 h or 18 h after LPS injection. Serum amylase, lipase, and cytokines [interleukin (IL)-6 and tumor necrosis factor (TNF)-α], pathological changes, acute lung injury, and apoptosis in the pancreas were evaluated. Serum amylase and lipase activities were detected using a medical automatic chemical analyzer. Enzyme-linked immunosorbent assay kits were used to evaluate TNF-α and IL-6 levels in mouse serum and lung tissue homogenates. Apoptotic cells in sections of pancreatic tissues were determined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) analysis. The mouse carotid arteries were cannulated and arterial blood samples were collected for PaO2 analysis. The oxygenation index was expressed as PaO2/FiO2.. Administration of Cn and LPS for 9 and 24 h induced severe acute pancreatitis in Chop(-/-) and WT mice. When comparing Chop(-/-) mice and WT mice, we observed that CHOP-deficient mice had greater increases in serum TNF-α (214.40 ± 19.52 pg/mL vs 150.40 ± 16.70 pg/mL; P = 0.037), amylase (4236.40 ± 646.32 U/L vs 2535.30 ± 81.83 U/L; P = 0.041), lipase (1678.20 ± 170.57 U/L vs 1046.21 ± 35.37 U/L; P = 0.008), and IL-6 (2054.44 ± 293.81 pg/mL vs 1316.10 ± 108.74 pg/mL; P = 0.046) than WT mice. The histopathological changes in the pancreases and lungs, decreased PaO2/FiO2 ratio, and increased TNF-α and IL-6 levels in the lungs were greater in Chop(-/-) mice than in WT mice (pancreas: Chop(-/-) vs WT mice, hemorrhage, P = 0.005; edema, P = 0.005; inflammatory cells infiltration, P = 0.005; total scores, P = 0.006; lung: hemorrhage, P = 0.017; edema, P = 0.017; congestion, P = 0.017; neutrophil infiltration, P = 0.005, total scores, P = 0.001; PaO2/FiO2 ratio: 393 ± 17.65 vs 453.8, P = 0.041; TNF-α: P = 0.043; IL-6, P = 0.040). Results from TUNEL analysis indicated increased acinar cell apoptosis in mice following the induction of acute pancreatitis. However, Chop(-/-) mice displayed significantly reduced pancreatic apoptosis compared with the WT mice (201.50 ± 31.43 vs 367.00 ± 47.88, P = 0.016).. These results suggest that CHOP can exert protective effects against acute pancreatitis and limit the spread of inflammatory damage to the lungs.

Topics: Acute Disease; Acute Lung Injury; Amylases; Animals; Apoptosis; Biomarkers; Ceruletide; Disease Models, Animal; Inflammation Mediators; Interleukin-6; Lipase; Lipopolysaccharides; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis; Severity of Illness Index; Transcription Factor CHOP; Tumor Necrosis Factor-alpha

To investigate whether miRNA-155 (miR-155) dysregulates apical junctional complex (AJC) protein expression in experimental severe acute pancreatitis (SAP).. Twenty-four male BALB/c mice were randomly assigned to two groups: the SAP group (n = 12) receiving sequential intraperitoneal injection of 50 µg/kg caerulein and 10 mg/kg lipopolysaccharide over 6 h, and the control group (n = 12) receiving intraperitoneal injection of normal saline. Animals were sacrificed 3 h following the last injection for collection of blood samples and pancreas and distal ileal segment specimens. Routine pancreas and intestine histology was used to assess SAP pathology and intestinal epithelial barrier damage. Levels of serum amylase, diamine oxidase (DAO), and tumor necrosis factor (TNF)-α were determined using commercial kits. Total RNA samples were isolated from intestinal epithelial specimens and reversely transcribed into cDNA. miR-155 and RhoA mRNA expression profiles were determined using quantitative real-time polymerase chain reaction. Target genes for miR-155 were predicted using the miRTarBase database, RNA22 and PicTar computational methods. Western blotting was performed to quantitate the protein expression levels of the target gene RhoA, as well as zonula occludens (ZO)-1 and E-cadherin, two AJC component proteins.. Intraperitoneal injection of caerulein and lipopolysaccharide successfully induced experimental acute pancreatic damage (SAP vs control, 10.0 ± 2.0 vs 3.2 ± 1.2, P < 0.01) and intestinal epithelial barrier damage (3.2 ± 0.7 vs 1.4 ± 0.7, P < 0.01). Levels of serum amylase (21.6 ± 5.1 U/mL vs 14.3 ± 4.2 U/mL, P < 0.01), DAO (21.4 ± 4.1 mg/mL vs 2.6 ± 0.8 mg/mL, P < 0.01), and TNF-α (61.0 ± 15.1 ng/mL vs 42.9 ± 13.9 ng/mL, P < 0.01) increased significantly in SAP mice compared to those in control mice. miR-155 was significantly overexpressed in SAP intestinal epithelia (1.94 ± 0.50 fold vs 1.03 ± 0.23 fold, P < 0.01), and RhoA gene containing three miR-155-specific binding sites in the three prime untranslated regions was one of the target genes for miR-155. RhoA (22.7 ± 5.8 folds vs 59.6 ± 11.6 folds, P < 0.01), ZO-1 (46 ± 18 folds vs 68 ± 19 folds, P < 0.01), and E-cadherin proteins (48 ± 15 folds vs 77 ± 18 folds, P < 0.01) were underexpressed in SAP intestinal epithelia although RhoA mRNA expression was not significantly changed in SAP (0.97 ± 0.18 folds vs 1.01 ± 0.17 folds, P > 0.05).. TNF-α-regulated miR-155 overexpression inhibits AJC component protein syntheses of ZO-1, and E-cadherin by downregulating post-transcriptional RhoA expression, and disrupts intestinal epithelial barrier in experimental SAP.

Topics: Acute Disease; Amine Oxidase (Copper-Containing); Amylases; Animals; Cadherins; Ceruletide; Disease Models, Animal; Epithelial Cells; Ileum; Intestinal Mucosa; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; MicroRNAs; Pancreatitis; Permeability; rho GTP-Binding Proteins; rhoA GTP-Binding Protein; RNA, Messenger; Severity of Illness Index; Tight Junctions; Tumor Necrosis Factor-alpha; Up-Regulation; Zonula Occludens-1 Protein

Acute pancreatitis accounts for almost 250.000 hospital admissions annually in the United States. Most promising treatment approaches are preventive; however, little is known about the early factors initiating acute pancreatitis. We aimed to evaluate the preventive effects of enoxaparin and hesperidin in cerulein-induced acute pancreatitis.. We used 70 Wistar albino rats for this study. Rats were divided into 7 groups: control group, and groups that were administered cerulein(Group 2), enoxaparin (Group 3), hesperidin (Group 4), cerulein with enoxaparin (Group 5), cerulein with hesperidin (Group 6), and cerulein with both enoxaparin and hesperidin (Group 7). Edema formation; leukocyte infiltration; measurement of the amylase level, pancreatic tissue weight, and pancreatic tissue oxidative capacity; and chemiluminescence using luminol, lucigenin, and nitric oxide levels as indices of tissue oxidative capacity were used to evaluate pancreatitis.. Acute edematous mild pancreatitis was induced in groups 2, 5, and 6 by cerulein injections. Enoxaparin and hesperidin significantly decreased (p < 0.001) all the tested parameters in these rats. Enoxaparin and hesperidin did not offer complete protection but showed 50% decrease in edema formation. The preventive agents showed no superiority to each other. Further, when enoxaparin and hesperidin were used in combination, no significant additive effects with regard to anti-inflammatory and anti-oxidative actions were present.. We showed that both enoxaparin and hesperidin exerted significant preventive effects in all the parameters related to acute pancreatitis in our experimental rat model.

Topics: Acute Disease; Amylases; Animals; Anticoagulants; Antioxidants; Ceruletide; Edema; Enoxaparin; Hesperidin; Male; Neutrophil Infiltration; Nitric Oxide; Pancreas; Pancreatitis; Rats, Wistar; Reactive Oxygen Species

In acute pancreatitis (AP), inflammatory cells and products disseminated in abdominal lymph and blood induce systemic inflammation. Interruption of abdominal lymph flow, and thereby reduction of lymphatic dissemination, could alter the course of the disease. Therefore, we investigated whether thoracic duct ligation (TDL) in a rat model of cerulein-induced AP results in reduced lung damage as a marker for reduction of systemic dissemination through the lymphatic system. Thirty-four male rats were assigned to TDL (TDL-rats, n=8), AP (AP-rats, n=8), TDL+AP (TDL+AP-rats, n=9) or sham TDL (Ctr-rats, n=9) groups. TDL and sham TDL were established first. Two days later, AP was induced in AP- and TDL+AP-rats by a series of subcutaneous injections of cerulein. Vehicle was injected in the same manner in Ctr- and TDL-rats as controls. Rats were sacrificed six hours after the end of the serial injections. Histological examination showed that AP-induced damage to the pancreas and ileum were similar in AP- and TDL+AP-rats whereas lung damage was less severe in TDL+AP-rats than in AP-rats. Assays demonstrated that: hepatic and pulmonary myeloperoxidase activities were increased in AP-rats but not in the TDL+AP-rats; more Il-6 was found in AP-rat than TDL+AP-rat lungs; and lung-lavage fluid from AP-rats yielded more angiopoietin-2 than TDL+AP-rats. In conclusion, prior TDL in the rat attenuates lung damage in acute pancreatitis.

Topics: Acute Disease; Acute Lung Injury; Animals; Ceruletide; Ligation; Male; Pancreatitis; Rats; Rats, Wistar; Thoracic Duct

The purpose of our study was to evaluate the protective effect of melatonin in a rat model of caerulein-induced acute pancreatitis. For the induction of experimental acute pancreatitis, four subcutaneous injections of caerulein (20 mgkg–1 body weight) were given to Wistar rats at 2-h intervals. Melatonin was injected intraperitoneally (25 mg kg–1 body weight) 30 min before each caerulein injection. After 12 h, rats were sacrificed by decapitation. Blood and pancreas samples were collected and processed for serological and histopathological studies,respectively. Lipase, a-amylase, corticosterone, total antioxidant power and cytokines interleukin (IL)-1b, IL-4 and tumour necrosis factor(TNF)-a were determined using commercial kits. ANOVA and Tukey tests (P<0.05) were performed for the statistical analysis of the results.Results showed that the administration of melatonin reduced histological damage induced by caerulein treatment as well as the hyperamylasemia and hyperlipidemia. Corticosterone and antioxidant total power were also reverted to basal activities. Furthermore, melatonin pre-treatment reduced pro-inflammatory cytokines IL-1b and TNF-a and increased the serum levels of anti-inflammatory cytokine IL-4. In conclusion,the findings suggest that the protective effect of melatonin in caerulein-induced acute pancreatitis is mediated by the anti-inflammatory ability of this indolamine. Thus, melatonin may have a protective effect against acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Antioxidants; Ceruletide; Cytokines; Female; Lipase; Male; Melatonin; Pancreatitis; Rats; Rats, Wistar

The synthetic tripeptide feG (D-Phe-D-Glu-Gly) is a novel pharmacologic agent that decreases neutrophil recruitment, infiltration, and activation in various animal models of inflammatory disease. We aimed to investigate the effect of feG as both a preventive treatment when administered before acute lung injury and as a therapeutic treatment administered following initiation of acute lung injury.. Lung injury was assessed following prophylactic or therapeutic intratracheal feG administration in a “two-hit” rodent model of acute pancreatitis plus intratracheal lipopolysaccharide.. Following both prophylactic and therapeutic feG administration, there were significant improvements in arterial blood oxygenation and respiratory mechanics and decreased lung edema, BAL protein concentration, histologic tissue injury scores, BAL cell infiltration, and lung myeloperoxidase activity. Most indices of lung damage were reduced to baseline control values.. feG reduced leukocyte infiltration, ameliorated the severity of inflammatory damage, and restored lung function when administered either prophylactically or therapeutically in a two-hit rat model of acute pancreatitis plus intratracheal lipopolysaccharide.

Topics: Acute Disease; Acute Lung Injury; Animals; Cell Movement; Ceruletide; Disease Models, Animal; Male; Neutrophils; Oligopeptides; Pancreatitis; Rats; Rats, Sprague-Dawley; Respiratory Mechanics; Severity of Illness Index; Treatment Outcome

The anti-inflammatory effects of O-1602 and cannabidiol (CBD), the ligands of G protein-coupled receptor 55 (GPR55), on experimental acute pancreatitis (AP) were investigated.. Acute pancreatitis was induced in C57BL mice by intraperitoneal injection of 50 μg/kg cerulein hourly, with a total of 6 times. Drugs (O-1602, 10 mg/kg, or CBD, 0.5 mg/kg) were given by intraperitoneal injection 2 times at 30 minutes before the first injection and immediately before the fifth cerulein injection. At 3 hours after the last injection, the blood, the lungs, and the pancreas were harvested for the pancreatic enzyme activity, myeloperoxidase activity, and pro-inflammatory cytokines measurement; and the expressions of GPR55 mRNA and protein in the pancreas were detected.. Cannabidiol or O-1602 treatment significantly improved the pathological changes of mice with AP and decreased the enzyme activities, IL-6 and tumor necrosis factor α; levels, and the myeloperoxidase activities in plasma and in the organ tissues. G protein-coupled receptor 55 mRNA and protein expressed in the pancreatic tissue, and the expressions were decreased in the mice with AP, and either CBD or O-1602 attenuated these changes to a certain extent.. Cannabidiol and O-1602 showed anti-inflammatory effects in mice with AP and improved the expression of GPR55 in the pancreatic tissue as well.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Blotting, Western; Cannabidiol; Ceruletide; Disease Models, Animal; Immunohistochemistry; Inflammation Mediators; Injections, Intraperitoneal; Interleukin-6; Lipase; Lung; Mice; Mice, Inbred C57BL; Pancreas; Pancreatitis; Peroxidase; Real-Time Polymerase Chain Reaction; Receptors, Cannabinoid; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors; Tumor Necrosis Factor-alpha

To investigate the role of bone morphogenetic protein (BMP) signaling in acute pancreatitis (AP) by administration of noggin, an endogenous BMP antagonist, in a cerulein-induced AP model.. Acute pancreatitis was induced by 9 hourly intraperitoneal injections of cerulein (50 μg/kg). Control mice received phosphate-buffered saline injections. In a separate group, noggin (0.5 mg/kg) was given intraperitoneally at 1 hour before and 2, 4, and 6 hours after AP induction. The mice were euthanized at 1 hour after completion of AP induction. The blood samples and the pancreas were harvested for analysis. Isolated pancreatic acini from normal mice and AR42J cells were treated with BMP2 and cerulein. AR42J cells were also treated with noggin. Phosphorylation of Smad1/5/8 was measured.. Bone morphogenetic protein signaling was up-regulated in AP mouse pancreas. Bone morphogenetic protein 2 and cerulein-induced phosphorylation of Smad1/5/8 in the acinar cells in vitro, which was blocked by noggin. Noggin administration in vivo attenuated AP induction, decreased vacuole formation in acinar cells, blocked LC3-II levels, and partially restored Beclin-1 and lysosomal-associated membrane protein 2 levels.. Bone morphogenetic protein signaling seems to promote AP induction and autophagy, as suggested by our study showing that noggin ameliorates AP and partially restores autophagic homeostasis.

Topics: Acute Disease; Animals; Apoptosis Regulatory Proteins; Autophagy; Beclin-1; Bone Morphogenetic Protein 2; Carrier Proteins; Cell Line; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Injections, Intraperitoneal; Lysosomal-Associated Membrane Protein 2; Male; Mice; Mice, Inbred C57BL; Microtubule-Associated Proteins; Pancreas; Pancreatitis; Phosphorylation; Rats; Recombinant Proteins; Signal Transduction; Smad1 Protein; Smad5 Protein; Smad8 Protein; Time Factors

To examine the effect of inflexinol on the development of acute pancreatitis (AP) and to investigate the mechanisms responsible for the protective effect against AP.. Acute pancreatitis was induced in mice by intraperitoneal injection of cerulein. Inflexinol was administered intraperitoneally 4 times every 6 hours from 1 hour before the first cerulein injection. Serum amylase activity and histology of the pancreas were measured. Determination of pancreatic nuclear factor-κB (NF-κB) p65 expression was conducted by Western blotting and immunohistochemistry to investigate the mechanisms responsible for the inflexinol effects.. Serum amylase activity in the cerulein group was significantly higher than that in the control group (P < 0.05). Pancreatic histology revealed marked inflammatory changes in the cerulein group such as interstitial edema, vacuolization, necrosis, and infiltration of inflammatory cells; and Western blotting and immunohistochemistry showed marked NF-κB p65 expression. Treatment with inflexinol significantly attenuated the inflammatory changes in pancreatic histology at 24, 48, and 72 hours (P < 0.05). Pancreatic NF-κB p65 expression decreased significantly after inflexinol treatment (P < 0.05).. Inflexinol reduced the severity of cerulein-induced AP by inhibiting NF-κB activation.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Biomarkers; Blotting, Western; Ceruletide; Disease Models, Animal; Diterpenes, Kaurane; Female; Immunohistochemistry; Mice; Mice, Inbred BALB C; Pancreas; Pancreatitis; Severity of Illness Index; Time Factors; Transcription Factor RelA

Our aim in the present study was to investigate the potential roles of the 78-kDa glucose-regulated protein (GRP78) and the X-linked inhibitor of apoptosis protein (XIAP) in the regulation of apoptosis during cerulein-induced acute pancreatitis (CAP). A rat CAP model was induced by injection of cerulein (50 µg/kg), and the severity of CAP was estimated by measuring serum amylase and lipase, pancreatic edema and histological changes. Pancreatic acinar cell apoptosis was determined by terminal-deoxynucleotidyl-transferase-mediated dUTP nick-end labeling (TUNEL) assay, and the expression of GRP78, XIAP and the apoptotic genes caspase-3, -7 and -9 were determined by real‑time quantitative PCR and western blotting. After induction with cerulein, increased serum amylase and lipase, pancreatic edema, inflammation and apoptosis were observed in CAP rats. Furthermore, the mRNA and protein levels of GRP78 and XIAP were significantly downregulated in CAP rats, while the mRNA levels of caspase-3, -7 and -9, as well as the cell apoptotic index were markedly increased when compared with control rats (P<0.05). The expression of GRP78 and XIAP was negatively correlated with caspase expression in CAP (P<0.05). This study suggests that the downregulation of GRP78 and XIAP were correlated with apoptosis in pancreatic acinar cells, and that this may occur through the regulation of caspase activation during CAP.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Caspase 3; Caspase 7; Caspase 9; Ceruletide; Disease Models, Animal; Down-Regulation; Endoplasmic Reticulum Chaperone BiP; Heat-Shock Proteins; Lipase; Male; Pancreatitis; Rats; Rats, Wistar; RNA, Messenger; X-Linked Inhibitor of Apoptosis Protein

Acute pancreatitis is a potentially fatal disease with no known cure. The initial events in acute pancreatitis may occur within the acinar cells. We examined the effect of sesamol on (i) a cerulein-induced pancreatic acinar cancer cell line, AR42J, and (ii) cerulein-induced experimental acute pancreatitis in rats. Sesamol inhibited amylase activity and increased cell survival. It also inhibited medium lipid peroxidation and 8-hydroxydeoxyguanosine in AR42J cells compared with the cerulein-alone groups. In addition, in cerulein-treated rats, sesamol inhibited serum amylase and lipase levels, pancreatic edema, and lipid peroxidation, but it increased pancreatic glutathione and nitric oxide levels. Thus, we hypothesize that sesamol attenuates cerulein-induced experimental acute pancreatitis by inhibiting the pancreatic acinar cell death associated with oxidative stress in rats.

Topics: Acute Disease; Amylases; Animals; Antioxidants; Benzodioxoles; Cell Line, Tumor; Cell Survival; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Lipid Peroxidation; Male; Mice; Oxidative Stress; Pancreatitis; Phenols; Rats; Rats, Wistar

The purpose of our study was to evaluate the protective effect of the strong antioxidant and anti-inflammatory agent, lycopene, on oxidative stress in a rat model of cerulein-induced acute edematous pancreatitis.. Sprague-Dawley rats were pretreated with lycopene (50 mg/kg, i.p.) or saline 15 min before cerulein was given 20 μg/kg (i.p.) at 1-h intervals within 4 h. Twelve hours after cerulein or saline injections, the animals were killed by decapitation. Blood samples were collected to analyze amylase, lipase, and proinflammatory cytokines (TNF-α and IL-1ß). Pancreatic tissues were taken for the determination of tissue glutathione (GSH) and malondialdehyde (MDA) levels, Na(+)/K(+)-ATPase, and myeloperoxidase (MPO) activities. Tissue samples were also examined histologically.. Acute pancreatitis caused significant decrease in tissue GSH levels and Na(+)/K(+)-ATPase activity, while pancreatic MDA levels and MPO activity were increased. Furthermore, TNF-α, IL-1ß, and amylase lipase levels were also significantly increased. On the other hand, lycopene pretreatment reserved all these biochemical indices as well as histopathologic alterations that were induced by cerulein.. According to the results, lycopene protects the pancreatic tissues from oxidative damage induced by cerulein, and this effect possibly involves the inhibition of neutrophil infiltration and lipid peroxidation. These results suggest that high dietary intake of tomatoes may have protective effects against acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Antioxidants; Carotenoids; Ceruletide; Cytokines; Disease Models, Animal; Female; Glutathione; Lipase; Lipid Peroxidation; Lycopene; Male; Malondialdehyde; Oxidative Stress; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley

Acute pancreatitis is a common inflammatory disease mediated by damage to acinar cells and subsequent pancreatic inflammation with recruitment of leukocytes. We investigated the pathologic roles of innate immune cells, especially macrophages, in cerulein- and L-arginine-induced acute pancreatitis in mice.. Acute pancreatitis was induced by sequential peritoneal administration of cerulein to mice. We determined serum concentrations of amylase and lipase, pancreatic pathology, and features of infiltrating mononuclear cells. We performed parabiosis surgery to assess the hemodynamics of pancreatic macrophages.. Almost all types of immune cells, except for CD11b(high)CD11c(-) cells, were detected in the pancreas of healthy mice. However, activated CD11b(high)CD11c(-) cells, including Gr-1(low) macrophages and Gr-1(high) cells (granulocytes and myeloid-derived suppressor cells), were detected in damaged pancreas after cerulein administration. CCL2(-/-) mice given cerulein injections developed significantly less severe pancreatitis, with less infiltration of CD11b(high)CD11c(-)Gr-1(low) macrophages, but comparable infiltration of myeloid-derived suppressor cells, compared with cerulein-injected wild-type mice. Parabiosis and bone marrow analyses of these mice revealed that the CD11b(high)CD11c(-)Gr-1(low) macrophages had moved out of the bone marrow. Furthermore, mice with macrophage-specific deletion of suppressor of cytokine signaling 3 given injections of cerulein developed less severe pancreatitis and Gr-1(low) macrophage produced less tumor necrosis factor-α than wild-type mice given cerulein, although the absolute number of CD11b(high)CD11c(-)Gr-1(low) macrophages was comparable between strains. Induction of acute pancreatitis by L-arginine required induction of macrophage migration by CCL2, via the receptor CCR2.. Cerulein induction of pancreatitis in mice involves migration of CD11b(high)CD11c(-)Gr-1(low) macrophage from the bone marrow (mediated by CCL2 via CCR2) and suppressor of cytokine signaling 3-dependent activation of macrophage. These findings might lead to new therapeutic strategies for acute pancreatitis.

Topics: Acute Disease; Animals; Arginine; Biomarkers; CD11b Antigen; CD11c Antigen; Ceruletide; Chemokine CCL2; Chemotaxis; Disease Models, Animal; DNA-Binding Proteins; Enzymes; Immunity, Innate; Lymphocyte Depletion; Macrophage Activation; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis; Parabiosis; Receptors, CCR2; Receptors, Chemokine; Signal Transduction; Suppressor of Cytokine Signaling 3 Protein; Suppressor of Cytokine Signaling Proteins

Although microcirculatory disturbances play pivotal role in the pathomechanism of acute pancreatitis (AP), very few papers can be found which had been tested any of hemorheological parameters. The aim of our study was to analyze the hemorheological changes in cerulein-induced experimental acute pancreatitis in rat in two doses (5 and 10 μg/kg, s.c.). Male and female rats were subjected to Control group, or AP with 5 or 10 μg/kg cerulein groups. Blood samplings (lateral caudal vein) were completed before cerulein administration, and 1, 2 and 24 hours later. Hematological parameters, amylase activity, erythrocyte deformability (ektacytometry) and aggregation (light-transmission method) were tested. The presence of AP could be confirmed by amylase testing and histological examination. The earliest impairment of the red blood cell deformability could be observed 1 hour after cerulein administration in 10 μg/kg dosage. Female animals had the worst rheological results with high mortality. In conclusion, subcutaneously administrated cerulein in dosage of 5 and 10 μg/kg resulted in AP in rats, with significant changes in red blood cell deformability and alterations in erythrocyte aggregation. This model seems to be suitable for further comparative studies.

Topics: Acute Disease; Animals; Blood Cell Count; Ceruletide; Erythrocyte Aggregation; Erythrocyte Deformability; Female; Hemorheology; Male; Microcirculation; Pancreatitis; Rats; Rats, Sprague-Dawley; Sex Factors

Endoplasmic reticulum (ER) stress and hypertriglyceridemia (HTG) have been implicated in acute pancreatitis (AP).. For cellular model, rat exocrine acinar cells were preincubated with palmitic acid (0.05 or 0.1 mmol/L, 3 h) and stimulated with a cholecystokinin analog, CCK-8 (100 pmol/L, 30 min). For animal model, rats fed a high-fat diet to cause HTG and AP was induced by injection of caerulein (20 μg/kg). Injury to pancreatic cells was estimated by measuring amylase secretion, intracellular calcium concentration, apoptosis and histological changes. Expression of genes involved in ER stress-induced unfolded protein response (UPR) was monitored by RT-PCR and immunohistology.. In CCK-8 stimulated rat acinar cells, preincubation with PA caused an increased secretion of amylase, a higher and prolonged accumulation of intracellular calcium and increased apoptosis. Rats on high-fat diet had significantly elevated serum triglyceride levels. Induction of AP led to increased apoptosis in pancreatic tissue on high-fat diet than controls. For favoring HTG, expression of UPR components, GRP78/Bip, XBP-1, GADD153/CHOP and caspase-12 was upregulated.. Levels of markers of AP pathogenesis and components of UPR were elevated in the presence of excess fatty acids in pancreatic acinar cells. HTG appears to aggravate ER-stress and pathogenesis of AP.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Calcium; Ceruletide; Endoplasmic Reticulum; Endoplasmic Reticulum Stress; Gene Expression Regulation; Hypertriglyceridemia; Immunohistochemistry; Male; Palmitic Acid; Pancreas, Exocrine; Pancreatitis; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; Sincalide; Time Factors; Tissue Culture Techniques; Unfolded Protein Response

Hydrogen sulfide (H(2)S), a novel gaseous messenger, is synthesized endogenously from L-cysteine by two pyridoxal-5'-phosphate-dependent enzymes, cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE). S-propargyl-cysteine (SPRC) is a slow H(2)S releasing drug that provides cysteine, a substrate of CSE. The present study was aimed to investigate the effects of SPRC in an in vivo model of acute pancreatitis (AP) in mice. AP was induced in mice by hourly caerulein injections (50 µg/kg) for 10 hours. Mice were treated with SPRC (10 mg/kg) or vehicle (distilled water). SPRC was administered either 12 h before or 3 h before the induction of pancreatitis. Mice were sacrificed 1 h after the last caerulein injection. Blood, pancreas and lung tissues were collected and processed to measure the plasma amylase, plasma H(2)S, myeloperoxidase (MPO) activities and cytokine levels in pancreas and lung. The results revealed that significant reduction of inflammation, both in pancreas and lung was associated with SPRC given 3 h prior to the induction of AP. Furthermore, the beneficial effects of SPRC were associated with reduction of pancreatic and pulmonary pro-inflammatory cytokines and increase of anti-inflammatory cytokine. SPRC administered 12 h before AP induction did not cause significant improvement in pancreatic and lung inflammation. Plasma H(2)S concentration showed significant difference in H(2)S levels between control, vehicle and SPRC (administered 3 h before AP) treatment groups. In conclusion, these data provide evidence for protective effects of SPRC in AP possibly by virtue of its slow release of endogenous H(2)S.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents, Non-Steroidal; Ceruletide; Cysteine; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Gases; Hydrogen Sulfide; Inflammation; Lung; Male; Mice; Pancreas; Pancreatitis; Peroxidase; Time Factors

Novel and effective drugs against acute pancreatitis are required. Therefore, we examined the changes in the metabolite levels in the serum and pancreatic tissue of mice with cerulein- and arginine-induced pancreatitis using gas-chromatography/mass-spectrometry (GC/MS) and investigated whether these alterations affected the severity of acute pancreatitis. In the cerulein-induced pancreatitis model, 93 and 129 metabolites were detected in the serum and pancreatic tissue, respectively. In the L-arginine-induced acute pancreatitis model, 120 and 133 metabolites were detected in the serum and pancreatic tissue, respectively. Among the metabolites, the concentrations of tricarboxylic acid (TCA) cycle intermediates and amino acids were altered in pancreatitis, and in pancreatic tissue, the levels of the intermediates involved in the initial part of the TCA cycle were increased and those of the intermediates involved in the latter part of the TCA cycle were decreased. Some metabolites exhibited similar changes in both pancreatitis mouse models, e.g., the levels of glutamic acid and O-phosphoethanolamine were significantly decreased in the pancreatic tissue. Supplementation with glutamic acid and O-phosphoethanolamine attenuated the severity of cerulein-induced acute pancreatitis. Our results suggest that GC/MS-based metabolomics is capable of accurately representing the status of acute pancreatitis, leading to the discovery of therapeutic agents for pancreatitis.

Topics: Acute Disease; Amino Acids; Animals; Arginine; Ceruletide; Citric Acid Cycle; Disease Models, Animal; Ethanolamines; Gas Chromatography-Mass Spectrometry; Glutamic Acid; Male; Metabolomics; Mice; Mice, Inbred C57BL; Pancreatitis

Acute pancreatitis is a common disease, which is divided into mild pancreatitis and severe pancreatitis. For the latter, a systemic inflammatory response may occur and lead to distant organ damage and the development of multiple organ dysfunction syndrome (MODS), which accounts for significant morbidity and mortality in humans. Chemokines and their receptors are being believed to play a pivotal role in the pathogenesis of acute pancreatitis. Chemokine receptor CXCR3 is reported to be involved in acute tissue injury, for example acute lung injury induced by cigarette smoking, but its role in acute pancreatitis is not yet known. In this study, two animal models of acute pancreatitis (cerulein- and arginine-induced pancreatitis) were applied in CXCR3⁻/⁻ mice and wild-type mice, in order to explore the role of CXCR3 in acute pancreatitis. Serum amylase, lipase and histological observations revealed that CXCR3 knockout did not affect the severity of acute pancreatitis. However, edema and inflammatory cell infiltrate in the lung tissue were attenuated in CXCR3⁻/⁻ mice when acute pancreatitis was induced. In conclusion, chemokine receptor CXCR3 is not involved in acute pancreatic injury, but has a connection with acute pancreatitis-associated lung injury. Acute pulmonary injury is attenuated in CXCR3 knockout mice in experimental acute pancreatitis.

Topics: Acute Disease; Acute Lung Injury; Amylases; Animals; Arginine; Ceruletide; Disease Models, Animal; Lipase; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreatitis; Receptors, CXCR3; Severity of Illness Index

Peroxisome proliferator-activated receptors (PPARs) are ligand activated transcription factors belonging to the nuclear receptor superfamily. PPARs activation has a profound impact on the local immune response with consequences affecting the progression of chronic inflammatory diseases. Relatively little is known on the role of PPAR-β/δ in the regulation of inflammatory responses. The aim of the present study was to evaluate the influence of PPAR-β/δ receptor in a model of edematous pancreatitis induced in mice by administration of cerulein at supramaximal doses, as well as in necrohemorrhagic model induced by intraductal administration of sodium taurocholate (STC).. Mice were treated with cerulein (50 μg/kg) or STC (5%). GW0742 (0.3 mg/kg) was intraperitoneally administered 1 and 6 hours after cerulein injection or was injected 2 hours before STC infusion. The pancreas and exopancreatic organs were carefully removed for microscopic examination. Pancreatic weight, serum amylase, lipase, tumor necrosis factor-α and interleukin-1β levels, as well as cytokines, adhesion molecules, nitrotyrosine, poly (ADP-ribose), inducible nitric oxide, FAS ligand, Bax, Bcl-2 expression by immunohistochemistry, and myeloperoxidase activity of the pancreas were assayed. Moreover, the involvement of nuclear factor-κB pathway was investigated by Western blot analysis.. Intraperitoneal injection of cerulein in mice resulted in severe, acute pancreatitis characterized by edema, neutrophil infiltration and apoptosis, and elevated serum levels of amylase and lipase. Taurocholate challenge caused a clear increase in serum amylase, neutrophil infiltration, and tissue damage in the pancreas. Tissue and inflammatory changes in the pancreata were significantly less in GW0742 group than in cerulein or STC groups. In addition, the pancreatic water content was reduced in mice treated with PPAR-β/δ agonist. In the mild pancreatitis, GW0742 was also able to decrease the expression of proinflammatory cytokines and enzymes, as well as of proteins involved in apoptosis and nuclear factor-Kappa B pathway.. GW0742 attenuated pancreatic damage in 2 different experimental models of pancreatitis in mice.

Topics: Acute Disease; Amylases; Animals; Cell Adhesion Molecules; Cell Movement; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Lipase; Male; Mice; Mice, Inbred Strains; Neutrophils; Pancreatitis; PPAR delta; PPAR-beta; Taurocholic Acid; Thiazoles; Time Factors

Bmi1 is a member of the Polycomb protein family and represses transcription by modifying chromatin organization at specific promoters. Bmi1 is implicated in the control of stem cell self-renewal and has been shown to regulate cell proliferation, tissue homeostasis, and differentiation. Bmi1 is present in a subpopulation of self-renewing pancreatic acinar cells and is expressed in response to pancreatic damage. We investigated the role of Bmi1 in regeneration of exocrine pancreas.. Acute pancreatitis was induced in Bmi1(-/-) mice with cerulein; pancreatic cell regeneration, differentiation, and apoptosis were assessed. Cultured Bmi1(-/-) and wild-type primary acini were analyzed in vitro to determine acinar-specific consequences of Bmi1 deletion. To investigate cell autonomous versus non-cell autonomous roles for Bmi1 in vivo, pancreatitis was induced in Bmi1(-/-) mice reconstituted with a wild-type hematopoietic system.. Bmi1 expression was up-regulated in the exocrine pancreas during regeneration after cerulein-induced pancreatitis. Exocrine regeneration was impaired following administration of cerulein to Bmi1(-/-) mice. Pancreata of Bmi1(-/-) mice were hypoplastic, and the exocrine pancreas was replaced with ductal metaplasia that had increased apoptosis and decreased cell proliferation compared with that of wild-type mice. Expression of Cdkn2a and p53-dependent apoptotic genes was markedly up-regulated in Bmi1(-/-) pancreas compared with wild-type mice after injury. Furthermore, after transplantation of bone marrow from wild-type to Bmi1(-/-) mice, the chimeric mice had intermediate levels of pancreatic hypoplasia and significant but incomplete rescue of impaired exocrine regeneration after cerulein injury.. Bmi1 contributes to regeneration of the exocrine pancreas after cerulein-induced injury through cell autonomous mechanisms, in part by regulating Cdkn2a expression, and non-cell autonomous mechanisms.

Topics: Acute Disease; Animals; Apoptosis; Bone Marrow Transplantation; Cell Differentiation; Cell Proliferation; Ceruletide; Choline Deficiency; Cyclin-Dependent Kinase Inhibitor p16; Disease Models, Animal; Ethionine; Female; Gene Expression Regulation; Green Fluorescent Proteins; Mice; Mice, Knockout; Mice, Transgenic; Nuclear Proteins; Pancreas, Exocrine; Pancreatitis; Polycomb Repressive Complex 1; Proto-Oncogene Proteins; Regeneration; Repressor Proteins; Time Factors; Tissue Culture Techniques; Transplantation Chimera; Tumor Suppressor Protein p53

To study the potential role of the 78kDa glucose regulated protein (GRP78) in the pathogenesis of acute pancreatitis (AP) in vitro.. AR42J cells were stimulated by cerulein or cerulein plus lipoplysaccharide (LPS). The severity of pancreatic inflammation was evaluated by amylase, lipase, TNF-a, and IL-6. Apoptosis was determined by flow cytometry; the expressions of apoptotic genes, GRP78 and the downstream molecules were determined by real-time quantitative PCR and Western blot.. After cerulein stimulation, the levels of amylase, lipase, TNF-a and IL-6 were all increased, with a more pronounced increase after cerulein plus LPS stimulation. Apoptosis was different in two cell models, high apoptosis in cerulein group; whereas cerulein plus LPS induced relatively less apoptosis. Apoptotic gene expressions revealed more pronounced increase in the cerulein group than those in cerulein plus LPS group. The expressions of GRP78 and downstream molecules were different in two cell models. GRP78 expression was down-regulated in cerulein group and upregulated in cerulein plus LPS group.. GRP78 expression was associated with apoptosis and the severity of cerulein-induced pancreatic inflammation, indicating that GRP78 might prevent apoptosis in pancreatic acinar cells thereby deteriorating the severity of AP.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Blotting, Western; Cell Line; Ceruletide; DNA-Binding Proteins; Flow Cytometry; Gene Expression Regulation; Heat-Shock Proteins; Interleukin-6; Lipase; Lipopolysaccharides; Necrosis; Pancreas, Exocrine; Pancreatitis; Rats; Real-Time Polymerase Chain Reaction; Regulatory Factor X Transcription Factors; RNA, Messenger; Severity of Illness Index; Signal Transduction; Time Factors; Transcription Factor CHOP; Transcription Factors; Tumor Necrosis Factor-alpha

The purpose of this study was to test the hypothesis that activation of endogenous peroxisome proliferator-activated receptor (PPARγ) inhibits induction of early growth response factor-1 (Egr-1), which is rapidly induced in the pancreas following cerulein intraperitoneal injection. Acute pancreatitis was induced in mice by hourly intraperitoneal injection of cerulein. Pioglitazone was administered prophylactically and pancreatic inflammation was assessed. AR42J cells were stimulated with caerulein 10⁻⁸ M co-incubated in presence of different concentration of pioglitazone. The expression of PPARγ, Egr-1, and the target genes of Egr-1 were studied by real-time reverse transcriptase polymerase chain reaction (PCR), Western blot, and immunohistochemistry. In vitro, a PPAR-γ activator (pioglitazone) strikingly diminished Egr-1 mRNA and protein expression corresponding to Egr-1. In vivo, treatment with pioglitazone prior to the intraperitoneal injection of cerulein induction of Egr-1 and its target genes such as, monocyte chemotactic protein-1 (MCP-1) and macrophage inflammatory protein-1 (MIP-1). The inhibitory effect of pioglitazone on Egr-1 expression induced by cerulein was almost fully restored by GW9662. Activation of PPAR-γ suppressed the activation of Egr-1 and its inflammatory gene targets and provided potent protection against pancreas injury. These data suggest a new mechanism in which PPAR-γ activation may decrease tissue inflammation in response to a cerulein insult.

Topics: Acinar Cells; Acute Disease; Anilides; Animals; Cell Line, Tumor; Ceruletide; Chemokine CCL2; Chemokines; Disease Models, Animal; Early Growth Response Protein 1; Hypoglycemic Agents; Male; Pancreatitis; Pioglitazone; PPAR gamma; Rats; Rats, Sprague-Dawley; RNA, Messenger; Severity of Illness Index; Thiazolidinediones; Translational Research, Biomedical

Alterations in protein expression within the initiation phase of acute pancreatitis (AP) might play an important role in the development of this disease, lysosomes being involved in its pathophysiology. The use of pancreatic subcellular fractions in proteomic analysis, simplifies protein maps and helps in the identification of new protein changes and biomarkers characterizing tissue damage. The present study aims to determine the differentially expressed acidic proteins in the pancreatic soluble and lysosomal+mitochondrial (L+M) fractions from rats during the early phase of the experimental model of cerulein (Cer)-induced AP. Subcellular pancreatic extracts from diseased and control rats were analyzed by 2-DE (3-5.6 pH range) and MALDI-TOF/TOF MS. Comparative analysis afforded the conclusive identification of 13 (soluble fraction) and 7 (L+M fraction) proteins or protein fragments occuring in different amounts between diseased and control pancreas, some of them being newly described in AP. In the soluble fraction, we detected changes related to inflammation and apoptosis (α1-inhibitor-3, α-1 antitrypsin, α-1 macroglobulin, haptoglobin, STRAP), oxidative stress and stress response (peroxiredoxin-2, thioredoxin-like 1, GRP94/TRA1, heat shock cognate 71kDa protein), digestive proteases (elastase 3B), serine protease inhibition (serpins B6 and A3L) and translation processes (EF 1-δ). In the L+M fraction, we detected changes mainly related to energy generation or cellular metabolism (ATP synthase β subunit, chymotrypsinogen B, triacylglycerol lipase), cell redox homeostasis (iodothyronine 5´monodeiodinase) and digestive proteases (carboxypeptidase B1). The data should provide valuable information for unraveling the early pathophysiologic mechanisms of Cer-induced AP.

Topics: Acute Disease; Animals; Apoptosis; Biomarkers; Ceruletide; Electrophoresis, Gel, Two-Dimensional; Hydrogen-Ion Concentration; Lysosomes; Male; Mitochondria; Pancreas; Pancreatitis; Proteome; Proteomics; Rats; Rats, Wistar; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Subcellular Fractions

The aim of the present study was to develop a model of hypertriglyceridemic (HTG) acute pancreatitis and to investigate the effects of probucol in this model.. Hamsters were fed a high-fat diet (HFD) or a normal diet for 3 weeks. Probucol was added at 1% to the HFD in the treated group. Pancreatitis was induced by 7 peritoneal injections of cerulein to the normal and HFD hamster groups. The severity of the pancreatitis and whole body oxidative stress were assessed.. The HFD induced severe HTG (>1000 mg/dL) in the hamsters. A more severe pancreatitis was observed in the HFD group. The HFD did not influence plasma-reduced glutathione level, but there was a significant increase after 1% probucol was provided in the diet. Plasma malonaldehyde levels in the HFD group were significantly higher than the normal chow group, whereas probucol administration significantly decreased plasma hydrogen peroxide and malonaldehyde levels. We also found that probucol significantly reduced levels of amylase and lipase in the plasma and pathological scores in pancreatic tissue.. This study presents a novel model of severe HTG acute pancreatitis, and our results support the potential therapeutic application of probucol in HTG acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Antioxidants; Biomarkers; Ceruletide; Cricetinae; Diet, High-Fat; Disease Models, Animal; Glutathione; Hydrogen Peroxide; Hypertriglyceridemia; Lipase; Malondialdehyde; Oxidative Stress; Pancreas; Pancreatitis; Probucol; Severity of Illness Index

To determine if the fraction of Nardostachys jatamansi (NJ) has the potential to ameliorate the severity of acute pancreatitis (AP).. Mice were administered the biologically active fraction of NJ, i.e., the 4th fraction (NJ4), intraperitoneally, and then injected with the stable cholecystokinin analogue cerulein hourly for 6 h. Six hours after the last cerulein injection, the pancreas, lung, and blood were harvested for morphological examination, measurement of cytokine expression, and examination of neutrophil infiltration.. NJ4 administration attenuated the severity of AP and lung injury associated with AP. It also reduced cytokine production and neutrophil infiltration and resulted in the in vivo up-regulation of heme oxygenase-1 (HO-1). Furthermore, NJ4 and its biologically active fraction, NJ4-2 inhibited the cerulein-induced death of acinar cells by inducing HO-1 in isolated pancreatic acinar cells.. These results suggest that NJ4 may be a candidate fraction offering protection in AP and NJ4 might ameliorate the severity of pancreatitis by inducing HO-1 expression.

Topics: Acute Disease; Animals; Cell Death; Ceruletide; Cytokines; Cytoprotection; Disease Models, Animal; Dose-Response Relationship, Drug; Enzymes; Female; Heme Oxygenase-1; Inflammation Mediators; Lung; Membrane Proteins; Mice; Mice, Inbred C57BL; Nardostachys; Neutrophil Infiltration; Pancreas; Pancreatitis; Plant Extracts; Plant Roots; Severity of Illness Index; Time Factors; Up-Regulation

Heat shock protein (Hsp) 72 is a molecular chaperone which is upregulated in response to a variety of stress situations and has a general cytoprotective function. Increased Hsp72 levels were implicated in protection from acute pancreatitis; a hypothesis which was not tested in a transgenic mouse model yet.. To analyze the role of Hsp72 during acute pancreatitis, well-characterized transgenic animals overexpressing rat Hsp72 (Hsp72 mice) under the control of the ß-actin promoter were subjected to caerulein- and L-arginine-induced acute pancreatitis. The severity of experimental pancreatitis was determined via serum lipase levels, morphometric evaluation and quantification of pancreatic edema/inflammation.. Hsp72 mice displayed ∼100-times Hsp72 overexpression, but no changes in the remaining chaperones. Robust Hsp72 signal was observed in pancreatic acini, but not in islets or ductal cells. In both models, elevated Hsp72 did not protect from development of acute pancreatitis and the pancreatitis-associated lung injury, but accelerated recovery from caerulein-induced tissue injury (lower lipase levels, edema, inflammation and necrosis 36 h after caerulein administration). The observed protective function of Hsp72 in caerulein-induced pancreatitis is likely due to an attenuated NF-κB signalling.. Hsp72 overexpression accelerates the recovery from acute pancreatitis and may represent a potential treatment strategy.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Disease Models, Animal; Gene Expression; Gene Expression Regulation; HSP72 Heat-Shock Proteins; Mice; Mice, Transgenic; NF-kappa B; Pancreas, Exocrine; Pancreatitis; Recovery of Function; Signal Transduction

Hedgehog signaling plays critical roles in pancreatic oncogenesis and chronic pancreatitis, but its roles in acute pancreatitis (AP) are largely ambiguous. In this study, we provide evidence that Sonic hedgehog (Shh), but neither Desert hedgehog (Dhh) nor Indian hedgehog (Ihh), is the main protein whose expression is activated during the development of cerulein-induced acute pancreatitis in mice, and the Shh serves as an anti-inflammation factor in an autocrine manner. Blocking autocrine Shh signaling with anti-Shh neutralizing antibody aggravates the progression of acute pancreatitis. Mechanistic insight into Shh signaling activation in acute pancreatitis indicates that inflammatory stimulation activates Shh expression and secretion, and subsequently upregulates the expression and secretion of interleukin-10 (IL-10). Moreover, inhibition of Shh signaling with neutralizing antibody abolishes IL-10 production in vivo and in vitro. Molecular biological studies show that autocrine Shh signaling activates the key transcriptional factor Gli1 so that the target gene IL-10 is upregulated, leading to the protective and anti-inflammatory functions in the mouse model of acute pancreatitis. Thus, this study suggests autocrine Shh signaling functions as a protective signaling in the progression of acute pancreatitis.

Topics: Acute Disease; Animals; Autocrine Communication; Cell Line; Ceruletide; Gene Expression; Gene Expression Regulation; Hedgehog Proteins; Interleukin-10; Kruppel-Like Transcription Factors; Male; Mice; Mice, Inbred C57BL; Neutrophil Infiltration; Pancreas; Pancreatitis; Signal Transduction; Up-Regulation; Zinc Finger Protein GLI1

Acute pancreatitis (AP) is a complicated inflammatory disease that has an unknown underlying pathogenesis. Because alpha-pinene can modulate inflammation, we examined whether alpha-pinene plays a role in AP.. Alpha-pinene was administered intraperitoneally 1h prior to the first injection of cerulein. Once AP developed, cerulein, a stable cholecystokinin analog, was injected hourly over a 6-h period. Blood samples were taken 6h later to determine serum amylase and lipase levels. The pancreas and lungs were rapidly removed for morphological examination, myeloperoxidase assay, and real-time reverse transcription polymerase chain reaction. We also isolated the pancreatic acinar cells using a collagenase solution. Cell viability, and cytokine productions were measured in pancreatic acini.. Intraperitoneal administration of alpha-pinene reduced the pancreatic weight (PW) to body weight (BW) ratio and the serum levels of amylase and lipase. Alpha-pinene treatment also reduced histological damage and myeloperoxidase activity in the pancreas and lungs. Furthermore, alpha-pinene pretreatment reduced the production of pancreatic tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 during cerulein-induced AP. In vitro, alpha-pinene inhibited cerulein-induced cell death and cytokine production in isolated cerulein-treated pancreatic acinar cells.. These findings suggest that alpha-pinene has an anti-inflammatory effect during cerulein-induced AP.

Topics: Acute Disease; Amylases; Animals; Bicyclic Monoterpenes; Body Weight; Cells, Cultured; Ceruletide; Female; Immunologic Factors; Injections, Intraperitoneal; Lipase; Lung; Mice; Mice, Inbred C57BL; Monoterpenes; Pancreas; Pancreatitis; Peroxidase

The type of immune response during development of acute pancreatitis (AP) determines disease severity. Pancreatic epithelial cells express the interleukin (IL)-22 receptor A1 (IL-22RA1). The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that regulates expression of IL-22. We investigated sources and role of IL-22 in the pancreas, along with the effects of AhR activation on IL-22 expression and AP progression in mice.. We analyzed the effects of recombinant IL-22, a monoclonal antibody against IL-22, and agonists and antagonists of AhR in mice with AP (induced with caerulein or a choline-deficient diet supplemented with DL-ethionine) and control mice. We also analyzed transgenic mice with AhR deficiency (AhR(d) and AhR(-/-) mice).. CD4(+) T cells were the main source of IL-22 in pancreatic tissues from healthy mice. During development of AP, numbers of IL-22(+) CD4(+) T cells were reduced, whereas IL-22RA1 was up-regulated. Consistent with high levels of IL-22RA1 expression, pancreatic acinar cells responded to IL-22 signaling via signal transducers and activators of transcription 3; administration of IL-22 reduced AP and associated lung injury in mice. AhR was required for production of IL-22 and protected mice from AP. Mice that did not respond to AhR activation developed AP, but administration of IL-22 reduced AP; blockade of IL-22 reversed the ability of activated AhR to protect against AP.. AhR activation protects mice from AP by inducing expression of IL-22. AhR therefore mediates interactions between pancreatic leukocytes and epithelial cells and might be developed as a therapeutic target.

Topics: Acute Disease; Animals; CD4-Positive T-Lymphocytes; Ceruletide; Choline Deficiency; Disease Models, Animal; Female; Interleukin-22; Interleukins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Pancreas; Pancreatitis; Phosphorylation; Receptors, Aryl Hydrocarbon; Signal Transduction; STAT3 Transcription Factor

The discovery of novel and effective treatment methods would be of great help to patients with acute pancreatitis. The aims of this study were to determine the inhibitory effects of vitamin K3 (VK3) against cerulein-induced acute pancreatitis in mice and to examine the mechanisms behind these effects.. Acute pancreatitis in mice was induced by intraperitoneal injection of cerulein 6 times at hourly intervals. Vitamin K3 was administered once before the first injection of cerulein or twice before and after the first injection of cerulein. The degrees of inflammation and autophagy in the pancreatic tissue were estimated by histological examination, measurement of enzyme activity, confocal microscopy, and Western blotting. The inhibitory effects of VK3 against rapamycin-induced autophagy were also examined using HeLa cells stably expressing green fluorescent protein LC3.. Cerulein-induced acute pancreatitis was markedly attenuated by the administration of VK3. In addition, VK3 led to the inhibition of cerulein-evoked autophagic changes and colocalization of autophagosomes and lysosomes in the pancreatic tissue. Vitamin K3 also reduced rapamycin-induced autophagy in HeLa/green fluorescent protein LC3 cells.. Our data suggest that the administration of VK3 reduces pancreatic inflammation in acute pancreatitis through inhibition of the autophagic pathway. Vitamin K3 may be an effective therapeutic strategy against acute pancreatitis.

Topics: Acute Disease; Animals; Autophagy; Ceruletide; Female; HeLa Cells; Humans; Lysosomes; Mice; Mice, Inbred C57BL; Pancreatitis; Phagosomes; Sirolimus; Vitamin K 2; Vitamin K 3

Lysosomes play an important role in acute pancreatitis (AP). Here we developed a method for the isolation of lysosome subpopulations from rat pancreas and assessed the stability of lysosomal membranes.. AP was induced by four subcutaneous injections of 20 μg caerulein/kg body weight at hourly intervals. The animals were killed 9h after the first injection. Marker enzymes [N-acetyl-β-D-glucosaminidase (NAG), cathepsin B and succinate dehydrogenase (SDH)] were assayed in subcellular fractions from control pancreas and in pancreatitis. Lysosomal subpopulations were separated by Percoll density gradient centrifugation and observed by electron microscopy. NAG molecular forms were determined by DEAE-cellulose chromatography.. AP was associated with: (i) increases in the specific activity of lysosomal enzymes in the soluble fraction, (ii) changes in the size and alterations in the morphology of the organelles from the lysosomal subpopulations, (iii) the appearance of large vacuoles in the primary and secondary lysosome subpopulations, (iv) the increase in the amount of the NAG form associated with the pancreatic lysosomal membrane as well as its release towards the soluble fraction.. Lysosome subpopulations are separated by a combination of differential and Percoll density gradient centrifugations. Primary lysosome membrane stability decreases in AP.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Lysosomes; Male; Pancreatitis; Rats; Rats, Wistar

Curcuma longa (CL) has been reported to possess a variety of pharmacological activities. However, the effects of CL on acute pancreatitis (AP) have not yet been determined. To this end, we examined the effects of CL on cerulein-induced AP. Cell viability and cytokine productions were measured in pancreatic acini. Mice were divided into 3 groups: i) Normal group, ii) normal saline-treated group, iii) group treated with CL at a dose of 0.05, 0.1, 0.5 and 1 g/kg. CL was administered orally to mice for 7 days. The mice were intraperitoneally injected with the stable cholecystokinin analogue, cerulein (50 μg/kg), every hour for a total of 6 h. The mice were sacrificed 6 h after the completion of the cerulein injections. Blood samples were obtained to determine serum amylase, lipase and cytokine levels. The pancreas was rapidly removed for morphological examination, measurement of tissue myeloperoxidase activity, as well as the level of cytokines and heme oxygenase-1 (HO-1). The CL treatment reduced cerulein-induced cell death and cytokine production in pancreatic acini. The administration of CL significantly ameliorated the severity of pancreatitis and pancreatitis-associated lung injury, as was shown by the reduction in pancreatic edema, neutrophil infiltration, vacuolization, necrosis, serum amylase, lipase and cytokine levels, and mRNA expression of multiple inflammatory mediators such as interleukin (IL)-1ß and -6 and tumor necrosis factor (TNF)-α. In order to identify the regulatory mechanism of CL on cerulein-induced pancreatitis, we examined the level of HO-1 in the pancreas. We found that the administration of CL induced HO-1. Our results suggest that CL plays a protective role in the development of AP and pancreatitis-associated lung injury.

Topics: Acute Disease; Animals; Ceruletide; Curcuma; Cytokines; Female; Heme Oxygenase-1; Lung Injury; Mice; Mice, Inbred C57BL; Pancreas; Pancreatitis; Peroxidase; Plant Extracts

Acute pancreatitis (AP) has a high mortality rate; repetitive AP induces chronic AP and pancreatic adenocarcinoma. Mesenchymal stem cells (MSCs) have immunoregulatory effects and reduce inflammation. We developed a protocol to isolate human bone marrow-derived clonal MSCs (hcMSCs) from bone marrow aspirate and investigated the effects of these cells in rat models of mild and severe AP.. Mild AP was induced in Sprague-Dawley rats by 3 intraperitoneal injections of cerulein (100 μg/kg), given at 2-hour intervals; severe AP was induced by intraparenchymal injection of 3% sodium taurocholate solution. hcMSCs were labeled with CM-1,1'-dioctadecyl-3,3,3'-tetramethylindo-carbocyanine perchloride and administered to rats through the tail vein.. hcMSCs underwent self-renewal and had multipotent differentiation capacities and immunoregulatory functions. Greater numbers of infused hcMSCs were detected in pancreas of rats with mild and severe AP than of control rats. Infused hcMSCs reduced acinar-cell degeneration, pancreatic edema, and inflammatory cell infiltration in each model of pancreatitis. The hcMSCs reduced expression of inflammation mediators and cytokines in rats with mild and severe AP. hcMSCs suppressed the mixed lymphocyte reaction and increased expression of Foxp3(+) (a marker of regulatory T cells) in cultured rat lymph node cells. Rats with mild or severe AP that were given infusions of hcMSCs had reduced numbers of CD3(+) T cells and increased expression of Foxp3(+) in pancreas tissues.. hcMSCs reduced inflammation and damage to pancreatic tissue in a rat model of AP; they reduced levels of cytokines and induced numbers of Foxp3(+) regulatory T cells. hcMSCs might be developed as a cell therapy for pancreatitis.

Topics: Acute Disease; Animals; Biomarkers; Bone Marrow Transplantation; CD3 Complex; Cell Differentiation; Cell Proliferation; Cells, Cultured; Ceruletide; Coculture Techniques; Cytokines; Disease Models, Animal; Forkhead Transcription Factors; Humans; In Situ Hybridization, Fluorescence; Inflammation Mediators; Mesenchymal Stem Cell Transplantation; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Rats, Wistar; Regeneration; Severity of Illness Index; T-Lymphocytes; Taurocholic Acid; Time Factors

Topics: Acute Disease; Animals; Bicarbonates; Ceruletide; Disease Progression; Galanin; Mice; Pancreatitis

Acute pancreatitis is an inflammatory condition that may lead to multisystemic organ failure. Melanocortin peptides have been successfully used in experimental models of organ failure and shock, and their protective effect occurs through the activation of a vagus nerve-mediated cholinergic anti-inflammatory pathway by acting at brain melanocortin 4 receptors. In the light of these observations, we studied the effects of the selective melanocortin 4 receptor agonist RO27-3225 in an experimental model of cerulein-induced pancreatitis.. Randomized experiment.. Research laboratory at a university hospital.. Experimental pancreatitis in rats.. Acute pancreatitis was induced in male Sprague-Dawley rats by intraperitoneal injections of cerulein (80 μg/kg, four injections at hourly intervals). Before pancreatitis induction, groups of animals were subjected to bilateral cervical vagotomy, pretreated with the nicotinic acetylcholine receptor antagonist chlorisondamine or the selective melanocortin 4 receptor antagonist HS024, or not pretreated. Thirty minutes after the first cerulein injection, rats were intraperitoneally treated with a nanomolar dose of RO27-3225 or vehicle. Some experimental groups were prepared for neural efferent activity recording along the vagus nerve starting 30 mins after treatment with RO27-3225 or vehicle, and for a 30-min period.. Serum lipase and amylase activity, tumor necrosis factor-α and interleukin-6 expression, pancreatic myeloperoxidase activity, and histologic damage were evaluated; neural efferent activity of vagal fibers was also assessed. RO27-3225 reduced cerulein-induced serum lipase and amylase activity, blunted the expression of tumor necrosis factor-α and interleukin-6, abated the increase in pancreatic myeloperoxidase activity, and protected against histologic damage. Furthermore, RO27-3225 markedly increased neural efferent activity along the vagus nerve. Vagotomy, chlorisondamine, and HS024 abated these protective effects of RO27-3225.. Our data show that melanocortin 4 receptor agonists reduce pancreatitis severity through the activation of the cholinergic anti-inflammatory pathway. These findings could be of particular interest in the clinical setting.

Topics: Acute Disease; Analysis of Variance; Animals; Blotting, Western; Ceruletide; Cholinergic Agents; Disease Models, Animal; Immunohistochemistry; Inflammation Mediators; Interleukin-6; Male; Pancreatitis; Peptides; Peroxidase; Random Allocation; Rats; Rats, Sprague-Dawley; Receptor, Melanocortin, Type 4; Receptors, Nicotinic; Sensitivity and Specificity; Severity of Illness Index; Signal Transduction; Tumor Necrosis Factor-alpha; Vagus Nerve

This study aimed to determine the effect of hydrogen sulfide (H2S) on Toll-like receptor 4 (TLR4)-mediated innate immune signaling in acute pancreatitis (AP) via substance P.. Male Swiss mice were treated with hourly intraperitoneal injections of cerulein (50 μg/kg) for 10 hours. dl-propargylglycine ([PAG] 100 mg/kg, intraperitoneally), an inhibitor of H2S formation, was administered 1 hour after the induction of AP. Pancreatic acinar cells from male preprotachykinin-A gene-knockout mice (PPTA) and their wild-type counterparts were incubated with or without cerulein (10 M for 60 minutes). To better understand the effect of H2S in inflammation, acinar cells were stimulated with cerulein after addition of H2S donor, sodium hydrosulfide. In addition, cerulein-treated pancreatic acinar cells were pretreated with PAG (30 μM) for 1 hour.. The H2S inhibitor PAG eliminated TLR4, interleukin 1 receptor-associated kinase 4, tumor necrosis factor receptor-associated factor 6, and nuclear factor-κB (NF-κB) levels in in vitro and in vivo models of cerulein-induced AP. PPTA gene deletion reduced TLR4, myeloid differentiation factor 88, interleukin 1 receptor-associated kinase 4, tumor necrosis factor receptor-associated factor 6, and NF-κB in cerulein-treated pancreatic acinar cells, whereas administration of sodium hydrosulfide resulted in a further rise in TLR4 and NF-κB levels in cerulein-treated pancreatic acinar cells.. The present findings show for the first time that in AP, H2S may up-regulate the TLR4 pathway and NF-κB via substance P.

Topics: Acute Disease; Animals; Base Sequence; Ceruletide; DNA Primers; Gene Deletion; Hydrogen Sulfide; Immunity, Innate; Interleukin-1 Receptor-Associated Kinases; Male; Mice; Myeloid Differentiation Factor 88; NF-kappa B; Pancreas; Pancreatitis; Protein Precursors; Signal Transduction; Substance P; Tachykinins; TNF Receptor-Associated Factor 6; Toll-Like Receptor 4; Up-Regulation

Acute pancreatitis is an inflammatory process of variable severity. Leucocytes are thought to play a key role in the development of pancreatitis and pancreatitis-associated lung injury. The interactions between inflammatory cells and their mediators are crucial for determining tissue damage. Monocyte chemoattractant protein-1 (or CCL-2), CCR-2 and CCR-4 are chemokines and chemokine receptors involved in leucocyte trafficking. The aim of the study was to evaluate the role of the CCL-2, CCR-2 and CCR-4 chemokine receptors in the pathogenesis of cerulein-induced pancreatitis and pancreatitis-associated lung injury. To address the role of CCL-2, CCR-2 and CCR-4 that attracts leucocytes cells in inflamed tissues, pancreatitis was induced by administering supramaximal doses of cerulein in mice that do not express CCL-2, CCR-2 or CCR-4.. The severity of pancreatitis was measured by serum amylase, pancreatic oedema and acinar cell necrosis. Lung injury was quantitated by evaluating lung microvascular permeability and lung myeloperoxidase activity. Chemokine and chemokine-receptor expression were quantitated by real-time PCR. The nature of inflammatory cells invading the pancreas and lungs was studied by immunostaining.. The authors have found that pancreas CCL-2 and CCR-2 levels rise during pancreatitis. Both pancreatitis and the associated lung injury are blunted, but not completely prevented, in mice deficient in CCL-2, whereas the deficiency in either CCR-2 or CCR-4 does not reduce the severity of both the pancreatitis and the lung injury. The amounts of neutrophils and monocyte/macrophages (MOMA)-2 cells were significantly lower in mice deficient in CCL-2 compared with their sufficient littermates.. These results suggest that CCL-2 plays a key role in pancreatitis by modulating the infiltration by neutrophils and MOMA-2 cells, and that its deficiency may improve the outcome of the disease.

Topics: Acute Disease; Animals; Ceruletide; Chemokine CCL2; Disease Models, Animal; Immunohistochemistry; Lung Injury; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Mice, Knockout; Neutrophil Infiltration; Organ Size; Pancreas; Pancreatitis; Receptors, CCR2; Receptors, CCR4

Excessive Ca2+ influx mediates many cytotoxic processes, including those associated with autoimmune inflammatory diseases such as acute pancreatitis and Sjögren syndrome. Transient receptor potential (canonical) channel (TRPC) 3 is a major Ca2+ influx channel in pancreatic and salivary gland cells. We investigated whether genetic or pharmacologic inhibition of TRPC3 protects pancreas and salivary glands from Ca2+-dependent damage.. We developed a Ca2+-dependent model of cell damage for salivary gland acini. Acute pancreatitis was induced by injection of cerulein into wild-type and Trpc3-/- mice. Mice were also given the Trpc3-selective inhibitor pyrazole 3 (Pyr3).. Salivary glands and pancreas of Trpc3-/- mice were protected from Ca2+-mediated cell toxicity. Analysis of Ca2+ signaling in wild-type and Trpc3-/- acini showed that Pyr3 is a highly specific inhibitor of Tprc3; it protected salivary glands and pancreas cells from Ca2+-mediated toxicity by inhibiting the Trpc3-mediated component of Ca2+ influx.. TRPC3-mediated Ca2+ influx mediates damage to pancreas and salivary glands. Pharmacologic inhibition of TRPC3 with the highly selective TRPC3 inhibitor Pyr3 might be developed for treatment of patients with acute pancreatitis and Sjögren syndrome.

Topics: Acute Disease; Animals; Calcium Channel Blockers; Calcium Signaling; Ceruletide; Disease Models, Animal; Epithelial Cells; Mice; Mice, Knockout; Pancreas; Pancreatitis; Pyrazoles; Salivary Gland Diseases; Salivary Glands; Severity of Illness Index; Time Factors; TRPC Cation Channels

Recent data suggest that platelets not only control thrombosis and hemostasis but may also regulate inflammatory processes such as acute pancreatitis. However, the specific role of platelet-derived mediators in the pathophysiology of acute pancreatitis is not known. Herein, we examined the role of CD40 ligand (CD40L, CD154) in different models of acute pancreatitis. Acute pancreatitis was induced by repetitive caerulein administration (50μg/kg, i.p.) or infusion of sodium taurocholate (5%-10μl) into the pancreatic duct in wild-type C57BL/6 and CD40L-deficient mice. Neutrophil infiltration, myeloperoxidase (MPO), macrophage inflammatory protein-2 (MIP-2) levels, acinar cell necrosis, edema and hemorrhage in the pancreas as well as serum amylase activity and lung levels of MPO were quantified 24h after induction of acute pancreatitis. Caerulein and taurocholate challenge caused a clear-cut pancreatic damage characterized by increased acinar cell necrosis, neutrophil infiltration, focal hemorrhage, edema formation as well as increased levels of serum amylase and MIP-2 in the pancreas and lung MPO and histological damage. Notably, CD40L gene-deficient animals exhibited a similar phenotype as wild-type mice after challenge with caerulein and taurocholate. Similarly, administration of an antibody directed against CD40L had no effect against acute pancreatitis. Our data suggest that CD40L does not play a functional role in experimental acute pancreatitis. Thus, other candidates than CD40L needs to be explored in order to identify platelet-derived mediators in the pathophysiology of acute pancreatitis.

Topics: Acute Disease; Animals; CD40 Ligand; Ceruletide; Disease Models, Animal; Male; Mice; Mice, Inbred C57BL; Pancreatitis; Taurocholic Acid

Acute pancreatitis is a life-threatening inflammatory disease characterized by abdominal pain of unknown etiology. Trypsin, a key mediator of pancreatitis, causes inflammation and pain by activating protease-activated receptor 2 (PAR(2)), but the isoforms of trypsin that cause pancreatitis and pancreatic pain are unknown. We hypothesized that human trypsin IV and rat P23, which activate PAR(2) and are resistant to pancreatic trypsin inhibitors, contribute to pancreatic inflammation and pain. Injections of a subinflammatory dose of exogenous trypsin increased c-Fos immunoreactivity, indicative of spinal nociceptive activation, but did not cause inflammation, as assessed by measuring serum amylase and myeloperoxidase activity and by histology. The same dose of trypsin IV and P23 increased some inflammatory end points and caused a more robust effect on nociception, which was blocked by melagatran, a trypsin inhibitor that also inhibits polypeptide-resistant trypsin isoforms. To determine the contribution of endogenous activation of trypsin and its minor isoforms, recombinant enterokinase (ENK), which activates trypsins in the duodenum, was administered into the pancreas. Intraductal ENK caused nociception and inflammation that were diminished by polypeptide inhibitors, including soybean trypsin inhibitor and a specific trypsin inhibitor (type I-P), and by melagatran. Finally, the secretagogue cerulein induced pancreatic nociceptive activation and nocifensive behavior that were reversed by melagatran. Thus trypsin and its minor isoforms mediate pancreatic pain and inflammation. In particular, the inhibitor-resistant isoforms trypsin IV and P23 may be important in mediating prolonged pancreatic inflammatory pain in pancreatitis. Our results suggest that inhibitors of these isoforms could be novel therapies for pancreatitis pain.

Topics: Abdominal Pain; Acute Disease; Amylases; Analgesics; Animals; Azetidines; Benzylamines; Ceruletide; Disease Models, Animal; Enteropeptidase; Enzyme Activation; Humans; Kinetics; Male; Pain Measurement; Pancreas; Pancreatitis; Peroxidase; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Receptor, PAR-2; Recombinant Proteins; Signal Transduction; Soybean Proteins; Spinal Cord; Trypsin; Trypsin Inhibitors

An overproduction of proinflammatory mediators in severe acute pancreatitis contributes to the systemic inflammatory response, which may lead to multiorgan damage and even death. Thus, inflammatory cytokines, e.g., tumor necrosis factor (TNF)-α and interleukin (IL)-1β, may be novel targets for the treatment of acute pancreatitis. The aim of this study was to investigate the therapeutic effects of pomalidomide (or CC-4047), a thalidomide analog and immunomodulatory agent, in acute pancreatitis.. Acute pancreatitis was induced in C57BL/6 mice by intraperitoneal administration of cerulein (100 μg/kg/h × 8). Pomalidomide was administered (0.5 mg/kg orally) 1 h before the first or 1 h after the last cerulein administration. The severity of the acute pancreatitis was evaluated biochemically and morphologically.. Pretreatment with pomalidomide significantly reduced the plasma levels of amylase and lipase; the histological injury; and the expression of TNF-α, IL-1β, monocyte chemotactic protein-1 (MCP-1), and inducible nitric oxide synthase (iNOS) in cerulein-induced acute pancreatitis. Post-treatment with pomalidomide also decreased the cerulein-induced elevation of plasma amylase and lipase and decreased the pancreatic damage.. Treatment with pomalidomide ameliorated the severity of cerulein-induced acute pancreatitis in mice. Our data suggest that pomalidomide may become a new therapeutic agent in future clinical trials for the treatment of acute pancreatitis.

Topics: Acute Disease; Administration, Oral; Amylases; Animals; Ceruletide; Disease Models, Animal; Immunologic Factors; Lipase; Male; Mice; Mice, Inbred C57BL; Pancreatitis; Severity of Illness Index; Thalidomide

Acute pancreatitis is characterized by early activation of intracellular proteases followed by acinar cell death and inflammation. Activation of damage-associated molecular pattern (DAMP) receptors and a cytosolic complex termed the inflammasome initiate forms of inflammation. In this study, we examined whether DAMP-receptors and the inflammasome provide the link between cell death and the initiation of inflammation in pancreatitis.. Acute pancreatitis was induced by caerulein stimulation in wild-type mice and mice deficient in components of the inflammasome (apoptosis-associated speck-like protein containing a caspase recruitment domain [ASC], NLRP3, caspase-1), Toll-like receptor 9 (TLR9), or the purinergic receptor P2X(7). Resident and infiltrating immune cell populations and pro-interleukin-1β expression were characterized in control and caerulein-treated adult murine pancreas. TLR9 expression was quantified in pancreatic cell populations. Additionally, wild-type mice were pretreated with a TLR9 antagonist before induction of acute pancreatitis by caerulein or retrograde bile duct infusion of taurolithocholic acid 3-sulfate.. Caspase-1, ASC, and NLRP3 were required for inflammation in acute pancreatitis. Genetic deletion of Tlr9 reduced pancreatic edema, inflammation, and pro-IL-1β expression in pancreatitis. TLR9 was expressed in resident immune cells of the pancreas, which are predominantly macrophages. Pretreatment with the TLR9 antagonist IRS954 reduced pancreatic edema, inflammatory infiltrate, and apoptosis. Pretreatment with IRS954 reduced pancreatic necrosis and lung inflammation in taurolithocholic acid 3-sulfate-induced acute pancreatitis.. Components of the inflammasome, ASC, caspase-1, and NLRP3, are required for the development of inflammation in acute pancreatitis. TLR9 and P2X(7) are important DAMP receptors upstream of inflammasome activation, and their antagonism could provide a new therapeutic strategy for treating acute pancreatitis.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Apoptosis; Apoptosis Regulatory Proteins; CARD Signaling Adaptor Proteins; Carrier Proteins; Caspase 1; Ceruletide; Cytoskeletal Proteins; Disease Models, Animal; DNA; Inflammasomes; Interleukin-1; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Necrosis; Neutrophil Infiltration; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreas; Pancreatitis; Pneumonia; Protein Precursors; Purinergic P2X Receptor Antagonists; Receptors, Purinergic P2X7; RNA, Messenger; Severity of Illness Index; Signal Transduction; Taurolithocholic Acid; Toll-Like Receptor 9

Multiple organ dysfunction is the main cause of death in severe acute pancreatitis. Primary mitochondrial dysfunction plays a central role in the development and progression of organ failure in critical illness. The present study investigated mitochondrial function in seven tissues during early experimental acute pancreatitis.. Twenty-eight male Wistar rats (463 ± 2 g; mean ± SEM) were studied. Group 1 (n= 8), saline control; Group 2 (n= 6), caerulein-induced mild acute pancreatitis; Group 3 (n= 7) sham surgical controls; and Group 4 (n= 7), taurocholate-induced severe acute pancreatitis. Animals were euthanased at 6 h from the induction of acute pancreatitis and mitochondrial function was assessed in the heart, lung, liver, kidney, pancreas, duodenum and jejunum by mitochondrial respirometry.. Significant early mitochondrial dysfunction was present in the pancreas, lung and jejunum in both models of acute pancreatitis, however, the Heart, liver, kidney and duodenal mitochondria were unaffected.. The present study provides the first description of early organ-selective mitochondrial dysfunction in the lung and jejunum during acute pancreatitis. Research is now needed to identify the underlying pathophysiology behind the organ selective mitochondrial dysfunction, and the potential benefits of early mitochondrial-specific therapies in acute pancreatitis.

Topics: Acute Disease; Animals; Biomarkers; Cell Respiration; Ceruletide; Disease Models, Animal; Energy Metabolism; Jejunum; Lung; Male; Mitochondria; Mitochondrial Diseases; Multiple Organ Failure; Pancreas; Pancreatitis; Rats; Rats, Wistar; Severity of Illness Index; Taurocholic Acid; Time Factors

The cellular mediators of acute pancreatitis are incompletely understood. Dendritic cells (DCs) can promote or suppress inflammation, depending on their subtype and context. We investigated the roles of DC in development of acute pancreatitis.. Acute pancreatitis was induced in CD11c.DTR mice using caerulein or L-arginine; DCs were depleted by administration of diphtheria toxin. Survival was analyzed using Kaplan-Meier method.. Numbers of major histocompatibility complex II(+)CD11c(+) DCs increased 100-fold in pancreata of mice with acute pancreatitis to account for nearly 15% of intrapancreatic leukocytes. Intrapancreatic DCs acquired a distinct immune phenotype in mice with acute pancreatitis; they expressed higher levels of major histocompatibility complex II and CD86 and increased production of interleukin-6, membrane cofactor protein-1, and tumor necrosis factor-α. However, rather than inducing an organ-destructive inflammatory process, DCs were required for pancreatic viability; the exocrine pancreas died in mice that were depleted of DCs and challenged with caerulein or L-arginine. All mice with pancreatitis that were depleted of DCs died from acinar cell death within 4 days. Depletion of DCs from mice with pancreatitis resulted in neutrophil infiltration and increased levels of systemic markers of inflammation. However, the organ necrosis associated with depletion of DCs did not require infiltrating neutrophils, activation of nuclear factor-κB, or signaling by mitogen-activated protein kinase or tumor necrosis factor-α.. DCs are required for pancreatic viability in mice with acute pancreatitis and might protect organs against cell stress.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Dendritic Cells; Diphtheria Toxin; Disease Models, Animal; Early Growth Response Protein 1; Interleukin-6; Kaplan-Meier Estimate; Male; Mice; Mice, Inbred C57BL; Pancreas; Pancreatitis; Phenotype; Time Factors; Tissue Survival

Reactive oxygen radicals, pro-inflammatory mediators and cytokines have been implicated in caerulein induced acute pancreatitis. Nordihydroguaiaretic acid (NDGA), a plant lignin, has marked anti-inflammatory properties. The present study aimed to investigate the possible protective effect of NDGA against caerulein induced pancreatitis. Acute pancreatitis was induced by intraperitoneal administration of eight doses of caerulein in male swiss albino mice. NDGA was administered after 9 h of acute pancreatitis induction. Pancreatic damage and the protective effect of NDGA were assessed by oxidative stress parameters and histopathology of pancreas. The mRNA expression of heat shock proteins (DNAJ C15 and HSPD1) was examined by real-time RT-PCR analysis. Expression of HSP 27, NF-κB, TNF-α, p-p38, Bcl-2, p-PP2A, procaspase-3, caspase-3 and histone modifications were examined by western blotting. NDGA attenuated the oxidative stress, led to increased plasma α-amylase and decreased IGF-1 in AP mice. It modulated the mRNA and protein levels of heat shock proteins and reduced the expression of NF-κB, TNF-α and p-p38. It increased the number of TUNEL positive apoptotic cells in the pancreas of AP mice. In addition, NDGA prevented the changes in modifications of histone H3 in acute pancreatitis. To best of our knowledge, this is the first report which suggests that NDGA prevents the progression of acute pancreatitis by involving alteration of histone H3 modifications and modulating the expression of genes involved in inflammatory/apoptotic cascade, which may be responsible for decreased necrosis and increased apoptosis in this model of acute pancreatitis.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Apoptosis; Caspase 3; Ceruletide; Chaperonin 60; Histones; In Situ Nick-End Labeling; Inflammation; Lipoxygenase Inhibitors; Male; Masoprocol; Mice; Mitochondrial Proteins; NF-kappa B; Oxidative Stress; Pancreas; Pancreatitis; Real-Time Polymerase Chain Reaction; Signal Transduction; Tumor Necrosis Factor-alpha

To study the potential role of tumor necrosis factor receptor-associated factor 6 (TRAF6) as the key adaptor of the toll-like receptor 4 (TLR4) signaling pathway in acute pancreatitis (AP) in mice.. Acute pancreatitis was induced by 7 intraperitoneal injections of cerulein in TLR4-deficient (TLR4-Def) and TLR4 wild-type (TLR4-WT) mice. Inflammatory severity was scored and evaluated based on pathological study. TRAF6 expression was determined by reverse transcriptase polymerase chain reaction, Western blot, and immunohistochemistry.. Acute pancreatitis was successfully induced in both mice strains, but the inflammatory progression was different. In TLR4-Def mice, pancreatic inflammation was blunt and mild first, then became increasingly intensive and peaked at the later stage, whereas in the TLR4-WT mice, the response was fast initiated and peaked at the early stage of AP, then alleviated gradually. TRAF6 expression in TLR4-Def mice was significantly higher than that in the TLR4-WT mice. Immunohistochemistry located TRAF6 expressed mainly in the pancreatic acinar cells.. The TLR4-TRAF6 signaling pathway is critically involved in AP. Other signaling pathways beyond TLR4 may participate in the pancreatic inflammatory process via TRAF6. As a convergence point of the TLR4-dependent and the TLR4-independent signaling pathways, TRAF6 plays an important role in AP.

Topics: Acute Disease; Animals; Ceruletide; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Pancreatitis; Signal Transduction; TNF Receptor-Associated Factor 6; Toll-Like Receptor 4

The inflammatory response during pancreatitis regulates necrotic and apoptotic rates of parenchymal cells. Neutrophil depletion by use of anti-polymorphonuclear serum (anti-PMN) increases apoptosis in experimental pancreatitis but the mechanism has not been determined. Our study was designed to investigate signaling mechanisms in pancreatic parenchymal cells regulating death responses with neutrophil depletion. Rats were neutrophil depleted with anti-PMN treatment. Then cerulein pancreatitis was induced, followed by measurements of apoptosis signaling pathways. There was greater activation of executioner caspases-3 in the pancreas with anti-PMN treatment compared with control. There were no differences between these groups of animals in mitochondrial cytochrome c release or in activities of initiator caspase-8 and -9. However, there was greater activation of caspase-2 with anti-PMN treatment during cerulein pancreatitis. The upstream regulation of caspases-2 includes p53, which was increased; the p53 negative regulator, Mdm2, was decreased by anti-PMN treatment during cerulein pancreatitis. In vitro experiments using isolated pancreatic acinar cells a pharmacological inhibitor of Mdm2 increased caspase-2/-3 activities, and an inhibitor of p53 decreased these activities during cholecystokinin-8 treatment. Furthermore, experiments using the AR42J cell line Mdm2 small interfering RNA (siRNA) increased caspase-2/-3 activities, and p53 siRNA decreased these activities during cholecystokinin-8 treatment. These results suggest that during acute pancreatitis the inflammatory response inhibits apoptosis. The mechanism of this inhibition involves caspase-2 and its upstream regulation by p53 and Mdm2. Because previous findings indicate that promotion of apoptosis decreases necrosis and severity of pancreatitis, these results suggest that strategies to inhibit Mdm2 or activate p53 will have beneficial effects for treatment of pancreatitis.

Topics: Acute Disease; Animals; Apoptosis; Caspase 3; Caspase 8; Caspase 9; Caspases; Cells, Cultured; Ceruletide; Cysteine Endopeptidases; Cytochromes c; Disease Models, Animal; Male; Necrosis; Neutrophils; Pancreatitis; Proto-Oncogene Proteins c-mdm2; Rats; Rats, Sprague-Dawley; RNA, Small Interfering; Tumor Suppressor Protein p53

The mechanisms by which reflux of bile acids into the pancreas induces pancreatitis are unknown. We reasoned that key events responsible for this phenomenon might be mediated by Gpbar1, a recently identified and widely expressed G-protein-coupled, cell surface bile acid receptor.. Acute pancreatitis was induced in wild-type and Gpbar1(-/-) mice by either retrograde ductal infusion of taurolithocholic acid-3-sulfate (TLCS) or supramaximal secretagogue stimulation with caerulein. In vitro experiments were performed in which acini obtained from wild-type and Gpbar1(-/-) mice were exposed to either submicellar concentrations of TLCS (200-500 microM) or a supramaximally stimulating concentration of caerulein (10 nM).. Gpbar1 is expressed at the apical pole of acinar cells and its genetic deletion is associated with reduced hyperamylasemia, edema, inflammation, and acinar cell injury in TLCS-induced, but not caerulein-induced, pancreatitis. In vitro, genetic deletion of Gpbar1 is associated with markedly reduced generation of pathological calcium transients, intracellular activation of digestive zymogens, and cell injury when these responses are induced by exposure to TLCS, but not when they are induced by exposure to caerulein.. Gpbar1 may play a critical role in the evolution of bile-acid-induced pancreatitis by coupling exposure to bile acids with generation of pathological intracellular calcium transients, intra-acinar cell zymogen activation, and acinar cell injury. Acute biliary pancreatitis may be a "receptor-mediated" disease and interventions that interfere with Gpbar1 function might prove beneficial in the treatment and/or prevention of biliary acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Bile Acids and Salts; Calcium Signaling; Ceruletide; Disease Models, Animal; Enzyme Precursors; GTP-Binding Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis; Receptors, G-Protein-Coupled; Severity of Illness Index; Taurolithocholic Acid

Nardostachys jatamansi belonging to the family Valerianaceae has been used as a remedy for stomach and skin ailments in Korea. The effect of N. jatamansi on acute pancreatitis (AP) has not been defined. Therefore, we investigated the effect of N. jatamansi on cerulein-induced AP.. In the pretreatment group, N. jatamansi was administered orally to mice at 10 and 20 mg/kg for 5 days, and the mice were intraperitoneally injected with the stable cholecystokinin analogue cerulein hourly for 6 hours. In the posttreatment group, cerulein was injected hourly for 6 hours, and N. jatamansi was administered at the indicated time (1, 3, and 5 hours after the first cerulein injection) and dose (10 and 20 mg/kg) during the cerulein injection. Blood samples were taken 6 hours later to determine the serum amylase, the lipase, and the cytokine levels. The pancreas and the lung were rapidly removed for morphologic examination, myeloperoxidase assay, and real-time reverse transcription polymerase chain reaction.. Nardostachys jatamansi treatment attenuated the AP, as shown by the histological examination results of the pancreas and the lung, reductions in pancreatic edema, neutrophil infiltration, serum amylase and lipase levels, serum cytokine levels, and messenger RNA expressions of inflammatory mediators.. These results suggest that N. jatamansi attenuates the severity of AP and pancreatitis-associated lung injury.

Topics: Acute Disease; Amylases; Animals; Body Weight; Cell Survival; Cells, Cultured; Ceruletide; Cytokines; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Female; Lipase; Mice; Mice, Inbred C57BL; Mitogen-Activated Protein Kinases; Nardostachys; Organ Size; Pancreas; Pancreatitis; Peroxidase; Phytotherapy; Plant Extracts; Reverse Transcriptase Polymerase Chain Reaction

Acute pancreatitis (AP) is characterized by pancreatic microcirculatory and secretory disturbances. As galanin can modulate pancreatic vascular perfusion, we sought to determine if galanin plays a role in AP.. Acute pancreatitis was induced in wild-type and galanin gene knockout mice by intraperitoneal injections of cerulein. The severity of AP was evaluated (plasma amylase and lipase, myeloperoxidase activity, and acinar cell necrosis) with and without treatment with galanin or the antagonist galantide. Galanin receptor messenger RNA expression in mouse pancreas was measured by reverse transcription-polymerase chain reaction and Western blot analysis.. Galantide ameliorated AP, reducing all indices by 25% to 40%, whereas galanin was without effect. In galanin knockout mice, all indices of AP were reduced 25% to 50% compared with wild-type littermates. Galanin administration to the knockout mice exacerbated AP such that it was comparable with the AP induced in the wild-type mice. Conversely, administration of galantide to the galanin knockout mice did not affect the AP, whereas AP was ameliorated in the wild-type mice. The 3 galanin receptor subtypes are expressed in mouse pancreas, with receptor subtype 3 expression predominating.. These data implicate a role for galanin in AP and suggest a potential clinical application for galanin antagonists in treatment.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Female; Galanin; Mice; Mice, Inbred BALB C; Mice, Knockout; Pancreas; Pancreatitis; Receptors, Galanin; RNA, Messenger; Severity of Illness Index; Substance P

The neuropeptide substance P (SP) has emerged to be an important proinflammatory mediator in acute pancreatitis (AP). The presence of substance P and its receptor, neurokinin-1 receptor (NK1R) has been shown in the pancreas and the pancreatic acinar cells. In this study, we investigated the unexplored mechanisms that mediate SP and NK1R expression using an in vitro AP model. Pancreatic acinar cells were obtained from pancreas of male Swiss mice. Isolated cells were treated with caerulein to mimic secretagogue pancreatitis. A concentration-dependent study that subjected the cells to 60 min of stimulation by caerulein showed that SP and the transcript from its gene preprotachykinin-A (PPT-A), and NK1R were up-regulated at a supraphysiological concentration of 10(-7) M. A concentration-dependent study on intracellular kinases, extracellular signal-regulated kinase (ERK1/2), and c-Jun N-terminal kinase (JNK) and also transcription factors nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) showed that they were activated when the caerulein concentration was 10(-7) M. Inhibition of JNK reversed the up-regulation of PPT-A, SP, and NK1R. However, inhibition of ERK1/2 reversed the up-regulation of NK1R but not of PPT-A and SP. Furthermore, we found that specific ERK1/2 and JNK inhibitors reduce NF-kappaB and AP-1 activity. Taken together, our results suggest that supraphysiological concentrations of caerulein up-regulate the expression of SP and NK1R in pancreatic acinar cells, and the signaling molecules that are involved in this up-regulation include ERK1/2, JNK, NF-kappaB, and AP-1.

Topics: Acute Disease; Animals; Ceruletide; JNK Mitogen-Activated Protein Kinases; Male; Mice; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; NF-kappa B; Pancreas; Pancreatitis; Phosphorylation; Protein Precursors; Receptor, Cholecystokinin A; Receptors, Neurokinin-1; Signal Transduction; Substance P; Tachykinins; Transcription Factor AP-1

We surveyed changes of the gene expression profile in caerulein-exposed pancreas using Affymetrix GeneChip system (39,000 genes). Up-regulation of genes coding for claudin 4, claudin 7, F11 receptor, cadherin 1, integrin beta 4, syndecan 1, heat shock proteins b1/90aa1, Serpinb6a, Serpinb6b, Serpinb9, Bax, Bak1, calpain 2, calpain 5, microtubule-associated protein 1 light chain 3 alpha, S100 calcium-binding proteins A4/A10 were found in mouse pancreas exposed to caerulein for 12h. In contrast, the anti-apoptotic gene Bcl2 was down-regulated. The functions of these genes concern tight junction formation, cell-cell/cell-matrix adhesions, stress response, protease inhibition, apoptosis, autophagy, and regulation of cytoskeletal dynamics. Caerulein-exposed pancreatic acinar cells were immunohistochemically stained for claudin 4, cadherin 1, integrin beta 4, heat shock protein b1, and Serpinb6a. In conclusion, we have newly identified a set of genes that are likely to be involved in endogenous self-protection mechanisms against acute pancreatitis.

Topics: Acute Disease; Animals; Apoptosis Regulatory Proteins; Autophagy; Ceruletide; Gene Expression; Gene Expression Profiling; Heat-Shock Proteins; Intercellular Junctions; Male; Mice; Mice, Inbred Strains; Pancreatitis; Proteinase Inhibitory Proteins, Secretory; S100 Proteins

To analyze susceptibility to acute pancreatitis, five mouse strains including Japanese Fancy Mouse 1 (JF1), C57BL/6J, BALB/c, CBA/J, and C3H/HeJ were treated with either a cholecystokinin analog, cerulein, or a choline-deficient, ethionine-supplemented (CDE) diet. The severity of acute pancreatitis induced by cerulein was highest in C3H/HeJ and CBA/J, moderate in BALB/c, and mildest in C57BL/6J and JF1. Basal protein expression levels of the serine protease inhibitor, Kazal type 3 (Spink3) were higher in JF1 and C57BL/6J mice than those of the other three strains under normal feeding conditions. After treatment with cerulein, expression level of Spink3 increased remarkably in JF1 and mildly in C57BL/6J, BALB/c, CBA/J, and C3H/HeJ strains. Increased proteinase, serine, 1 (Prss1) protein expression accompanied by increased trypsin activity with cerulein treatment was observed in susceptible strains such as CBA/J and C3H/HeJ. Similar results were obtained with a CDE diet. In the 3 kb Spink3 promoter region, 92 or 8 nucleotide changes were found in JF1 or C3H vs C57BL/6J, respectively, whereas in the Prss1 promoter region 39 or 46 nucleotide changes were found in JF1 or C3H vs C57BL/6J, respectively. These results suggest that regulation of Prss1 and Spink3 expression is involved in the susceptibility to experimentally induced pancreatitis. The JF1 strain, which is derived from the Japanese wild mouse, will be useful to examine new mechanisms that may not be found in other laboratory mouse strains.

Topics: Acute Disease; Animals; Base Sequence; Blotting, Northern; Blotting, Western; Ceruletide; Diet; Gene Expression; Genetic Predisposition to Disease; Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Inbred CBA; Molecular Sequence Data; Pancreatitis; Prostatic Secretory Proteins; Sequence Homology, Nucleic Acid; Severity of Illness Index; Species Specificity; Trypsin; Trypsin Inhibitor, Kazal Pancreatic

We have previously shown that galantide ameliorates mild acute pancreatitis (AP), and the salivary tripeptide analogue, feG, ameliorates severe AP in mice. In this study, we compared the efficacy of combining galantide and feG with that of the individual agents in treating mild AP induced in mice with 7-hourly caerulein injections. Galantide was co-administered with each caerulein injection commencing with the first injection. feG was co-administered with the first injection of caerulein as a single intraperitoneal injection. Combination of the agents was also administered. Control animals received galantide, feG, or saline alone. Pancreata were harvested for histological examination and estimation of myeloperoxidase (MPO) activity. Plasma enzyme activities were measured. Galantide significantly reduced AP-induced hyperenzymemia by 41-49%. The combination of galantide and feG significantly reduced AP-induced hyperenzymemia by 39-40%, whereas feG alone was without effect. Plasma enzyme activity in the control groups was comparable with pre-treatment activity. Galantide, feG, and their combination significantly reduced MPO activity by 83, 44 and 74% respectively, and % abnormal acinar cells by 32, 29 and 36% respectively. This study demonstrates for the first time the beneficial effect of feG in mild caerulein-induced AP. Moreover the data indicate that the hyperenzymemia in mild caerulein-induced AP at 12h possibly reflect a larger secretory component as compared to enzyme release due to neutrophil-mediated acinar cell damage. The effects of the treatment with both peptides indicate a possible role for galantide in modulating neutrophil chemotaxis/activation and supports the hypothesis that galantide may influence neurogenic inflammation in AP.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Drug Therapy, Combination; Galanin; Lipase; Mice; Oligopeptides; Pancreas; Pancreatitis; Stereoisomerism; Substance P

Topics: Acute Disease; Animals; Ceruletide; Cytoplasm; Dyneins; Golgi Apparatus; Kinesins; Male; Pancreatitis; Rats; Rats, Wistar

This study was designed to evaluate the protective effect of the cysteinyl leukotriene receptor antagonist montelukast against pancreatic injury during acute pancreatitis.. Acute pancreatitis was induced in rats by 20-μg/kg (intraperitoneal) cerulein given at 1-hour intervals within 4 hours. Montelukast was administered intraperitoneally at a dose of 10 mg/kg 15 minutes before the first cerulein injection. Six hours after the cerulein or saline injections, the animals were killed by decapitation. Blood samples were collected to analyze amylase, lipase, and the proinflammatory cytokines tumor necrosis factor α and interleukin 1β. Pancreas tissues were taken for the determination of tissue glutathione and malondialdehyde levels and Na,K-adenosine triphosphatase and myeloperoxidase activities. The extent of tissue injury was analyzed microscopically.. Acute pancreatitis caused significant decreases in tissue glutathione level and Na,K-adenosine triphosphatase activity, which were accompanied with significant increases in the pancreatic malondialdehyde level, myeloperoxidase activity, and plasma cytokine level. On the other hand, montelukast treatment reversed all these biochemical indices and histopathological alterations that were induced by cerulein.. These results suggest that cysteinyl leukotrienes may be involved in the pathogenesis of acute pancreatitis and that the cysteinyl leukotriene receptor antagonist, montelukast, might be of therapeutic value for treatment of acute pancreatitis.

Topics: Acetates; Acute Disease; Animals; Ceruletide; Cyclopropanes; Cytokines; Female; Glutathione; Leukotriene Antagonists; Leukotriene B4; Lipid Peroxidation; Male; Pancreas; Pancreatitis; Peroxidase; Quinolines; Rats; Rats, Sprague-Dawley; Receptors, Leukotriene; Sodium-Potassium-Exchanging ATPase; Sulfides

Previously, we found that the University of North Carolina cystic fibrosis (UNC-CF) mouse had more severe experimental acute pancreatitis (AP) than wild-type (WT) mice characterized by exuberant pancreatic inflammation and impaired acinar apoptosis. Because exon 10 CFTR gene mutations exhibit different phenotypes in tissues such as the mouse lung, we tested the hypothesis that DeltaF508-CF mice also develop severe AP associated with an antiapoptotic acinar phenotype, which requires indirect effects of the extracellular milieu. We used cerulein hyperstimulation models of AP. More severe pancreatitis occurred in cerulein-injected DeltaF508-CF vs. WT mice based on histological severity (P < 0.01) and greater neutrophil sequestration [P < 0.0001; confirmed by myeloperoxidase activity (P < 0.005)]. In dispersed acini cerulein-evoked necrosis was greater in DeltaF508-CF acini compared with WT (P < 0.05) and in WT acini pretreated with CFTR(inh)-172 compared with vehicle (P < 0.05). Cerulein-injected DeltaF508-CF vs. WT mice had less apoptosis based on poly(ADP-ribose) polymerase (PARP) cleavage (P < 0.005), absent DNA laddering, and reduced terminal deoxynucleotidyltransferase biotin-dUTP nick end labeling (TUNEL) staining (P < 0.005). Unexpectedly, caspase-3 activation was greater in DeltaF508-CF vs. WT acini at baseline (P < 0.05) and during AP (P < 0.0001). Downstream, DeltaF508-CF pancreas overexpressed the X-linked inhibitor of apoptosis compared with WT (P < 0.005). In summary, the DeltaF508-CF mutation, similar to the UNC-CF "null" mutation, causes severe AP characterized by an exuberant inflammatory response and impaired acinar apoptosis. Enhanced acinar necrosis in DeltaF508-CF occurs independently of extracellular milieu and correlates with loss of CFTR-Cl conductance. Although both exon 10 models of CF inhibit acinar apoptosis execution, the DeltaF508-CF mouse differs by increasing apoptosis signaling. Impaired transduction of increased apoptosis signaling in DeltaF508-CF acini may be biologically relevant to the pathogenesis of AP associated with CFTR mutations.

Topics: Acute Disease; Animals; Apoptosis; Caspase 3; Ceruletide; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Enzyme Activation; Genotype; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Mutation; Necrosis; Pancreas; Pancreatitis; Peroxidase; RNA, Messenger; Severity of Illness Index; Signal Transduction; X-Linked Inhibitor of Apoptosis Protein

We used a peptidomic approach for the analysis of the low molecular weight proteome in rat pancreatic tissue extracts. The goal was to develop a method that allows identifying endogenous peptides produced in the pancreas in the course of acute pancreatitis. The workflow combines peptides enrichment by centrifugal ultrafiltration, fractionation by isoelectric focusing, and LC-MS/MS analysis without prior enzymatic digestion. The method was assessed on pancreatic extracts from 3 rats with caerulein-induced pancreatitis and 3 healthy controls. A qualitative analysis of the peptide patterns obtained from the different samples was performed to determine the main biological processes associated to the identified peptides. Comparison of peptidomic and immunoblot data for alpha-tubulin, beta-tubulin and coatomer gamma showed that the correlation between the number of identified peptides and the protein abundance was variable. Nevertheless, peptidomic analysis highlighted inflammatory and stress proteins, which peptide pattern was related to acute pancreatitis pathobiology. For these proteins, the higher number of peptides in pancreatitis samples reflected an increase in protein abundance. Moreover, for murinoglobulin-1 or carboxypeptidase B, peptide pattern could be related to protein function. These data suggest that peptidomic analysis is a complementary approach to proteomics for investigating pathobiological processes involved in acute pancreatitis.

Topics: Acute Disease; Amino Acid Sequence; Animals; Ceruletide; Chromatography, Liquid; Disease Models, Animal; Heat-Shock Proteins; Immunoblotting; Inflammation; Male; Molecular Sequence Data; Molecular Weight; Pancreatitis; Peptides; Proteins; Proteome; Proteomics; Rats; Rats, Sprague-Dawley; Tandem Mass Spectrometry

We have previously shown that galantide, a non-specific galanin receptor antagonist, ameliorates acute pancreatitis (AP) induced in mice. Octreotide, a somatostatin analogue, has been used in the treatment of AP with inconsistent outcomes. This study set out to compare the efficacy of a combined treatment of galantide and octreotide with the efficacy of each agent individually in experimental AP.. Acute pancreatitis was induced in mice with 7-hourly caerulein injections. Galantide and/or octreotide were co-administered with each caerulein injection commencing with the first injection. Control animals received galantide, octreotide or saline alone. Pancreata were harvested for histological examination and estimation of myeloperoxidase (MPO) activity. Plasma amylase and lipase activities were measured.. Galantide significantly reduced AP-induced hyperenzymaemia by 39-45%. Octreotide alone, or in combination with galantide, did not significantly alter AP-induced hyperenzymaemia. Plasma enzyme activity in the control groups was comparable with pre-treatment activity. Galantide and octreotide administered individually reduced MPO activity by 79% and 50%, respectively; however their combination was without effect. Galantide, octreotide and their combination significantly reduced the percentage of abnormal acinar cells by 28-45%.. Treatment with galantide alone ameliorated most of the indices of AP studied, whereas treatment with octreotide reduced pancreatic MPO activity and acinar cell damage. Combining the two peptides appears to negate their individual benefits, which suggests an interaction in their mechanism of action.

Topics: Acute Disease; Amylases; Animals; Biomarkers; Ceruletide; Disease Models, Animal; Drug Therapy, Combination; Galanin; Lipase; Male; Mice; Octreotide; Pancreas; Pancreatitis; Peroxidase; Substance P; Time Factors

Recent studies have shown that pretreatment with ghrelin exhibits protective effect in the gut. Administration of ghrelin reduces gastric mucosal damage, as well as inhibits the development of experimental pancreatitis. However, this protective effect requires administration of ghrelin before gastric or pancreatic damage and thus has a limited clinical value. The aim of present study was to assess the influence of ghrelin administered after development of acute pancreatitis on the course of this disease. Acute pancreatitis was induced by cerulein. Ghrelin was administered twice a day for 1, 2, 4, 6 or 9 days at the dose of 4, 8 or 16 nmol/kg/dose. The first dose of ghrelin was given 24 hours after last injection of cerulein. The severity of acute pancreatitis was assessed between 0 h and 10 days after cessation of cerulein administration. Administration of caerulein led to the development of acute edematous pancreatitis and maximal severity of this disease was observed 24 hours after induction of pancreatitis. Treatment with ghrelin reduced morphological signs of pancreatic damage such as pancreatic edema, leukocyte infiltration and vacuolization of acinar cells, and led to earlier regeneration of the pancreas. Also biochemical indexes of the severity of acute pancreatitis, serum activity of lipase and amylase were significantly reduced in animals treated with ghrelin. These effects were accompanied by an increase in the pancreatic DNA synthesis and a decrease in serum level of pro-inflammatory interleukin-1b. Administration of ghrelin improved pancreatic blood flow in rats with acute pancreatitis. We conclude that: (1) treatment with ghrelin exhibits therapeutic effect in caerulein-induced experimental acute pancreatitis; (2) this effect is related, at least in part, to the improvement of pancreatic blood flow, reduction in proinflammatory interleukin-1beta and stimulation of pancreatic cell proliferation.

Topics: Acute Disease; Animals; Blood Flow Velocity; Blood Glucose; Cell Proliferation; Ceruletide; Dose-Response Relationship, Drug; Down-Regulation; Ghrelin; Inflammation Mediators; Insulin; Interleukin-1beta; Male; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Up-Regulation

Acute pancreatitis is an inflammatory disease of the pancreas, which can result in serious morbidity or death. Acute pancreatitis severity can be reduced in experimental models by preconditioning animals with a short hyperthermia prior to disease induction. Heat shock proteins 27 and 70 are key effectors of this protective effect. In this study, we performed a comparative proteomic analysis using a combination of liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis and isobaric tagging to investigate changes in pancreatic proteins expression that were associated with thermal stress, both in healthy rats and in a model of caerulein-induced pancreatitis. In agreement with previous studies, we observed modulation of heat shock and inflammatory proteins expression in response to heat stress or pancreatitis induction. We also identified numerous other proteins, whose pancreatic level changed following pancreatitis induction, when acute pancreatitis severity was reduced by prior thermal stress, or in healthy rats in response to hyperthermia. Interestingly, we showed that the expression of various proteins associated with the secretory pathway was modified in the different experimental models, suggesting that modulation of this process is involved in the protective effect against pancreatic tissue damage.

Topics: Acute Disease; Animals; Ceruletide; Fever; Heat-Shock Response; Pancreatitis; Protective Agents; Proteomics; Rats

Proteinase-activated receptor-2 (PAR2) and transient receptor potential vanilloid-1 (TRPV1) are co-localized in the primary afferents, and the trans-activation of TRPV1 by PAR2 activation is involved in processing of somatic pain. Given evidence for contribution of PAR2 to pancreatic pain, the present study aimed at clarifying the involvement of TRPV1 in processing of pancreatic pain by the proteinase/PAR2 pathway in mice.. Acute pancreatitis was created by repeated administration of cerulein in conscious mice, and the referred allodynia/hyperalgesia was assessed using von Frey filaments. Injection of PAR2 agonists into the pancreatic duct was achieved in anesthetized mice, and expression of Fos in the spinal cord was determined by immunohistochemistry.. The established referred allodynia/hyperalgesia following cerulein treatment was abolished by post-treatment with nafamostat mesilate, a proteinase inhibitor, and with capsazepine, a TRPV1 antagonist, in mice. Injection of trypsin, an endogenous PAR2 agonist, or SLIGRL-NH(2), a PAR2-activating peptide, into the pancreatic duct caused expression of Fos protein in the spinal superficial layers at T8-T10 levels in the mice. The spinal Fos expression caused by trypsin and by SLIGRL-NH(2) was partially blocked by capsazepine, the former effect abolished by nafamostat mesilate.. Our data thus suggest that the proteinase/PAR2/TRPV1 cascade might impact pancreatic pain, in addition to somatic pain, and play a role in the maintenance of pancreatitis-related pain in mice.

Topics: Acute Disease; Animals; Benzamidines; Capsaicin; Ceruletide; Disease Models, Animal; Gene Expression Regulation; Guanidines; Hyperalgesia; Male; Mice; Oligopeptides; Pain; Pancreatitis; Proto-Oncogene Proteins c-fos; Receptor, PAR-2; Spinal Cord; TRPV Cation Channels

The endoplasmic reticulum (ER) is abundant in the acinar cells of the exocrine pancreas. To test the role of ER homeostasis in acute pancreatitis, we manipulated GRP78 levels, a major ER chaperone, in mice. Grp78(+/+) and (+/-) littermates were fed either a regular diet (RD) or a high-fat diet. Acinar cells were examined for ER structure by electron microscopy, and ER chaperone levels were assessed by immunoblotting. Pancreatitis was induced by cerulein injection, and multiple pathological parameters were analyzed. Grp78(+/-) mice showed decreased GRP78 expression in acinar cells. Exocrine pancreata of RD-fed Grp78(+/-) mice in an outbred C57BL/6 × 129/sv genetic background exhibited ER lumen dilation, a reduction in chaperones calnexin (CNX) and calreticulin (CRT), and exacerbated pancreatitis associated with high CHOP induction. With the high-fat diet regimen, Grp78 heterozygosity triggered GRP94 up-regulation and restoration of GRP78, CNX, and CRT to wild-type levels, corresponding with mitigated pancreatitis on cerulein insult. Interestingly, after backcrossing into the C57BL/6 background, RD-fed Grp78(+/-) mice exhibited an increase in GRP94 and levels of CNX and CRT equivalent to wild type, associated with decreased experimental pancreatitis severity. Administration of a chemical chaperone, 4-phenolbutyrate, was protective against cerulein-induced death. Thus, in exocrine pancreata, Grp78 heterozygosity regulates ER chaperone balance, in dietary- and genetic background-dependent manners, and improved ER protein folding capacity might be protective against pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Dietary Fats; Endoplasmic Reticulum; Endoplasmic Reticulum Chaperone BiP; Gastrointestinal Agents; Heat-Shock Proteins; Heterozygote; Mice; Mice, Inbred C57BL; Mice, Transgenic; Molecular Chaperones; Pancreas, Exocrine; Pancreatitis; Stress, Physiological; Unfolded Protein Response

Toll-like receptor 4 (TLR4) plays an important role in the occurrence and development of acute pancreatitis (AP). Apoptosis of pancreatic cells is closely related to the severity of AP. TLR4 is known to induce apoptosis in some cell types and therefore it is of importance to investigate potential associations between TLR4 activity and apoptosis in the setting of AP.. A total of 50 wild-type (C57BL/10J) and TLR4-deficient (C57BL/10ScNJ) mice were divided into three groups: 2-hour, 4-hour, and control groups. Each group was divided into two equal subgroups: TLR4-wild-type mice and TLR4-deficient mice. AP was experimentally induced by 7 intraperitoneal injections of 50 μg/kg cerulein at hourly intervals. Control mice received 7 injections of equal volumes of saline. The severity of pancreatic injury during AP was assessed by serum amylase concentration and histopathology. The level of apoptosis of pancreatic cells in response to AP was evaluated by calculating the apoptotic index (AI) and comparing the expression levels of cytochrome C and Fas-associated protein with death domain (FADD) between TLR4-wild-type and TLR4-deficient mice at 2 time points.. The AI was found to be significantly lower in the pancreas of TLR4-deficient mice with AP compared to TLR4-wild-type mice at two hours after the last treatment injection. Enzyme-linked immunosorbent assay and real-time reverse transcription-polymerase chain reaction also revealed significantly lower expression of cytochrome C and FADD in the pancreas of TLR4-deficient mice than in TLR4-wild-type animals at the same time point. Serum amylase concentration and morphological severity of AP in pancreatic tissue were found to be similar in the two strains of mice at both time points.. We postulate that TLR4 can mediate apoptosis of pancreatic cells during the early stages of AP, via the activation of both intrinsic and extrinsic apoptotic signaling pathways.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Ceruletide; Fas-Associated Death Domain Protein; Female; In Situ Nick-End Labeling; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Pancreas; Pancreatitis; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Toll-Like Receptor 4

The renin-angiotensin system contributes to pathological processes in a variety of organs. In the pancreas, blocking the angiotensin II (AII) type 1 receptor (AT1) attenuates pancreatic fibrogenesis in animal models of pancreatitis. Because the role of the AII type 2 receptor (AT2) in modulating pancreatic injury is unknown we investigated the role of AT2 in pancreatic injury and fibrosis. Pancreatic fibrosis was induced by repetitive cerulein administration in C57BL/6 wild-type (WT) or AT2-deficient (AT2-/-) mice and assessed by morphology and gene expression at 10 days. There was no difference between WT and AT2-/- mice in the degree of acute pancreatic injury as assessed by amylase release at 9 and 12 h and by histological examination of the pancreas at 12 h. In contrast, parenchymal atrophy and fibrosis were more pronounced in AT2-/- mice compared with WT mice at 10 days. Fibrosis was accompanied by activation of pancreatic stellate cells (PSC) evaluated by Western blot analysis for alpha-smooth muscle actin and by immunocytochemistry; PSC activation was further increased in AT2-/- mice compared with WT mice. The level of pancreatic transforming growth factor-beta1 mRNA and protein after repetitive cerulein treatment was higher in AT2-/- mice than in WT mice. Our results demonstrate that, in contrast to AT1 receptor signaling, AT2 receptor signaling modulates protective antifibrogenic effects in a mouse model of cerulein-induced pancreatic fibrogenesis. We propose that the effects of AII on injury-induced pancreatic fibrosis may be determined by the balance between AT1 and AT2 receptor signaling.

Topics: Actins; Acute Disease; Amylases; Angiotensins; Animals; Ceruletide; Collagen; Disease Models, Animal; Female; Fibrosis; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis; Receptor, Angiotensin, Type 1; Receptor, Angiotensin, Type 2; RNA, Messenger; Severity of Illness Index; Signal Transduction; Time Factors; Transforming Growth Factor beta1

The role of nitric oxide (NO) has been increasingly implicated in the pathophysiology of acute pancreatitis (AP). Studies have shown increased NO production in AP although not all are agreeable on whether NO is beneficial or detrimental in AP. This study aims to profile NO production and NO synthase (NOS) expression in the pancreas and lungs in the progression of AP in mice to gain insights to the role played by different NOS isoforms.. AP was induced in mice by hourly administration of cerulein. NO production was determined by measuring the total nitrite and nitrate (NOx) content while NOS expression was measured by Western blot.. Pancreatic NO production increased sharply and was sustained throughout AP. iNOS expression was greatly increased while eNOS was downregulated at the later stages. In the lungs, there was an unexpected early increase in the constitutive NOS expression; however iNOS was also significantly overexpressed at the later time point along with a significant increase in NO. Acinar cells were found to overproduce NO in response to cerulein hyperstimulation with iNOS again being the major contributor.. These data show that NO production and NOS expression are differentially regulated temporally and in magnitude in the pancreas and lungs in response to cerulein hyperstimulation which suggests differing roles for each NOS isoform. and IAP.

Topics: Acute Disease; Animals; Ceruletide; Lung; Lung Injury; Mice; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Pancreas; Pancreatitis

Inflammatory cytokines and oxidative stress have a central role in the pathogenesis of acute pancreatitis. Propolis is a resinous hive product collected by honeybees from various plant sources and has anti-inflammatory and anti-oxidant effects. The present work aimed to investigate the therapeutic role of ethanolic extract of propolis on a cerulein-induced acute pancreatitis model in rats.. Seventy male Wistar albino rats were used in the study. Acute edematous pancreatitis was induced by subcutaneous cerulein injection (20 microg/kg) four times at one-hour intervals. Ethanolic extract of propolis 300 mg/kg was given subcutaneously at the beginning of the procedure (ethanolic extract of propolis-1 group) or 12 h after the last cerulein injection (ethanolic extract of propolis-2 group). Serum amylase and lipase levels, white blood cell count and serum tumor necrosis factor-alpha levels were measured and pancreatic tissue was evaluated histologically.. In the acute pancreatitis group, serum amylase and lipase levels were found to be elevated and the histopathological evaluation of the tissue revealed massive edema and inflammation with less fatty necrosis when compared to the sham and control groups. Serum amylase and lipase levels and edema formation were significantly decreased in the ethanolic extract of propolis-treated groups (p<0.001). In the ethanolic extract of propolis-2 group, in particular, tissue edema was improved markedly (p=0.001). Tissue inflammation and fatty necrosis were decreased with ethanolic extract of propolis treatment; however, the improvement was not statistically significant.. Treatment with ethanolic extract of propolis improved the biochemical and histopathological findings in a rat model of experimental pancreatitis. Although our findings suggest that ethanolic extract of propolis might be considered an effective agent for the treatment of acute pancreatitis, this notion should be supported with further experimental and clinical investigations.

Topics: Acute Disease; Amylases; Animals; Anti-Infective Agents; Ceruletide; Disease Models, Animal; Edema; Gastrointestinal Agents; Lipase; Male; Pancreas; Pancreatitis; Propolis; Rats; Rats, Wistar; Treatment Outcome

Systemic inflammatory mediators, including the protein high-mobility group box 1 (HMGB1), play an important role in the development of acute pancreatitis. Anticoagulants, such as antithrombin III (AT III), inhibit inflammation resulting from various causes, but their mechanism of action is not well understood. Because acute pancreatitis is a severe inflammatory disease, we hypothesized that AT III would inhibit inflammation and prevent cerulein-induced acute pancreatitis.. Experimental animals received or were saline injected with a bolus of 250 IU/kg of AT III followed by intraperitoneal injections of 50 mg/kg of cerulein. Levels of cytokines (interleukin 6 and tumor necrosis factor alpha), nitric oxide (NO), and HMGB1 were measured in serum and pancreatic tissue at regular intervals for 12 hours after the cerulein injection.. Pancreas histopathology and wet-dry ratio significantly improved in the AT III-injected (250 IU/kg) animals compared with the saline-injected rats. Serum and pancreas HMGB1 levels decreased over time in AT III-treated animals. Antithrombin III also decreased cytokine, NO, and HMGB1 levels during cerulein-induced inflammation. As a result, AT III ameliorated the pathologic pancreas in the rat model of cerulein-induced acute pancreatitis.. Antithrombin III treatment inhibited the secretion of cytokines, NO, and HMGB1 and prevented cerulein-induced acute pancreatitis in the rat model.

Topics: Acute Disease; Animals; Anticoagulants; Antithrombin III; Blotting, Western; Ceruletide; HMGB1 Protein; Immunohistochemistry; Interleukin-6; Male; Nitric Oxide; Pancreas; Pancreatitis; Random Allocation; Rats; Rats, Wistar

Receptor-stimulated Ca(2+) influx is a critical component of the Ca(2+) signal and mediates all cellular functions regulated by Ca(2+). However, excessive Ca(2+) influx is highly toxic, resulting in cell death, which is the nodal point in all forms of pancreatitis. Ca(2+) influx is mediated by store-operated channels (SOCs). The identity and function of the native SOCs in most cells is unknown.. Here, we determined the role of deletion of Trpc3 in mice on Ca(2+) signaling, exocytosis, intracellular trypsin activation, and pancreatitis.. Deletion of TRPC3 reduced the receptor-stimulated and SOC-mediated Ca(2+) influx by about 50%, indicating that TRPC3 functions as an SOC in vivo. The reduced Ca(2+) influx in TRPC3(-/-) acini resulted in reduced frequency of the physiologic Ca(2+) oscillations and of the pathologic sustained increase in cytosolic Ca(2+) levels caused by supramaximal stimulation and by the toxins bile acids and palmitoleic acid ethyl ester. Consequently, deletion of TRPC3 shifted the dose response for receptor-stimulated exocytosis and prevented the pathologic inhibition of digestive enzyme secretion at supramaximal agonist concentrations. Accordingly, deletion of TRPC3 markedly reduced intracellular trypsin activation and excessive actin depolymerization in vitro and the severity of pancreatitis in vivo.. These findings establish the native TRPC3 as an SOC in vivo and a role for TRPC3-mediated Ca(2+) influx in the pathogenesis of acute pancreatitis and suggest that TRPC3 should be considered a target for prevention of pancreatic damage in acute pancreatitis.

Topics: Actins; Acute Disease; Animals; Calcium Signaling; Carbachol; Ceruletide; Cholinergic Agonists; Disease Models, Animal; Dose-Response Relationship, Drug; eIF-2 Kinase; Enzyme Activation; Enzyme Inhibitors; Exocytosis; Indoles; Membrane Potentials; Mice; Mice, Knockout; Pancreas; Pancreatitis; Phosphorylation; Sarcoplasmic Reticulum; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Severity of Illness Index; Sincalide; Taurocholic Acid; TRPC Cation Channels; Trypsin

The cytosolic cysteine protease calpain is implicated in a multitude of cellular functions but also plays a role in cell damage. Our previous results suggest that an activation of calpain accompanied by a decrease in its endogenous inhibitor calpastatin may contribute to pancreatic damage during cerulein-induced acute pancreatitis. The present study aimed at the time course of secretagogue-induced calpain activation and cellular substrates of the protease. Isolated rat pancreatic acini were incubated with a supramaximal concentration of cholecystokinin (0.1 microM CCK) for 30 min in the presence or absence of the calpain inhibitor Z-Val-Phe methyl ester (100 microM ZVP). The activation of calpain and the expression of calpastatin and the actin cytoskeleton-associated proteins alphaII-spectrin, E-cadherin and vinculin were studied by immunoblotting. The cell damage was assessed by lactate dehydrogenase release and ultrastructural analysis including fluorescence-labelled actin filaments. Immediately after administration, CCK led to activation of both calpain isoforms, mu- and m-calpain. The protease activation was accompanied by a decrease in the E-cadherin level and formation of calpain-specific breakdown products of alphaII-spectrin. A calpain-specific cleavage product of vinculin appeared concomitantly with changes in the actin filament organization. No effect of CCK on calpastatin was found. Inhibition of calpain by ZVP reduced CCK-induced damage of the actin-associated proteins and the cellular ultrastructure including the actin cytoskeleton. The results suggest that CCK-induced acinar cell damage requires activation of calpain and that the actin cytoskeleton belongs to the cellular targets of the protease.

Topics: Actins; Acute Disease; Animals; Blotting, Western; Cadherins; Calcium-Binding Proteins; Calpain; Ceruletide; Cholecystokinin; Cysteine Proteinase Inhibitors; Cytoskeletal Proteins; Cytoskeleton; Dipeptides; Enzyme Activation; Female; Gene Expression; Microscopy, Confocal; Microscopy, Electron; Models, Animal; Organ Culture Techniques; Pancreas; Pancreatitis; Rats; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spectrin; Stimulation, Chemical; Time Factors; Vinculin

Development of human chronic pancreatitis is associated with intrapancreatic accumulation of pituitary adenylate cyclase-activating polypeptide (PACAP) accompanied with an altered inflammatory response (Michalski et al., Am J Physiol Gastrointest Liver Physiol. 2008;294:G50-G57). To investigate the role of pancreatic PACAP in the development of acute pancreatitis, we employed transgenic mice over-expressing PACAP in pancreatic beta-cells (PACAP-Tg). In comparison to wild-type mice, PACAP-Tg mice exhibited more severe pathophysiological signs of the cerulein-induced pancreatitis at 12 h, as evidenced by higher serum amylase and lipase levels accompanied by the exacerbation of pancreatic edema, necrosis, and inflammation. Cerulein treatment increased mRNA expression of several proinflammatory cytokines (TNFalpha, IL-1beta, and IL-6) at 12 h with similar magnitude both in wild-type and PACAP-Tg mice. In addition, the mRNA and protein levels of regenerating gene III beta (RegIIIbeta), a key factor in the pancreatic response to acute pancreatitis, were up-regulated at 24 h in wild-type mice upon cerulein administration, whereas they were attenuated in PACAP-Tg mice. These data indicate that over-expressed PACAP in pancreas enhances the cerulein-induced inflammatory response of both acinar cells, leading to aggravated acute pancreatitis, which was accompanied by a down-regulation of RegIIIbeta, an anti-inflammatory factor.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cytokines; Disease Models, Animal; Down-Regulation; Insulin-Secreting Cells; Lipase; Mice; Mice, Transgenic; Pancreatitis; Pancreatitis-Associated Proteins; Pituitary Adenylate Cyclase-Activating Polypeptide; Proteins; RNA, Messenger; Up-Regulation

Green tea polyphenols (GTPs) are naturally occurring antioxidants acting through pathways that include reactive oxygen species and nuclear factor kappa B (NF-kappaB). This study investigates the effect of GTPs in a cerulein-induced murine model of acute pancreatitis (AP).. Male CD mice (median weight, 37.7 g) were divided into 4 groups: mice administered with cerulein alone, cerulein and GTP, saline alone (sham), and GTP alone. Acute pancreatitis was induced by serial intraperitoneal administration of cerulein (50 microg/kg, x6). Green tea polyphenol was administered intraperitoneally at 25 mg/kg on the first, third, and sixth hours after pancreatitis induction.We analyzed histologic and biochemical features of AP, NF-kappaB pathway activity, leukocyte-mediated damage, cytokine levels, oxidative stress injury, lipid peroxidation, expression of poly-(adenosine diphosphate-ribose) synthetase, and presence of apoptosis.. Treatment with GTP reduced the histologic and biochemical features of AP. Western blot revealed significant NF-kappaB inactivation. Immunostaining for P selectin and intercellular adhesion molecule 1, tumor necrosis factor alpha, transforming growth factor beta, vascular endothelial growth factor, nitrotirosine, poly-(adenosine diphosphate ribose) synthetase, and malondialdheide levels were significantly reduced. There was a significant down-regulation of apoptotic markers.. Our results demonstrated that GTP significantly ameliorated the effects of cerulein-induced AP in mice. These effects of GTP are mediated by actions at the NF-kappaB/IkB (inhibitor kB) proteins and oxidative stress pathways.

Topics: Acute Disease; Animals; Apoptosis; Blotting, Western; Ceruletide; I-kappa B Proteins; Immunohistochemistry; In Situ Nick-End Labeling; Injections, Intraperitoneal; Intercellular Adhesion Molecule-1; Lipid Peroxidation; Male; Mice; Neutrophil Infiltration; NF-kappa B; P-Selectin; Pancreas; Pancreatitis; Phenol; Poly(ADP-ribose) Polymerases; Tea; Time Factors; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

Aquaporin 12 (AQP12) is the most recently identified member of the mammalian AQP family and is specifically expressed in pancreatic acinar cells. In vitro expression studies have revealed that AQP12 is localized at intracellular sites. To determine the physiological roles of AQP12 in the pancreas, we generated knockout mice for this gene (AQP12-KO). No obvious differences were observed under normal conditions between wild-type (WT) and AQP12-KO mice in terms of growth, blood chemistry, pancreatic fluid content, or histology. However, when we induced pancreatitis through the administration of a cholecystokinin-8 (CCK-8) analog, the AQP12-KO mice showed more severe pathological damage to this organ than WT mice. Furthermore, when we analyzed exocytosis in the pancreatic acini using a two-photon excitation imaging method, the results revealed larger exocytotic vesicles (vacuoles) in the acini of AQP12-KO mice at a high CCK-8 dose (100 nM). From these results, we conclude that AQP12 may function in the mechanisms that control the proper secretion of pancreatic fluid following rapid and intense stimulation.

Topics: Acute Disease; Amylases; Animals; Aquaporins; Ceruletide; Cholecystokinin; Diet; Disease Susceptibility; Endoplasmic Reticulum; Exocytosis; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreas, Exocrine; Pancreatitis; Peptide Fragments; Permeability; Photons; Tissue Distribution; Water

The aim of this study was to evaluate the role of oxidative damage in pancreatitis-induced hepatic injury. Thirty-five rats were divided into five groups (each of 7 rats): control, cerulein (100 microg/kg body weight), cerulein and pentoxifylline (12 mg/kg body weight), cerulein plus L-NAME (10 mg/kg body weight) and cerulein plus L-arginine (160 mg/kg body weight). The degree of hepatic cell degeneration differed significantly between groups. Mean malondialdehyde levels were 7.00 +/- 2.29, 20.89 +/- 10.13, 11.52 +/- 4.60, 18.69 +/- 8.56, and 8.58 +/- 3.68 nmol/mg protein for the control, cerulein, pentoxifylline, L-NAME, and L-arginine groups, respectively. Mean catalase activity was 3.20 +/- 0.83, 1.09 +/- 0.35, 2.05 +/- 0.91, 1.70 +/- 0.60, and 2.85 +/- 0.47 U/mg protein for the control, cerulein, pentoxifylline, L-NAME, and L-arginine groups, respectively, and mean glutathione peroxidase activity was 0.72 +/- 0.25, 0.33 +/- 0.09, 0.37 +/- 0.04, 0.34 +/- 0.07 and 0.42 +/- 0.1 U/mg protein for the control, cerulein, pentoxifylline, L-NAME, and L-arginine groups, respectively. Cerulein-induced liver damage was accompanied by a significant increase in tissue malondialdehyde levels (P < 0.05) and a significant decrease in catalase (P < 0.05) and GPx activities (P < 0.05). L-arginine and pentoxifylline, but not L-NAME, protected against this damage. Oxidative injury plays an important role not only in the pathogenesis of AP but also in pancreatitis-induced hepatic damage.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Female; Free Radical Scavengers; Lipid Peroxidation; Liver Diseases; NG-Nitroarginine Methyl Ester; Pancreatitis; Pentoxifylline; Rats; Rats, Wistar; Reactive Oxygen Species

Obestatin is a peptide derived from the proghrelin, a common prohormone for ghrelin and obestatin. Obestatin, like the ghrelin has been originally extracted from rat stomach, and the stomach seems to be a major source of circulating obestatin. Previous studies have shown that administration of ghrelin exhibits protective effect in the pancreas, inhibiting the development of acute pancreatitis. Recent study has shown that obestatin promotes survival of beta-cells and pancreatic islets. Aim of the present study was to investigate the influence of obestatin administration on the development of cerulein-induced pancreatitis. Studies were performed on male Wistar rats. Acute pancreatitis was induced by cerulein given intraperitoneally 5 times at a dose of 50 microg/kg/dose with 1-h intervals. Obestatin was injected twice intraperitoneally at the dose of 4, 8 or 16 nmol/kg/dose. In control saline-treated rats, obestatin was without effect on pancreatic morphology, serum activity of pancreatic enzymes, serum level of pro-inflammatory interleukin-1beta or pancreatic cells proliferation. In animals with induction of acute pancreatitis, morphological examination showed that administration of obestatin decreased pancreatic leukocyte infiltration and vacuolization of acinar cells. These effects were accompanied by reduction in the pancreatitis-evoked increase in serum level of pancreatic digestive enzymes, lipase amylase and poly-C ribonuclease. Obestatin administered at the highest dose of 16 nmo/kg/dose reduced serum activity of these enzymes by 33, 42 and 44%, respectively. Also serum concentration of pro-inflammatory interleukin-1beta was decreased by obestatin in rats with acute pancreatitis; whereas the pancreatitis-evoked decrease in pancreatic blood flow and pancreatic DNA synthesis was partially reversed. Administration of obestatin reduces the severity of cerulein-induced acute pancreatitis. This effect is related, at least in part, to the improvement of pancreatic blood flow and reduction in proinflammatory interleukin-1beta release.

Topics: Acute Disease; Amylases; Animals; Cell Proliferation; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Injections, Intraperitoneal; Interleukin-1beta; Lipase; Male; Pancreas; Pancreatitis; Peptide Hormones; Protective Agents; Rats; Rats, Wistar; Splanchnic Circulation

To evaluate the therapeutic role of caffeic acid phenethyl ester (CAPE) in a rat model of cerulean-induced acute pancreatitis (AP).. Seventy male Wistar albino rats were divided into seven groups. Acute edematous pancreatitis was induced by subcutaneous cerulein injection (20 microg/kg) four times at 1-h intervals. CAPE (30 mg/kg) was given by subcutaneous injection at the beginning (CAPE 1 group) and 12 h after the last cerulein injection (CAPE 2 group). Serum amylase, lipase, white blood cell count, and tumor necrosis factor (TNF)-alpha levels were measured, and pancreatic histopathology was assessed.. In the AP group, amylase and lipase levels were found to be elevated and the histopathological evaluation showed massive edema and inflammation of the pancreas, with less fatty necrosis when compared with sham and control groups. Amylase and lipase levels and edema formation decreased significantly in the CAPE therapy groups (P < 0001); especially in the CAPE 2 group, edema was improved nearly completely (P = 0001). Inflammation and fatty necrosis were partially recovered by CAPE treatment. The pathological results and amylase level in the placebo groups were similar to those in the AP group. White blood cell count and TNF-alpha concentration was nearly the same in the CAPE and placebo groups.. CAPE may be useful agent in treatment of AP but more experimental and clinical studies are needed to support our observation of beneficial effects of CAPE before clinical usage of this agent.

Topics: Acute Disease; Amylases; Animals; Caffeic Acids; Ceruletide; Cytotoxins; Disease Models, Animal; Edema; Leukocyte Count; Lipase; Male; Pancreas; Pancreatitis; Phenylethyl Alcohol; Rats; Rats, Wistar; Treatment Outcome; Tumor Necrosis Factor-alpha

Protease-activated receptor-2 (PAR2) is a 7-transmembrane G-protein-coupled tethered ligand receptor that is expressed by pancreatic acinar and ductal cells. It can be physiologically activated by trypsin. Previously reported studies (Namkung, W., Han, W., Luo, X., Muallem, S., Cho, K. H., Kim, K. H., and Lee, M. G. (2004) Gastroenterology 126, 1844-1859; Sharma, A., Tao, X., Gopal, A., Ligon, B., Andrade-Gordon, P., Steer, M. L., and Perides, G. (2005) Am. J. Physiol. 288, G388-G395) have shown that PAR2 activation exerts a protective effect on the experimental model of pancreatitis induced by supramaximal secretagogue (caerulein) stimulation. We now show that PAR2 exerts a worsening effect on a different model of experimental pancreatitis, i.e. one induced by retrograde pancreatic ductal infusion of bile salts. In vitro studies using freshly prepared pancreatic acini show that genetic deletion of PAR2 reduces bile salt-induced pathological calcium transients, acinar cell injury, and activation of c-Jun N-terminal kinase, whereas genetic deletion of PAR2 has the opposite or no effect on these pancreatitis-related events when they are elicited, in vitro, by caerulein stimulation. Studies employing a combination of trypsin inhibition and activation of PAR2 with the activating peptide SLIGRL show that all these differences indeed depend on the activation of PAR2. These studies are the first to report that a single perturbation can have model-specific and opposite effects on pancreatitis, and they underscore the importance of performing mechanistic pancreatitis studies using two dissimilar models of the disease to detect idiosyncratic, model-specific events. We suggest PAR2 activation exerts a worsening effect on the severity of clinical pancreatitis and that interventions interfering with PAR2 activation may be of benefit in the treatment of patients with severe pancreatitis.

Topics: Acute Disease; Animals; Bile Acids and Salts; Ceruletide; Enzyme Activation; Female; Male; Mice; Mice, Inbred C57BL; Models, Biological; Pancreas; Pancreatitis; Peptides; Protein Structure, Tertiary; Receptor, PAR-2

Obesity is a risk factor for acute pancreatitis (AP), but the molecular mechanism remains unclear. Adiponectin, an adipose tissue-derived secretory factor, has anti-inflammatory properties in addition to various biological functions, and its plasma concentrations are reduced in obese subjects. However, the role of adiponectin in AP has not been investigated.. To determine the effects of adiponectin on AP.. We investigated the effects of adiponectin on experimental AP by using adiponectin-knockout (APN-KO) mice and adenovirus-mediated adiponectin over-expression. AP was induced by 10 hourly intraperitoneal injections of low-dose caerulein (10 microg/kg) after 2 week feeding of normal chow or a high-fat diet (HFD) in wild-type (WT) and APN-KO mice. We evaluated the severity of AP biochemically and morphologically.. Low-dose caerulein treatment did not induce pancreatic damage in either WT or APN-KO mice under normal chow feeding. APN-KO mice, but not WT mice, fed a HFD and then treated with caerulein developed pancreatic damage and inflammation, accompanied by increased macrophage/neutrophil infiltration and upregulation of pro-inflammatory mediators such as tumour necrosis factor alpha in the pancreas. Adenovirus-mediated over-expression of adiponectin attenuated the severity of HFD/caerulein-induced AP in APN-KO mice.. Adiponectin plays a protective role in caerulein-induced AP in HFD-fed mice.

Topics: Acute Disease; Adiponectin; Adipose Tissue; Animals; Ceruletide; Dietary Fats; Humans; Mice; Mice, Inbred C57BL; Mice, Knockout; Obesity; Pancreatitis; Reverse Transcriptase Polymerase Chain Reaction; Up-Regulation

Obesity is clearly an independent risk factor for increased severity of acute pancreatitis (AP), although the mechanisms underlying this association are unknown. Adipokines (including leptin and adiponectin) are pleiotropic molecules produced by adipocytes that are important regulators of the inflammatory response. We hypothesized that the altered adipokine milieu observed in obesity contributes to the increased severity of pancreatitis. Lean (C57BL/6J), obese leptin-deficient (LepOb), and obese hyperleptinemic (LepDb) mice were subjected to AP by six hourly intraperitoneal injections of cerulein (50 microg/kg). Severity of AP was assessed by histology and by measuring pancreatic concentration of the proinflammatory cytokines IL-1beta and IL-6, the chemokine MCP-1, and the marker of neutrophil activation MPO. Both congenitally obese strains of mice developed significantly more severe AP than wild-type lean animals. Severity of AP was not solely related to adipose tissue volume: LepOb mice were heaviest; however, LepDb mice developed the most severe AP both histologically and biochemically. Circulating adiponectin concentrations inversely mirrored the severity of pancreatitis. These data demonstrate that congenitally obese mice develop more severe AP than lean animals when challenged by cerulein hyperstimulation and suggest that alteration of the adipokine milieu exacerbates the severity of AP in obesity.

Topics: Acute Disease; Adipokines; Adiponectin; Amylases; Animals; Blood Glucose; Body Weight; Ceruletide; Chemokines; Cytokines; Disease Models, Animal; Female; Insulin; Leptin; Lung; Mice; Mice, Inbred C57BL; Mice, Obese; Obesity; Pancreas; Pancreatitis; Peroxidase; Severity of Illness Index

Some recent studies indicate that cannabis may induce acute pancreatitis in humans and administration of anandamide increases the severity of acute pancreatitis; whereas another study exhibits some therapeutic effects in acute pancreatitis. Aim of the present study was to discover what is the reason for these opposite confusing results and to determine the role of sensory nerves in this effect. Acute pancreatitis was induced in rats by cerulein. Anandamide, an endogenous cannabinoid, was administered i.p. (1.5 micromol/kg) before or 2 h after cerulein administration. Stimulation of sensory nerves was performed by capsaicin (0.5 mg/kg s.c.). In rats treated with combination of anandamide plus capsaicin, capsaicin was given 10 min after each dose of anandamide. After the last injection of cerulein or 4 h later, the study was terminated. In our study we observed that stimulation of sensory nerves by capsaicin, before administration of cerulein, reduced the severity of acute pancreatitis. Anandamide, administered alone before cerulein, increased pancreatic damage in acute pancreatitis. Anandamide administered in combination with capsaicin, before cerulein, abolished the capsaicin-induced protective effect on the pancreas. Opposite effects were observed when capsaicin and anandamide were administered after injection of cerulein. Capsaicin increased the severity of acute pancreatitis, whereas anandamide reduced pancreatic damage and reversed the deleterious effect of capsaicin. We conclude that the effect of anandamide on the severity of acute pancreatitis depends on the phase of this disease. Administration of anandamide, before induction of pancreatitis, aggravates pancreatic damage; whereas anandamide administered after induction of pancreatitis, reduces the severity of acute pancreatitis. Sensory nerves are involved in the mechanism of this biphasic effect of anandamide.

Topics: Acute Disease; Animals; Arachidonic Acids; Cannabinoid Receptor Modulators; Ceruletide; Disease Models, Animal; Drug Administration Schedule; Endocannabinoids; Male; Neurons, Afferent; Pancreatitis; Polyunsaturated Alkamides; Rats; Rats, Wistar; Severity of Illness Index; Time Factors

Some authors have found beneficial effect of statins in certain inflammatory conditions, but the effect of statins on acute pancreatitis is not yet defined.. The aim of this study was to evaluate the effect of simvastatin on an experimental model of mild and severe acute pancreatitis.. One hundred and one Wistar rats with cerulein or taurocholate-induced acute pancreatitis were used in this study.. The rats were divided into two groups: Group I (n=51) received two previously i.p. injections (18+/-2 and 3+/-1 hours) of simvastatin (200 microg/kg) and Group II (n=50) received two previously i.p. injections of saline. Both groups were subdivided into two subgroups: mild pancreatitis (cerulein-induced; IA, n=10; IIA, n=10) and severe pancreatitis (taurocholate-induced; IB, n=41; IIB, n=40).. The parameters evaluated were: pancreatic vascular permeability, tissue water content, histologic lesion, amylase serum levels in rats with mild pancreatitis (subgroups A); mortality rate, serum levels of IL-6, IL-10, amylase, pulmonary myeloperoxidase activity and ascitic levels of TNF-alpha in rats with severe pancreatitis (subgroups B).. Serum levels of IL-10 were significantly lower in the simvastatin-treated group as well as the myeloperoxidase activity. There was no significant difference in any of other studied parameters.. Simvastatin appears to reduce inflammatory cytokines and pulmonary neutrophilic activation in the severe acute pancreatitis model, but there is no significant effect on survival curve, in spite of a clear trend towards a better survival in the simvastatin group.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Interleukin-10; Interleukin-6; Lung; Male; Pancreatitis; Peroxidase; Rats; Simvastatin; Survival Rate; Taurocholic Acid; Tumor Necrosis Factor-alpha

Topics: Acute Disease; Animals; Antioxidants; Ceruletide; Male; Oxidative Stress; Pancreatitis; Pyruvic Acid; Rats; Rats, Wistar

Acute pancreatitis (AP) is an inflammatory disease of the pancreas, which evolves in approximately 20% of the patients to a severe illness associated with a high mortality rate. In this study, we performed a comparative proteomic analysis of pancreatic tissue extracts from rats with AP and healthy rodent controls in order to identify changes in protein expression related to the pathobiological processes of this disease. Pancreatic extracts from diseased and controls rats were analyzed by 2-DE and MS/MS. A total of 125 proteins were identified from both samples. Comparative analysis allowed the detection of 42 proteins or protein fragments differentially expressed between diseased and control pancreas, some of them being newly described in AP. Interestingly, these changes were representative of the main pathobiological pathways involved in this disease. We observed activation of digestive proteases and increased expression of various inflammatory markers, including several members of the alpha-macroglobulin family. We also detected changes related to oxidative and cell stress responses. Finally, we highlighted modifications of 14-3-3 proteins that could be related to apoptosis regulation. These results showed the interest of proteomic analysis to identify changes characterizing pancreatic tissue damage and, therefore, to highlight new potential biomarkers of AP.

Topics: 14-3-3 Proteins; Acute Disease; Animals; Antigens, Neoplasm; Biomarkers; Biomarkers, Tumor; Ceruletide; Disease Models, Animal; Electrophoresis, Gel, Two-Dimensional; Lectins, C-Type; Lithostathine; Male; Oxidative Stress; Pancreas; Pancreatitis; Pancreatitis-Associated Proteins; Proteomics; Rats; Rats, Sprague-Dawley; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tandem Mass Spectrometry

Autodigestion of the pancreas by its own prematurely activated digestive proteases is thought to be an important event in the onset of acute pancreatitis. Although lysosomal hydrolases, such as cathepsin B, play a key role in intrapancreatic trypsinogen activation, it remains unclear where and how trypsinogen meets these lysosomal enzymes. Autophagy is an intracellular bulk degradation system in which cytoplasmic components are directed to the lysosome/vacuole by a membrane-mediated process. To analyze the role of autophagy in acute pancreatitis, we produced a conditional knockout mouse that lacks the autophagy-related (Atg) gene Atg5 in the pancreatic acinar cells. The severity of acute pancreatitis induced by cerulein is greatly reduced in these mice. In addition, Atg5-deficient acinar cells show a significantly decreased level of trypsinogen activation. These data suggest that autophagy exerts a detrimental effect in pancreatic acinar cells by activation of trypsinogen to trypsin. We propose a theory in which autophagy accelerates trypsinogen activation by lysosomal hydrolases under acidic conditions, thus triggering acute pancreatitis in its early stage.

Topics: Acute Disease; Animals; Autophagy; Autophagy-Related Protein 5; Ceruletide; Cytoplasm; Enzyme Activation; Mice; Mice, Knockout; Microtubule-Associated Proteins; Pancreatitis; Trypsinogen; Vacuoles

To investigate the mechanism of Chaiqin Chengqi Decoction (CQCQD), a compound of traditional Chinese herbal medicine, acting on the pancreatic acinar cell calcium overload in rats with acute pancreatitis (AP).. A total of 30 SD rats were randomly divided into normal control group, untreated group and CQCQD group (n=10, respectively). AP was induced in rats by caerulein (5x50 mug/kg) intraperitoneal injection within 4 h. The pancreatic tissue SERCA1 and SERCA2 mRNA expressions were detected by fluorescent quantization polymerase chain reaction method; intracellular calcium fluorescence intensity (FI) of pancreatic acinar cells and the pancreatic pathological score were measured by laser scanning confocal microscopy and light microscopy respectively.. There were no SERCA1 mRNA expressions in pancreatic acinar cells of rats in the normal control group and the untreated group. The expression of pancreatic SERCA2 mRNA in the untreated group was down-regulated compared with that in the normal control group (expression ratio=0.536; P=0.001); the expression of pancreatic SERCA2 mRNA in the CQCQD group was up-regulated compared with that in the untreated group (expression ratio=2.00; P=0.012). The pancreatic pathological score in the CQCQD group was lower than that in the untreated group and the FI of Ca(2+) was also lower.. CQCQD can up-regulate the expression of pancreatic SERCA2 mRNA, release the calcium overload, and hence reduce the pathological changes in pancreatic tissue.

Topics: Acute Disease; Animals; Calcium; Calcium Channel Blockers; Ceruletide; Drugs, Chinese Herbal; Male; Pancreas, Exocrine; Pancreatitis; Random Allocation; Rats; Rats, Sprague-Dawley; RNA, Messenger; Sarcoplasmic Reticulum Calcium-Transporting ATPases

Bacterial endotoxin (lipopolysaccharide, LPS), is the component of the cellular wall of Gram negative bacteria. Endotoxemia (sepsis) could produce multiorgan failure and could be particularly danger in the early period of life. The effects of endotoxemia induced in the neonatal period of life on the pancreatic secretory function and on pancreatic defense of adult organism have not been investigated yet. To induce endotoxemia suckling rats (30 g) have been injected intraperitoneally with LPS from E. coli (5, 10 or 15 mg/kg-day) during 5 consecutive days. Three months later in these animals (300 g) the studies on pancreatic secretion and acute pancreatitis were carried out. In the adult rats, which have been subjected in infancy to endotoxemia, basal pancreatic secretion was unaffected, whereas amylase secretions stimulated by caerulein or by diversion of pancreatic-biliary juice to the exterior were significantly, and dose-dependently reduced as compared to the untreated control. In the rats pretreated with LPS in the suckling period of life caerulein-induced amylase release from isolated pancreatic acini was significantly decreased, and dose-dependent reduction of mRNA signal for CCK1 receptor on pancreatic acini have been observed. Caerulein infusion (25 microg/kg) produced caerulein induced pancreatitis (AP) in all animals tested, that was confirmed by histological examination. In the rats, which have been subjected in the neonatal period of life to LPS (10 or 15 mg/kg-day x 5 days) all manifestations of AP have been reduced. In these animals acute inflammatory changes of pancreatic tissue have been significantly diminished. Pancreatic weight and plasma lipase activity, have been markedly decreased in these animals as compared to the control rats, subjected in the infancy to saline injection instead of LPS. Caerulein-induced fall in an antioxidative enzyme; SOD concentration was reversed and accompanied by significant reduction of lipid peroxidation products; MDA+ 4 HNE in the pancreatic tissue.. 1/ neonatal endotoxemia reduces gene expression for CCK1 receptor and could produce impairment of the exocrine pancreatic function at adult age; 2/ Prolonged exposition of suckling rats to bacterial endotoxin attenuated acute pancreatitis induced in these animals at adult age and this effect could be related to the increased concentration of antioxidative enzyme SOD in the pancreatic tissue.

Topics: Actins; Acute Disease; Amylases; Animals; Animals, Newborn; Animals, Suckling; Ceruletide; Cytokines; Dose-Response Relationship, Drug; Endotoxemia; Heat-Shock Proteins; Lipase; Lipid Peroxidation; Lipopolysaccharides; Pancreas; Pancreatitis; Rats; Receptor, Cholecystokinin A; Superoxide Dismutase

In patients suffering from acute pancreatitis, the pathogenesis is not completely understood, and several recent studies in vitro suggested that heat shock proteins might play an important role in cell signaling. To investigate the possible role of extracellular heat shock protein 70 (Hsp70) in pancreatitis, toll-like receptor-4 (TLR4)-deficient and wild-type mice were administered with exogenous Hsp70 during the course of cerulein-induced pancreatitis (CIP).. Acute pancreatitis was induced by 5 intraperitoneal injections of cerulein at hourly intervals, and then treated with recombinant Hsp70 through the caudal vein 4 hours after the start of cerulein injections. Subsequently serum amylase and serum cytokines levels were detected. Histologic alteration of the pancreas was evaluated. Tumor necrosis factor alpha (TNF-alpha) concentrations and myeloperoxidase (MPO) activity in both pancreas and lungs were analyzed. The nuclear factor kappa B (NF-kappaB) activation in pancreatic tissue was measured using a sensitive RelA enzyme-linked immunosorbent assay.. Treatment with recombinant Hsp70 to wild-type mice in CIP resulted in significant aggravation of inflammation in pancreas, elevated levels of serum cytokines, up-regulation of pulmonary MPO activity and increase of lung tissues TNF-alpha concentrations. In contrast, treatment with Hsp70 to TLR4-deficient mice had little effect on serum cytokines levels, pancreatic inflammation, pulmonary MPO activity and TNF-alpha concentrations.. The results suggest that extracellular Hsp70 might induce systemic inflammatory response syndrome (SIRS)-like response in vivo and TLR4 might be involved in the Hsp70-mediated activation of inflammatory reaction in the progression of CIP without infection.

Topics: Acute Disease; Animals; Ceruletide; Female; HSP70 Heat-Shock Proteins; Male; Mice; Mice, Inbred C57BL; Pancreatitis; Systemic Inflammatory Response Syndrome; Toll-Like Receptor 4

To investigate the effect of Gardenia jasminoides (GJ) on cerulein-induced acute pancreatitis (AP) in mice.. C57BL/6 mice weighing 18-20 g were divided into three groups. (1) Normal saline-treated group, (2) treatment with GJ at a dose of 0.1 g/kg, (3) treatment with GJ at a dose of 1 g/kg. GJ was administered orally (n = 6 per group) for 1 wk. Three hours later, the mice were given an intraperitoneal injection of cerulein (50 microg/kg), a stable cholecystokinin (CCK) analogue, every hour for a total of 6 h as described previously. The mice were sacrificed at 6 h after completion of cerulein injections. Blood samples were obtained to determine serum amylase, lipase and cytokine levels. The pancreas was rapidly removed for morphologic examination and scoring. A portion of pancreas was stored at -70 degree and prepared for the measurement of tissue myeloperoxidase (MPO) activity, an indicator of neutrophil sequestration, and for reverse-transcriptase PCR (RT-PCR) and real-time PCR measurements.. Treatment with GJ decreased significantly the severity of pancreatitis and pancreatitis-associated lung injury. Treatment with GJ attenuated the severity of AP compared with saline-treated mice, as shown by reduction in pancreatic edema, neutrophil infiltration, serum amylase and lipase levels, serum cytokine levels, and mRNA expression of multiple inflammatory mediators.. These results suggest that GJ attenuated the severity of AP as well as pancreatitis-associated lung injury.

Topics: Acute Disease; Administration, Oral; Amylases; Animals; Anti-Inflammatory Agents; Body Weight; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Gardenia; Inflammation Mediators; Interleukin-1beta; Interleukin-6; Lipase; Lung; Lung Injury; Mice; Mice, Inbred C57BL; Neutrophil Infiltration; Organ Size; Pancreas; Pancreatitis; Peroxidase; Plant Extracts; RNA, Messenger; Tumor Necrosis Factor-alpha

The prognosis of acute pancreatitis (AP) depends upon the degree of pancreatic necrosis and the intensity of multisystem organ failure. The liver contributes to the systemic manifestations of AP by releasing some cytokines. This study was undertaken to examine comparative effects of melatonin, antioxidant mixture containing L(+)-ascorbic acid and N-acetyl cysteine, pentoxifylline and L-arginine on hepatic damage induced by caerulein-pancreatitis.. The liver specimens of all groups showed histopathological alterations such as hepatocyte necrosis, intracellular vacuolization, vascular congestion, sinusoidal dilatation and inflammatory infiltration. TEM studies revealed vacuole formation, mitochondrial degeneration, lysosome accumulation and necrosis. The mean histopathological score of the caerulein group was significantly different from that of each treatment group.. L-Arginine and antioxidant administration be important for reducing hepatic damage induced by AP. Improvement of hepatic damage, in turn, might be beneficial for the prognosis of AP.

Topics: Acetylcysteine; Acute Disease; Animals; Antioxidants; Arginine; Ascorbic Acid; Ceruletide; Female; Liver Diseases; Melatonin; Pancreatitis; Pentoxifylline; Rats; Rats, Wistar; Statistics, Nonparametric

The proteins expressed in pancreatic acinar cells during the initiation of acute pancreatitis may determine the severity of the disease. Cerulein pancreatitis is one of the best characterized models for acute pancreatitis. Present study aims to determine the differentially expressed proteins in cerulein-stimulated pancreatic acinar cells as an in vitro model for acute pancreatitis. Rat pancreatic acinar AR42J cells were treated with 10(-8)M cerulein for 12h. The protein patterns separated by two-dimensional electrophoresis using pH gradients of 5-8 were compared between the cells treated without cerulein and those with cerulein. The changed proteins were conclusively identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis of the peptide digests. As a result, 10 proteins (Orp150 protein, protein disulfide isomerase related protein, dnaK-type molecular chaperone hsp72-ps1, mitochondrial glutamate dehydrogenase, similar to chaperonin containing TCP-1 beta subunit, RuvB-like protein 1, heterogeneous nuclear ribonucleoprotein H1, aldehyde reductase 1, triosephosphate isomerase 1, peroxiredoxin 2) were up-regulated while four proteins (vasolin-containing protein, 78 kDa glucose-regulated protein precursor, heat shock protein 8, adenosylhomocysteinase) were down-regulated by cerulein in pancreatic acinar AR42J cells. These proteins are related to chaperone, cell defense mechanism against oxidative stress or DNA damage, anti-apoptosis and energy generation. The differentially expressed proteins by ceruein share their functional roles in pancreatic acinar cells, suggesting the possible involvement of oxidative stress, DNA damage, and anti-apoptosis in pathogenesis of acute pancreatitis. Proteins involved in cellular defense mechanism and energy production may protect pancreatic acinar cells during the development of pancreatitis.

Topics: Acute Disease; Animals; Cell Line, Tumor; Ceruletide; Disease Models, Animal; Down-Regulation; Electrophoresis, Gel, Two-Dimensional; Pancreatitis; Protein Array Analysis; Proteins; Rats; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Up-Regulation

The mechanisms for tissue regeneration and renewal after acute pancreatitis are not well understood but may involve activation of Notch signaling. To study the effect of Notch signaling ablation during acute experimental pancreatitis, we used a chemical and genetic approach to ablate Notch signaling in cerulein-induced pancreatitis in mice.. Acute pancreatitis was induced by cerulein treatment in mice treated with the gamma-secretase inhibitor dibenzazepine or in conditional Notch1 knockout mice. Mice were characterized using immunohistologic, biochemical, and molecular methods. To investigate Notch and beta-catenin interaction, acinar 266-6 cells were analyzed using transfection and biochemical assays.. Loss of Notch signaling results in impaired regeneration after acute pancreatitis with fewer mature acinar cells in dibenzazepine-treated and Notch1-deficient mice in the regenerative phase 3 days after induction. beta-catenin expression was increased and prolonged during exocrine regeneration. Crosstalk between Notch and beta-catenin-mediated signaling was identified, with Notch1-IC inhibiting beta-catenin-mediated transcriptional activity. This inhibition was dependent on a functional RAM domain.. Inhibition of Notch signaling in vivo leads to impaired regeneration of the exocrine pancreas after acute pancreatitis. Our results suggest an interaction of Notch and Wnt signaling in pancreatic acinar cells, providing evidence for a role of these pathways in the regulation of the maturation process of acinar cells.

Topics: Acute Disease; Amyloid Precursor Protein Secretases; Animals; beta Catenin; Cell Line, Tumor; Ceruletide; Dibenzazepines; Disease Models, Animal; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas, Exocrine; Pancreatic Neoplasms; Pancreatitis; Receptor, Notch1; Regeneration; Signal Transduction; Wnt Proteins

Acute pancreatitis (AP) is associated with significant morbidity and mortality; however, there is no specific treatment for this disease. A novel salivary tripeptide analog, feG, reduces inflammation in several different animal models of inflammation. The aims of this study were to determine whether feG reduced the severity of AP and modifies the expression of pancreatic ICAM-1 mRNA during AP in a mouse model. AP was induced in mice by hourly (x12) intraperitoneal injections of caerulein. A single dose of feG (100 microg/kg) was coadministered with caerulein either at time 0 h (prophylactic) or 3 h after AP induction (therapeutic). Plasma amylase and pancreatic MPO activities and pancreatic ICAM-1 mRNA expression (by RT-PCR) were measured. Pancreatic sections were histologically assessed for abnormal acinar cells and interstitial space. AP induction produced a sevenfold increase in plasma amylase, a tenfold increase in pancreatic MPO activity, and a threefold increase in interstitial space, and 90% of the acinar cells were abnormal. Prophylactic treatment with feG reduced the AP-induced plasma amylase activity by 45%, pancreatic MPO by 80%, the proportion of abnormal acinar cells by 30%, and interstitial space by 40%. Therapeutic treatment with feG significantly reduced the AP-induced abnormal acinar cells by 10% and the interstitial space by 20%. Pancreatic ICAM-1 mRNA expression was upregulated in AP and was reduced by 50% with prophylactic and therapeutic treatment with feG. We conclude that feG ameliorates experimental AP acting at least in part by modulating ICAM-1 expression in the pancreas.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents; Ceruletide; Disease Models, Animal; Injections, Intraperitoneal; Intercellular Adhesion Molecule-1; Male; Mice; Oligopeptides; Pancreas; Pancreatitis; Peroxidase; RNA, Messenger; Severity of Illness Index; Time Factors

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Pancreas; Pancreatitis; Rats; RNA, Messenger; Time Factors; Toll-Like Receptor 9; Up-Regulation

This study was planned to observe the effects of nitric oxide synthesis on the antioxidative defense enzymes and pancreatic tissue histology in caerulein-induced acute pancreatitis. Acute pancreatitis was induced by intraperitoneal injections of 50 microg/kg caerulein, L-arginine used for NO induction and N(omega)-nitro-L-arginine methyl ester (L-NAME) used for NO inhibition. In the caerulein group acinar cell degeneration, interstitial inflammation, oedema and haemorrhage were detected. Pancreatic damage scores were decreased with both NO induction and inhibition (p<0.05). MDA, GSH-Px, CAT, GSH and SOD activities were significantly changed in the caerulein group and indicated increased oxidative stress. Both NO induction and inhibition decreased this oxidative stress. It is concluded that both nitric oxide induction and inhibition ameliorated caerulein-induced acute pancreatitis. The findings indicate that a certain amount of NO production has beneficial effects in experimental acute pancreatitis, but uncontrolled over-production of NO may be detrimental.

Topics: Acute Disease; Amylases; Animals; Arginine; Ceruletide; Female; Gene Expression Regulation; Lipase; Microscopy, Electron, Transmission; NG-Nitroarginine Methyl Ester; Nitric Oxide; Oxidative Stress; Pancreatitis; Rats; Rats, Sprague-Dawley

To investigate the effect of Chaiqinchengqi decoction (CQCQD) on sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) mRNA expression of pancreatic tissues in acute pancreatitis (AP) rats.. Thirty Sprague-Dawley (SD) rats were randomized into control group, AP group and CQCQD group (n = 3 x 10). The rats in the CQCQD group were intragastrically administered with CQCQD (10 mL/kg every 2 h) after induction of AP by intraperitoneal injection of caerulein (50 microg/kg.h x 5) within 4 h. At 6 h after the induction of AP model, pancreatic tissues were collected for the pathological observation, mRNA extraction for determination of SERCA1 and SERCA2 mRNA expression or pancreatic acinar cell isolation for measurement of fluorescence intensity (FI) of intracellular calcium ion concentration [Ca2+]i.. There was no expression of pancreatic SERCA1 mRNA in the control group and the AP group. The expression of pancreatic SERCA2 mRNA in the AP group was down-regulated (expression ratio = 0.536; P = 0.001) compared with the control group, while that in the CQCQD group was up-regulated (expression ratio = 2.00; P = 0.012) compared with AP group. The FI of intracellular [Ca2+] of pancreatic acinar cells in the AP group (138.2 +/- 23.1) was higher than the C group (111.0 +/- 18.4) and the CQCQD group (118.7 +/- 15.2 ) (P < 0.05) and the pancreatic pathological score in the CQCQD group was lower than that in the AP group (5.7 +/- 1.9 vs 9.2 +/- 2.7, P < 0.05).. CQCQD can up-regulate the expression of SERCA2 mRNA of pancreatic tissues, reduce intracellular calcium overload and relieve pancreatic tissue lesions.

Topics: Acute Disease; Animals; Calcium; Ceruletide; Disease Models, Animal; Drugs, Chinese Herbal; Gene Expression Regulation, Enzymologic; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; RNA, Messenger; Sarcoplasmic Reticulum Calcium-Transporting ATPases

Hydrogen sulphide (H(2)S), a novel gasotransmitter, has been recognized to play an important role in inflammation. Cystathionine-gamma-lyase (CSE) is a major H(2)S synthesizing enzyme in the cardiovascular system and DL-propargylglycine (PAG) is an irreversible inhibitor of CSE. Substance P (SP), a product of preprotachykinin-A (PPT-A) gene, is a well-known pro-inflammatory mediator which acts principally through the neurokinin-1 receptor (NK-1R). We have shown an association between H(2)S and SP in pulmonary inflammation as well as a pro-inflammatory role of H(2)S and SP in acute pancreatitis. The present study was aimed to investigate the interplay between pro-inflammatory effects of H(2)S and SP in a murine model of caerulein-induced acute pancreatitis. Acute pancreatitis was induced in mice by 10 hourly intraperitoneal injections of caerulein (50 (g/kg). PAG (100 mg/kg, i.p.) was administered either 1 hr before (prophylactic) or 1 hr after (therapeutic) the first caerulein injection. PAG, given prophylactically as well as therapeutically, significantly reduced plasma H(2)S levels and pancreatic H(2)S synthesizing activities as well as SP concentrations in plasma, pancreas and lung compared with caerulein-induced acute pancreatitis. Furthermore, prophylactic as well as therapeutic administration of PAG significantly reduced PPT-A mRNA expression and NK-1R mRNA expression in both pancreas and lung when compared with caerulein-induced acute pancreatitis. These results suggest that the pro-inflammatory effects of H(2)S may be mediated by SP-NK-1R pathway in acute pancreatitis.

Topics: Acute Disease; Alkynes; Animals; Ceruletide; Cystathionine beta-Synthase; Glycine; Hydrogen Sulfide; Inflammation; Male; Mice; Pancreatitis; Random Allocation; Receptors, Neurokinin-1; RNA, Messenger; Substance P

The present study investigated whether chemokines are involved in hydrogen sulfide (H2S)-associated pathogenesis of acute pancreatitis and associated lung injury.. We have examined the effect of DL-propargylglycine, a cystathionine gamma-lyase inhibitor, on the synthesis of CC chemokine monocyte chemotactic protein 1, Regulated upon Activation, Normal T-cell Expressed, and Secreted, and macrophage inflammatory protein-1alpha (MIP-1alpha), and CXC chemokine MIP-2 in an in vitro and in vivo model of cerulein-induced acute pancreatitis and associated lung injury. In addition, the pancreatic acinar cells were treated with H2S donor drug, sodium hydrosulfide. The expression of these chemokines in the pancreatic acini, pancreas, and lungs was determined by quantitative real-time reverse transcriptase polymerase chain reaction, enzyme-linked immunosorbent assay, and immunohistochemistry.. After treatment with DL-propargylglycine, reverse transcriptase polymerase chain reaction, and enzyme-linked immunosorbent assay demonstrated down-regulation of cerulein-induced increase in monocyte chemotactic protein 1, MIP-1alpha, and MIP-2 expression but had no apparent effect on Regulated upon Activation, Normal T-cell Expressed, and Secreted expression.. These results suggest that the proinflammatory effect of H2S may be mediated by chemokines.

Topics: Acute Disease; Alkynes; Animals; Ceruletide; Chemokine CCL5; Chemokines; Cystathionine gamma-Lyase; DNA Primers; Glycine; Hydrogen Sulfide; Male; Mice; Pancreatitis; Respiratory Distress Syndrome; Reverse Transcriptase Polymerase Chain Reaction

Transient receptor potential subtype vanilloid 1 (TRPV1) is an ion channel that is primarily expressed by primary sensory neurons where it mediates pain and heat sensation and participates in neurogenic inflammation. In this study, we examined the role of TRPV1 during neurogenic activation of pancreatic inflammation using a secretagogue-induced model in mice.. A supramaximal dose of caerulein (50 microg/kg) was injected hourly for 12 hours. Mice lacking TRPV1 were compared to wild-type animals.. All the parameters: serum amylase, pancreatic myeloperoxidase activity, histological scoring, pancreatic wet weight/body weight ratio, and quantification of neurokinin-1 receptor internalization indicated that null mice were not protected from acute pancreatitis. However, when primary sensory neurons were ablated by injection of the neurotoxin and TRPV1 agonist, resiniferatoxin, pancreatitis was ameliorated in wild-type mice but not in null mice, indicating that nerves bearing TRPV1 are part of the inflammatory pathway in acute pancreatitis because disappearance significantly reduced the inflammatory response.. Nerves expressing TRPV1 participate in the neurogenic inflammation during acute pancreatitis. The lack of protection in TRPV1 null mice suggests that an alternate pathway to TRPV1 coexists in the same neurons.

Topics: Acute Disease; Animals; Ceruletide; Crosses, Genetic; Disease Models, Animal; Endocytosis; Female; Gene Deletion; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Neurons, Afferent; Pancreatitis; TRPV Cation Channels

Smad6 is implicated in the inhibition of bone morphogenetic protein signalling. However, the function of Smad6 in the pancreas remains obscure.. To elucidate the unknown function of Smad6, we developed transgenic mice selectively expressing Smad6 in pancreatic acinar cells using a plasmid construct coding rat elastase 1 enhancer/promoter.. Smad6 transgenic mice had no specific distinguishing phenotype such as body weight, pancreatic wet weight and concentrations of pancreatic protein. However, Smad6 transgenic mice reacted to hyperstimulation by caerulein injection or a diet containing 0.5% ethionine. Maximal amylase release stimulated by CCK-8 was significantly decreased in Smad6 transgenic mice acini, and trypsin activities in transgenic mice acini were significantly increased after stimulation of CCK-8. There was no difference in effect of CCK-8 stimulation on the subsequent increase in intracellular free Ca2+ concentration ([Ca2+](i)) between wild-type and transgenic mice acini. These findings suggest that reduced pancreatic enzyme secretion was caused by the disorder of its downstream signal transduction pathways in acinar cells. The amino acid sequence at the N-terminus of Smad6 was similar to that of synaptosome-associated protein (SNAP) 25 interacting protein, which plays an important role in regulating exocytosis of pancreatic enzymes in acinar cells. Pancreatic SNAP25 protein levels in transgenic mice were decreased after caerulein-induced pancreatitis.. These results suggest that elevated expression of Smad6 inhibits normal function of SNAP25-interacting protein and SNAP25, reduces amylase secretion in acinar cells, and increases the susceptibility of acinar cells to the onset of pancreatitis.

Topics: Acute Disease; Amino Acid Sequence; Amylases; Animals; Ceruletide; Disease Models, Animal; Disease Progression; Genetic Predisposition to Disease; Mice; Mice, Inbred C57BL; Mice, Transgenic; Molecular Sequence Data; Pancreas, Exocrine; Pancreatitis; Phenotype; RNA, Messenger; Sequence Alignment; Smad6 Protein; Synaptosomal-Associated Protein 25; Transforming Growth Factor beta1; Up-Regulation

Accumulating evidence suggests the neuropeptide substance P (SP) and its receptor neurokinin-1 receptor (NK-1R) play a pivotal role in the pathogenesis of acute pancreatitis (AP). However, the mechanisms remain unclear. The present study investigated whether chemokines as proinflammatory molecules are involved in SP-NK-1R-related pathogenesis of this condition. We observed temporally and spatially selective chemokine responses in secretagogue caerulein-induced AP in mice. CC chemokines monocyte chemotactic protein (MCP)-1 and macrophage inflammatory protein-1alpha (MIP-1alpha) and CXC chemokine MIP-2 were elevated after AP induction. Time-dependent, tissue-specific analysis of their mRNA and protein expression suggested that they are early mediators in the condition and mediate local as well as systemic inflammatory responses. In contrast, another CC chemokine regulated on activation, T cells expressed and secreted (RANTES) was only involved in local pancreatic inflammation at a later stage of the disease. Either prophylactic or therapeutic treatment with a potent selective NK-1R antagonist CP-96,345 significantly suppressed caerulein-induced increase in MCP-1, MIP-1alpha, and MIP-2 expression but had no apparent effect on RANTES expression. The suppression effect of CP-96,345 on MCP-1, MIP-1alpha, and MIP-2 expression was concordantly demonstrated by immunohistochemistry, which, additionally, suggested that chemokine immunoreactivity was localized to acinar cells and the infiltrating leukocytes in the pancreas and alveolar macrophages, epithelial cells, and endothelial cells in the lungs. Our data suggest that SP, probably by acting via NK-1R on various chemokine-secreting cells in the pancreas and lungs, stimulates the release of chemokines that aggravate local AP and the development of its systemic sequelae.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Biphenyl Compounds; Ceruletide; Chemokine CCL2; Chemokine CCL3; Chemokine CCL4; Chemokine CXCL2; Chemokines; Disease Models, Animal; Gene Expression Regulation; Lung; Lung Injury; Macrophage Inflammatory Proteins; Macrophages, Alveolar; Male; Mice; Neurokinin-1 Receptor Antagonists; Pancreatitis; Reverse Transcriptase Polymerase Chain Reaction; RNA; Transcription, Genetic

The STAT pathways are integral to the inflammatory response and these proteins provide a direct link between the cytokine receptors and cytokine-induced gene transcription. We examined the roles of STAT4 and STAT6 in lung injury after caerulein-induced severe acute pancreatitis. We hypothesized that a modified organ expression of cytokines and chemokines that occurs in transgenic mice may affect the systemic response to severe acute pancreatitis.. Acute pancreatitis [13-hourly intraperitoneal injections of caerulein (50 microg/kg body weight, 0.2 mL) or the same volume of saline] was induced in wild-type (BALB/c) and transgenic (STAT4 or STAT6) mice of the same background, 7 to 8 weeks old. The pancreatic and lung tissues were collected at 1, 6, 12, and 24 h after the completion of caerulein administration. Tissue leukocyte sequestration was assessed by myeloperoxidase (MPO) activity. Standard histological staining hematoxylin and eosin was performed and blindly scored by a pathologist for evidence of lung injury (pulmonary edema, accumulations of neutrophils and mononuclear cells, thickness of alveolar-capillary membrane, perivascular infiltrate, and hemorrhage).. Caerulein-treated wild-type mice exhibited increased lung injury score at 1 through 12 h, as compared to saline controls. As compared to wild-type, STAT6-deficient mice had increased lung injury from 1 to 6 h, with full recovery by 12 h. An opposite pattern was observed in STAT4-deficient mice with mild injury seen at 1 and 6 h, and maximal injury at 12 h. MPO activity was significantly increased at 6 h in caerulein-treated wild-type mice compared to saline-treated controls. Caerulein-treated STAT6 and STAT4 mice had markedly increased MPO activity as compared with their saline controls during the first 6 h. Both caerulein-treated STAT4- and STAT6-deficient mice had significantly increased MPO activity in comparison with wild-type mice with pancreatitis at 6 h.. We found the maximal lung injury after caerulein-induced pancreatitis occurred at different time-points in STAT4 and STAT6-deficient mice. These temporal differences may suggest alternative roles in the systemic inflammatory response associated with pancreatitis.

Topics: Acute Disease; Animals; CD4-Positive T-Lymphocytes; Ceruletide; Disease Models, Animal; Female; Lung; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Pancreas; Pancreatitis; Peroxidase; Respiratory Distress Syndrome; STAT4 Transcription Factor; STAT6 Transcription Factor

Protease-activated receptor-2 (PAR-2) is present in the pancreas, where it has been shown to play a protective role during pancreatitis. However, the mechanism by which it protects against pancreatitis still remains to be elucidated. Acute pancreatitis is associated with premature zymogen activation and a blockage in digestive enzyme secretion.. To examine the effects of PAR-2 activation on the severity of pancreatitis, and to determine whether its protective effects are mediated by affecting either premature activation or secretory blockage, or both.. The results confirmed that PAR-2 -/- mice have more severe pancreatitis than wild-type mice. Interestingly, intrapancreatic trypsin levels in the PAR-2 knockouts remained high after 6 h of pancreatitis, whereas they reverted to normal in the wild types. During pancreatitis, PAR-2 mRNA levels were upregulated in wild-type mice in response to supramaximal caerulein administration. Further, after a single injection of supramaximal caerulein, PAR-2 mRNA levels were also elevated, reaching a peak at 3 h. Stimulating PAR-2 with trypsin or the PAR-2-activating peptide, serine-leucine-isoleucine-glycine-arginine-leucine (SLIGRL), induced significantly more secretion from the acini of these caerulein-sensitised mice compared with the controls. PAR-2 activation also reversed the inhibition of secretion observed in both the caerulein and arginine models.. Trypsin released during the early stages of pancreatitis activates PAR-2 receptors on the acinar cells and stimulates secretion from these cells. Thus, PAR-2 activation may decrease pancreatic injury and limit the severity of pancreatitis by allowing extracellular trypsin to act as a secretagogue.

Topics: Acute Disease; Amylases; Animals; Arginine; Ceruletide; Disease Models, Animal; Enzyme Activation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas, Exocrine; Pancreatitis; Receptor, PAR-2; RNA, Messenger; Tissue Culture Techniques; Transcription Factors; Trypsin; Up-Regulation

Transforming growth factors betas (TGF-betas) are implicated in pancreatic tissue repair but their role in acute pancreatitis is not known. To determine whether endogenous TGF-betas modulate the course of caerulein induced acute pancreatitis, caerulein was administered to wild-type (FVB-/-) and transgenic mice that are heterozygous (FVB+/-) for expression of a dominant negative type II TGF-beta receptor.. After 7 hourly supramaximal injections of caerulein, the pancreas was evaluated histologically and serum was assayed for amylase and lipase levels. Next, the effects of caerulein on amylase secretion were determined in mouse pancreatic acini, and cholecystokinin (CCK) receptor expression was assessed.. The normal mouse pancreas was devoid of inflammatory cells whereas the pancreas from transgenic mice contained lymphocytic infiltrates. Caerulein injection in wild-type mice resulted in 6- and 36-fold increases in serum amylase and lipase levels, respectively, increased serum trypsinogen activation peptide (TAP) levels, gross oedema and a marked inflammatory response in the pancreas that consisted mainly of neutrophils and macrophages. By contrast, FVB+/- mice exhibited minimal alterations in response to caerulein with attenuated neutrophil-macrophage infiltrates. Moreover, acini from FVB+/- mice did not exhibit restricted stimulation at high caerulein concentrations, even though CCK receptor mRNA levels were not decreased.. Our findings indicate that a functional TGF-beta signalling pathway may be required for caerulein to induce acute pancreatitis and for the CCK receptor to induce acinar cell damage at high ligand concentrations. Our results also support the concept that restricted stimulation at high caerulein concentrations contributes to the ability of caerulein to induce acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Blotting, Northern; Ceruletide; Edema; Ligands; Lipase; Mice; Mice, Transgenic; Oligopeptides; Pancreatic Diseases; Pancreatitis; Receptors, Cholecystokinin; Severity of Illness Index; Signal Transduction; Transforming Growth Factor beta

Current markers of exocrine pancreatic toxicity have historically been poor indicators for both early diagnosis of disease and prediction of disease severity. Recently we identified two peptide markers (RA1609 and RT2864) of pancreatic toxicity that are target organ specific. In order to evaluate sensitivity of these markers versus current standard tests for pancreatic damage (i.e., lipase), we measured amylase and lipase, as well as RA1609 and RT2864 marker levels, in serum from rats treated with four doses (50-200 mg/kg) of the model pancreatic toxicant cyanohydroxybutene (CHB). In addition, to determine whether these peptide markers could detect pancreatic injury induced by different toxicants and in different species, we measured RA1609 and RT2864 marker levels in rats treated with the pancreatic toxicant caerulein, and in mice treated with CHB. RA1609 and RT2864 peptide markers proved to be more sensitive than amylase or lipase in detecting pancreatic damage, especially at an early time point (8 h) following CHB administration. The peptide markers also accurately predicted pancreatic injury induced by caerulein in rats. These markers were sensitive in detecting very mild pancreatic damage following CHB administration in mice, which are less susceptible to CHB-induced pancreatic toxicity. In addition, a species comparison of the RA1609 albumin fragment sequence indicated that cleavage of albumin from pancreatic proteases produces a similar fragment marker in several species, including humans. To determine whether the comparable human albumin fragment could be detected in sera from pancreatitis patients, we analyzed sera from normal individuals and from patients with diabetes, vasculitis, pancreatic cancer, and pancreatitis. It was found that markers corresponding to the fragments found in rat serum (RA1609 and RT2864) were present in human serum, and changes in these were indicative of and specific to pancreatitis. In conclusion, the RA1609 and RT2864 peptides are sensitive indicators of exocrine pancreatic damage that may be useful as safety markers for general pancreatic toxicity in multiple species.

Topics: Acute Disease; Alkenes; Amylases; Animals; Biomarkers; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Humans; Lipase; Male; Mice; Nitriles; Pancreas, Exocrine; Pancreatitis; Peptide Fragments; Predictive Value of Tests; Protein Array Analysis; Proteomics; Rats; Rats, Sprague-Dawley; Sensitivity and Specificity; Severity of Illness Index; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Time Factors

Several animal models have been developed to investigate the pathobiology of pancreatitis, but few studies have examined the effects that altered pancreatic gene expression have in these models. In this study, the sensitivity to secretagogue-induced pancreatitis was examined in a mouse line that has an altered acinar cell environment due to the targeted deletion of Mist1. Mist1 is an exocrine specific transcription factor important for the complete differentiation and function of pancreatic acinar cells. Mice lacking the Mist1 gene [Mist1 knockout (KO) mice] exhibit cellular disorganization and functional defects in the exocrine pancreas but no gross morphological defects. Following the induction of pancreatitis with caerulein, a CCK analog, we observed elevated serum amylase levels, necrosis, and tissue damage in Mist1 KO mice, indicating increased pancreatic damage. There was also a delay in the regeneration of acinar tissue in Mist1 KO animals. Molecular profiling revealed an altered activation of stress response genes in Mist1 KO pancreatic tissue compared with wild-type (WT) tissue following the induction of pancreatitis. In particular, Western blot analysis for activating transcription factor 3 and phosphorylated eukaryotic initiation factor 2alpha (eIF2alpha), mediators of endoplasmic reticulum (ER) stress, indicated limited activation of this pathway in Mist1 KO animals compared with WT controls. Conversely, Mist1 KO pancreatic tissue exhibits increased expression of growth arrest and DNA damage inducible 34 protein, an inhibitor of eIF2alpha phosphorylation, before and after the induction of pancreatitis. These finding suggest that activation of the ER stress pathway is a protective event in the progression of pancreatitis and highlight the Mist1 KO mouse line as an important new model for studying the molecular events that contribute to the sensitivity to pancreatic injury.

Topics: Activating Transcription Factor 3; Acute Disease; Amylases; Animals; Antigens, Differentiation; Apoptosis; Basic Helix-Loop-Helix Transcription Factors; Cell Cycle Proteins; Cells, Cultured; Ceruletide; Cholecystokinin; Disease Models, Animal; Dose-Response Relationship, Drug; Endoplasmic Reticulum; Eukaryotic Initiation Factor-2; Gene Expression; Immediate-Early Proteins; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreas, Exocrine; Pancreatitis; Protein Phosphatase 1; Regeneration; RNA, Messenger; Severity of Illness Index; Stress, Physiological; Time Factors

The present study was conducted to examine the contribution of substance P to the pancreatic microcirculatory dysfunction during acute pancreatitis.. Pancreatitis was elicited by up to 6 hourly injections of cerulein (50 microg/kg IP) in male C57Bl/6 mice. At 0, 1, 3, and 6 hours after cerulein treatment, the pancreatic microvasculature in anesthetized mice was studied using established high-resolution in vivo microscopic methods.. Treatment of mice with cerulein for 6 hours caused a 30% decrease in capillary perfusion and the diameter of the capillaries and an increase in microvascular permeability (20%) and interstitial space (30-fold). The administration of the substance P receptor antagonist (D-Arg1, D-Pro2, D-Trp7,9, Leu11) (2 mg/kg IP) minimized the pancreatic microcirculatory dysfunction 3 hours after cerulein treatment. The superfusion of substance P for 0.5 hours decreased the diameter (by 22%) and increased microvascular permeability (by 23%) along with interstitial space (22-fold increase). Blockade of substance P receptor attenuated substance P-induced pancreatic microcirculatory dysfunction.. These results suggest that substance P mediates pancreatic microcirculatory dysfunction during the development of acute pancreatitis.

Topics: Acute Disease; Animals; Capillary Permeability; Ceruletide; Male; Mice; Mice, Inbred C57BL; Microcirculation; Microscopy, Video; Pancreas; Pancreatitis; Substance P

We investigated the effect of a specific neurokinin-1 receptor (NK1R) antagonist, CP-96,345, on the regulation of the expression of adhesion molecules ICAM-1, VCAM-1, E-selectin, and P-selectin as well as leukocyte recruitment during acute pancreatitis (AP). AP was induced in male Balb/C mice by 10 consecutive hourly intraperitoneal injections of caerulein. In the treatment groups, CP-96,345 was administered at 2.5 mg/kg ip either 30 min before or 1 h after the first caerulein injection. Animals were killed, and the lungs and pancreas were isolated for RNA extraction and RT-PCR or for immunohistochemical staining. mRNA expression of the four adhesion molecules was upregulated in the pancreas during AP. Treatment with CP-96,345 effectively reduced the mRNA expression of P-selectin and E-selectin but not ICAM-1 and VCAM-1. In the lung, ICAM-1, E-selectin, and P-selectin mRNA expression increased during AP. Antagonist treatment suppressed this elevation. Similar expression patterns were seen in the immunohistochemical stainings. Intravital microscopy of the pancreatic microcirculation revealed the effect of CP-96,345 on leukocyte recruitment. The present study provides important information on the relationship between NK1R activation and the regulation of adhesion molecules. Also, this study points to the differential regulation of inflammation in the pancreas and lung with AP.

Topics: Acute Disease; Amylases; Animals; Biphenyl Compounds; Cell Adhesion; Ceruletide; Immunohistochemistry; Intercellular Adhesion Molecule-1; Lung; Mice; Mice, Inbred BALB C; Neurokinin-1 Receptor Antagonists; Pancreas; Pancreatitis; Peroxidase; Substance P; Vascular Cell Adhesion Molecule-1

We have hypothesized that the colocalization of digestive zymogens with lysosomal hydrolases, which occurs during the early stages of every experimental pancreatitis model, facilitates activation of those zymogens by lysosomal hydrolases such as cathepsin B and that this activation triggers acute pancreatitis by leading to acinar cell injury. Some, however, have argued that the colocalization phenomenon may be the result, rather than the cause, of zymogen activation during pancreatitis. To resolve this controversy and explore the causal relationships between zymogen activation and other early pancreatitis events, we induced pancreatitis in mice by repeated supramaximal secretagogue stimulation with caerulein. Some animals were pretreated with the cathepsin B inhibitor CA-074 me to inhibit cathepsin B, prevent intrapancreatic activation of digestive zymogens, and reduce the severity of pancreatitis. We show that inhibition of cathepsin B by pretreatment with CA-074 me prevents intrapancreatic zymogen activation and reduces organellar fragility, but it does not alter the caerulein-induced colocalization phenomenon or subcellular F-actin redistribution or prevent caerulein-induced activation of NF-kappaB, ERK1/2, and JNK or upregulated expression of cytochemokines. We conclude 1) that the colocalization phenomenon, F-actin redistribution, activation of proinflammatory transcription factors, and upregulated expression of cytochemokines are not the results of zymogen activation, and 2) that these early events in pancreatitis are not dependent on cathepsin B activity. In contrast, zymogen activation and increased subcellular organellar fragility during caerulein-induced pancreatitis are dependent on cathepsin B activity.

Topics: Actins; Acute Disease; Amylases; Animals; Arylsulfatases; Cathepsin B; Ceruletide; Chemokine CCL2; Dipeptides; Disease Models, Animal; Enzyme Activation; Enzyme Inhibitors; Interleukin-6; JNK Mitogen-Activated Protein Kinases; Lysosomes; Mice; Mice, Inbred C57BL; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; NF-kappa B; Pancreas; Pancreatitis; Protein Transport; Secretory Vesicles; Severity of Illness Index; Time Factors; Trypsin; Trypsinogen

PAP/HIP was first reported as an additional pancreatic secretory protein expressed during the acute phase of pancreatitis. It was shown in vitro to be anti-apoptotic and anti-inflammatory. This study aims to look at whether PAP/HIP plays the same role in vivo.. A model of caerulein-induced pancreatitis was used to compare the outcome of pancreatitis in PAP/HIP(-/-) and wild-type mice.. PAP/HIP(-/-) mice showed the normal phenotype at birth and normal postnatal development. Caerulein-induced pancreatic necrosis was, however, less severe in PAP/HIP(-/-) mice than in wild-type mice, as judged by lower amylasemia and lipasemia levels and smaller areas of necrosis. On the contrary, pancreas from PAP/HIP(-/-) mice was more sensitive to apoptosis, in agreement with the anti-apoptotic effect of PAP/HIP in vitro. Surprisingly, pancreatic inflammation was more extensive in PAP/HIP(-/-) mice, as judged from histological parameters, increased myeloperoxidase activity and increased pro-inflammatory cytokine expression. This result, in apparent contradiction with the limited necrosis observed in these mice, is, however, in agreement with the anti-inflammatory function previously reported in vitro for PAP/HIP. This is supported by the observation that activation of the STAT3/SOCS3 pathway was strongly decreased in the pancreas of PAP/HIP(-/-) mice and by the reversion of the apoptotic and inflammatory phenotypes upon administration of recombinant PAP/HIP to PAP/HIP(-/-) mice.. The anti-apoptotic and anti-inflammatory functions described in vitro for PAP/HIP have physiological relevance in the pancreas in vivo during caerulein-induced pancreatitis.

Topics: Acute Disease; Animals; Apoptosis; Autoantigens; Ceruletide; Disease Models, Animal; Lithostathine; Mice; Mice, Knockout; Necrosis; Pancreas; Pancreatitis; Pancreatitis-Associated Proteins; Phenotype; Proteins; STAT3 Transcription Factor; Suppressor of Cytokine Signaling 3 Protein; Suppressor of Cytokine Signaling Proteins

TNF-alpha plays a pivotal role in the pathogenesis of acute pancreatitis. Recent studies have shown that TNF-alpha inhibition significantly ameliorates the course of experimental acute pancreatitis, but in this context, the effects of Etanercept, a novel anti-TNF-alpha agent, have not been investigated so far. The aims of the present study are (i) to assess the effects of pharmacological inhibition of TNF-alpha by means of Etanercept on the inflammatory response and apoptosis in a murine model of necrotizing acute pancreatitis and (ii) to compare the results to those observed in TNF-alpha receptor 1 knockout (TNFR1-KO) mice. Necrotizing acute pancreatitis was induced in TNF-alpha wild type for TNFR1 (WT) and TNFR1-KO mice by intraperitoneal injection of cerulein (hourly x5, 50 microg/kg). In another group of WT mice, Etanercept was administered (5 or 10 mg/kg, s.c.) at 1 h after first cerulein injection. Control groups received saline treatment. After 24 h, biochemical, histological, and immunohistochemical evidences of acute pancreatitis developed in all cerulein-treated mice; apoptosis was also present in the pancreas. Contrarily, pancreatitis histological features, amylase and lipase levels, pancreas water content, and myeloperoxidase activity were reduced in a similar degree in Etanercept-treated and TNFR1-KO mice. Likewise, in these two groups, immunohistochemical stainings and terminal deoxynucleotidyltransferase-mediated UTP nick-end labeling assay were found negative. TNF-alpha receptor 1 gene deletion and Etanercept administration ameliorate the course of experimental acute pancreatitis in a similar degree. Future studies on clinical applications of Etanercept in pancreatitis seem promising.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; bcl-2-Associated X Protein; bcl-Associated Death Protein; Ceruletide; Enzyme-Linked Immunosorbent Assay; Etanercept; Fas Ligand Protein; Gene Deletion; Immunoglobulin G; Immunohistochemistry; Inflammation; Intercellular Adhesion Molecule-1; Lymphotoxin-alpha; Mice; Mice, Knockout; P-Selectin; Pancreatitis; Peroxidase; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

Bacterial endotoxin (lipopolysaccharide, LPS), at high concentration is responsible for sepsis, and neonatal mortality, however low concentration of LPS protected the pancreas against acute damage. The aim of this study was to investigate the effect of exposition of suckling rats to LPS on the course of acute pancreatitis at adult age. Suckling rat (30-40g) received intraperitoneal (i.p.) injection of saline (control) or LPS from Escherichia coli or Salmonella typhi (5, 10 or 15 mg/kg-day) during 5 consecutive days. Two months later these rats have been subjected to i.p. cearulein infusion (25 microg/kg) to produce caerulein-induced pancreatitis (CIP). The following parameters were tested: pancreatic weight and morphology, plasma amylase and lipase activities, interleukin 1beta (IL-1 beta), interleukin 6 (IL-6), and interleukin 10 (IL-10) plasma concentrations. Pancreatic concentration of superoxide dismutase (SOD) and lipid peroxidation products; malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) have been also measured. Caerulein infusion produced CIP in all animals tested, that was confirmed by histological examination. In the rats, which have been subjected in the neonatal period of life to LPS at doses 10 or 15 mg/kg-day x 5 days, all manifestations of CIP have been reduced. In these animals acute inflammatory infiltration of pancreatic tissue and pancreatic cell vacuolization have been significantly diminished. Also pancreatic weight, plasma lipase and alpha-amylase activities, as well as plasma concentrations of IL-1beta and IL-6 have been markedly decreased, whereas plasma anti-inflammatory IL-10 concentration was significantly increased in these animals as compared to the control rats, subjected in the infancy to saline injection instead of LPS. Caerulein-induced fall in pancreatic SOD concentration was reversed and accompanied by significant reduction of MDA + 4 HNE in the pancreatic tissue. The effects of LPS derived from E. coli or S. typhi were similar. Pretreatment of suckling rats with LPS at dose of 10 mg/kg-day x 5 days resulted in the most prominent attenuation of acute pancreatitis at adult age, whereas LPS at dose of 5 mg/kg-day x 5 days given to the neonatal rats failed to affect significantly acute pancreatitis induced in these animals 2 months later. We conclude that: 1/ Prolonged exposition of suckling rats to bacterial endotoxin attenuated acute pancreatitis induced in these animals at adult age. 2/ This effect could be related to

Topics: Acute Disease; Aldehydes; alpha-Amylases; Animals; Animals, Newborn; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Endotoxemia; Interleukins; Lipase; Lipid Peroxidation; Lipopolysaccharides; Male; Malondialdehyde; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Severity of Illness Index; Superoxide Dismutase; Time Factors

Topics: Acute Disease; Animals; Ascitic Fluid; Ceruletide; Deoxycholic Acid; Disease Models, Animal; High Mobility Group Proteins; HMGB1 Protein; Intestine, Small; Kidney; Liver; Lung; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Repressor Proteins; Severity of Illness Index; Time Factors

The functional involvement of the endocannabinoid system in modulation of pancreatic inflammation, such as acute pancreatitis, has not been studied to date. Moreover, the therapeutic potential of cannabinoids in pancreatitis has not been addressed.. We quantified endocannabinoid levels and expression of cannabinoid receptors 1 and 2 (CB1 and CB2) in pancreas biopsies from patients and mice with acute pancreatitis. Functional studies were performed in mice using pharmacological interventions. Histological examination, serological, and molecular analyses (lipase, myeloperoxidase, cytokines, and chemokines) were performed to assess disease pathology and inflammation. Pain resulting from pancreatitis was studied as abdominal hypersensitivity to punctate von Frey stimuli. Behavioral analyses in the open-field, light-dark, and catalepsy tests were performed to judge cannabinoid-induced central side effects.. Patients with acute pancreatitis showed an up-regulation of cannabinoid receptors and elevated levels of endocannabinoids in the pancreas. HU210, a synthetic agonist at CB1 and CB2, abolished abdominal pain associated with pancreatitis and also reduced inflammation and decreased tissue pathology in mice without producing central, adverse effects. Antagonists at CB1- and CB2-receptors were effective in reversing HU210-induced antinociception, whereas a combination of CB1- and CB2-antagonists was required to block the anti-inflammatory effects of HU210 in pancreatitis.. In humans, acute pancreatitis is associated with up-regulation of ligands as well as receptors of the endocannabinoid system in the pancreas. Furthermore, our results suggest a therapeutic potential for cannabinoids in abolishing pain associated with acute pancreatitis and in partially reducing inflammation and disease pathology in the absence of adverse side effects.

Topics: Acute Disease; Animals; Biopsy; Cannabinoid Receptor Modulators; Cannabinoids; Ceruletide; Dronabinol; Female; Gene Expression Regulation; Humans; Male; Mice; Mice, Inbred C57BL; Neuroprotective Agents; Pain; Pancreas; Pancreatitis; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2

We explored the pathophysiological roles of IFN-gamma in cerulein-induced acute pancreatitis. In wild-type (WT) mice, cerulein injection caused acute pancreatitis as evidenced by increased serum amylase levels and pathological changes such as interstitial edema, vacuolization, acinar cell necrosis, and neutrophil infiltration in pancreas. Concomitantly, cerulein treatment augmented intrapancreatic gene expression of TNF-alpha, KC/CXCL1, MIP-2/CXCL2, cyclooxygenase-2 (COX-2), and IFN-gamma in WT mice. In situ hybridization combined with immunofluorescence analyses demonstrated that infiltrating neutrophils expressed IFN-gamma mRNA. Unexpectedly, IFN-gamma(-/-) mice exhibited exacerbated cerulein-induced pancreatic injury, with enhanced neutrophil recruitment. Moreover, intrapancreatic gene expression of TNF-alpha, KC/CXCL1, MIP-2/CXCL2, and COX-2 were significantly exaggerated in IFN-gamma(-/-) mice, compared with WT mice. Cerulein activated NF-kappaB, an indispensable transcription factor for gene transcription of TNF-alpha, KC/CXCL1, MIP-2/CXCL2, and COX-2, in pancreas of cerulein-treated WT mice as evidenced by the increases in nuclear amount and DNA-binding activity of NF-kappaB p65. In comparison with WT mice, IFN-gamma(-/-) mice exhibited exaggerated and prolonged NF-kappaB activation, probably due to reduced acetylation of Stat1, a main signal transducer of IFN-gamma, because acetylated Stat1 can inhibit NF-kappaB activation. Indeed, IFN-gamma acetylated Stat1 and reciprocally reduced NF-kappaB activation and COX-2 expression in neutrophils. Finally, even when administered 4 h after the first cerulein injection, IFN-gamma remarkably attenuated acute pancreatitis in both WT and IFN-gamma(-/-) mice, with reduced NF-kappaB activation and COX-2 expression. Thus, IFN-gamma can have anti-inflammatory effects on acute pancreatitis by depressing the proinflammatory consequences of NF-kappaB activation.

Topics: Acute Disease; Adjuvants, Immunologic; Animals; Cell Movement; Ceruletide; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Injections, Intraperitoneal; Interferon-gamma; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Neutrophils; NF-kappa B; Pancreas; Pancreatitis; Repressor Proteins; RNA, Messenger; Up-Regulation

To determine the effect of exogenous leptin on acute lung injury (ALI) in cerulein-induced acute pancreatitis (AP).. Forty-eight rats were randomly divided into 3 groups. AP was induced by intraperitoneal (i.p.) injection of cerulein (50 microg/kg) four times, at 1 h intervals. The rats received a single i.p. injection of 10 mug/kg leptin (leptin group) or 2 mL saline (AP group) after cerulein injections. In the sham group, animals were given a single i.p. injection of 2 mL saline. Experimental samples were collected for biochemical and histological evaluations at 24 h and 48 h after the induction of AP or saline administration. Blood samples were obtained for the determination of amylase, lipase, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, macrophage inflammatory peptide (MIP)-2 and soluble intercellular adhesion molecule (sICAM)-1 levels, while pancreatic and lung tissues were removed for myeloperoxidase (MPO) activity, nitric oxide (NOx) level, CD40 expression and histological evaluation.. Cerulein injection caused severe AP, confirmed by an increase in serum amylase and lipase levels, histopathological findings of severe AP, and pancreatic MPO activity, compared to the values obtained in the sham group. In the leptin group, serum levels of MIP-2, sICMA-1, TNF-alpha, and IL-1beta, pancreatic MPO activity, CD40 expression in pancreas and lung tissues, and NOx level in the lung tissue were lower compared to those in the AP group. Histologically, pancreatic and lung damage was less severe following leptin administration.. Exogenous leptin attenuates inflamma-tory changes, and reduces pro-inflammatory cytokines, nitric oxide levels, and CD40 expression in cerulein-induced AP and may be protective in AP associated ALI.

Topics: Acute Disease; Animals; CD40 Antigens; Ceruletide; Chemokine CXCL2; Chemokines, CXC; Female; Interleukin-1beta; Leptin; Lung; Nitric Oxide; Pancreas; Pancreatitis; Peroxidase; Random Allocation; Rats; Rats, Wistar; Respiratory Distress Syndrome; Tumor Necrosis Factor-alpha

Previous studies have shown that ischemic preconditioning protects several organs, including the pancreas, from ischemia/reperfusion-induced injury. The aim of the investigation was to determine whether ischemic preconditioning affects the course edematous pancreatitis.. In rats, ischemic preconditioning was performed by short-term clamping the celiac artery. Acute pancreatitis was induced by caerulein. The severity of acute pancreatitis was evaluated between the first and tenth day of inflammation.. Ischemic preconditioning applied alone caused a mild pancreatic damage. Combination of ischemic preconditioning with caerulein attenuated the severity of pancreatitis in histological examination and reduced the pancreatitis-evoked increase in plasma lipase and pro-inflammatory interleukin-1beta. This effect was associated with an increase in plasma level of anti-inflammatory interleukin-10 and partial reversion of the pancreatitis-evoked drop in pancreatic DNA synthesis and pancreatic blood flow. In secretory studies, ischemic preconditioning in combination with induction of acute pancreatitis attenuated the pancreatitis-evoked decrease in secretory reactivity of isolated pancreatic acini to stimulation by caerulein. In the initial period of acute pancreatitis, ischemic preconditioning alone and in combination with caerulein-induced acute pancreatitis prolonged the activated partial thromboplastin time (APTT), increased plasma level of D-dimer and shortened the euglobulin clot lysis time. The protective effect of ischemic preconditioning was observed during entire time of experiment and led to acceleration of pancreatic regeneration.. Ischemic preconditioning reduces the severity of caerulein-induced pancreatitis and accelerates pancreatic repair; and this effect is related to the activation of fibrinolysis and reduction of inflammatory process.

Topics: Acute Disease; Amylases; Animals; Blood Coagulation; Ceruletide; Fibrin Fibrinogen Degradation Products; Fibrinolysis; Interleukin-10; Ischemic Preconditioning; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar

Protein kinase C-zeta (PKC-zeta) regulates cell death via NF-kappaB; therefore, we tested the hypothesis that PKC-zeta plays a critical role in pancreatitis-induced Kupffer cell apoptosis. Acute pancreatitis was induced in rats by cerulein injection 24 h later, livers were assayed for PKC-zeta, IKKalpha, IKKbeta, IKKgamma, NF-kappaB, Fas/FasL, and apoptosis was assessed with Caspase-3 and DNA fragmentation. Kupffer cells from unoperated rats were infected with a PKC-zeta domain-negative adenovirus (AdPKCzeta-DN) to inhibit PKC-zeta, or transfected with pCMVPKC-zeta to overexpress PKC-zeta, and then stimulated with pancreatic elastase; cellular extracts were assayed for PKC-zeta, IKKalpha, IKKbeta, IKKgamma, NF-kappaB, Fas/FasL, Caspase-3, and DNA fragmentation. Cerulein-induced pancreatitis upregulated PKC-zeta protein and activity, IKKbeta, IKKgamma, NF-kappaB, Fas/FasL, Caspase-3 and increased DNA fragmentation in rat livers (all p < 0.001 vs control). AdPKCzeta-DN abolished elastase-induced upregulation of PKC-zeta activity, IKKbeta, IKKgamma, NF-kappaB, Fas/FasL, Caspase-3 and DNA fragmentation (all p < 0.001 vs infection control), whereas overexpression of PKC-zeta augmented elastase-induced upregulation of IKKbeta, IKKgamma, Fas/FasL, Caspase-3 and DNA fragmentation (p < 0.001 vs control). PKC-zeta plays a critical role in pancreatitis-induced Kupffer cell apoptosis via NF-kappaB and Fas/FasL. The ability of Kupffer cells to autoregulate their stress response by upregulating their death receptor/ligand and key proapoptotic cell signaling systems warrants further investigation.

Topics: Acute Disease; Animals; Apoptosis; Caspase 3; Ceruletide; DNA Fragmentation; I-kappa B Kinase; In Vitro Techniques; Isoenzymes; Kupffer Cells; Male; NF-kappa B; Pancreatitis; Phosphorylation; Protein Kinase C; Protein Serine-Threonine Kinases; Rats; Rats, Sprague-Dawley; Up-Regulation

Hypothermia is a frequent event in severe acute pancreatitis (AP) and its real effects on the normal pancreas have not been well demonstrated. Moreover, neither have its effects on the outcome of acute pancreatitis been fully investigated. One hypothesis is that oxidative stress may be implicated in lesions caused or treated by hypothermia.. To investigate the effect of hypothermia in cerulein-induced acute pancreatitis (CIAP) in rats and the role played by oxidative stress in this process.. Male Wistar rats were divided into hypothermic and normothermic groups. Hypothermia was induced with a cold mattress and rectal temperature was kept at 30 masculineC for one hour. Acute pancreatitis was induced with 2 doses of cerulein (20 ìg/kg) administered at a one-hour interval. Serum amylase, pancreas vascular permeability by Evan's blue method, pancreas wet-to-dry weight ratio and histopathology were analyzed in each group.. When compared with normothermic rats, hypothermic animals, with cerulein-induced acute pancreatitis, showed higher levels of pancreatic vascular permeability (p < 0.05), pancreas wet-to-dry weight ratio (p = 0.03), and histologically verified edema (p < 0.05), but similar serum amylase levels. The hypothermic group showed a higher oxidized-reduced glutathione ratio than the normothermic group.. Moderate hypothermia produced a greater inflammatory response in established acute pancreatitis induced by cerulein in rats. Moreover, this study suggests that oxidative stress may be one of the mechanisms responsible for the worse outcome in hypothermic rats with cerulein-induced acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Capillary Permeability; Ceruletide; Glutathione; Hypothermia; Male; Oxidative Stress; Pancreatitis; Rats; Rats, Wistar

Ghrelin and its receptor are expressed abundantly in the stomach and pituitary. Recently, a ghrelin system, consisting of both ligand and receptor, has also been found to exist in the endocrine cells of pancreatic islets. This ghrelin system may play a role in regulating insulin secretion and glucose homeostasis. The aim of the present study was to investigate whether a functional ghrelin system also exists in the exocrine pancreas.. Precise localization and expression of ghrelin and its receptor in rat pancreatic acinar cells were examined by immunocytochemistry and Western blot, whereas messenger RNA levels were examined by semiquantitative reverse transcription-polymerase chain reaction. The roles of physiological and pathophysiological conditions, such as gastric acid inhibition, starvation, and acute pancreatitis, in regulation of ghrelin and its receptor were also examined.. Both ghrelin and its receptor were detected, at both protein and messenger RNA levels, in the acinar cells of the exocrine pancreas. Ghrelin receptor expression was up-regulated by gastric acid inhibition and down-regulated by acute pancreatitis, whereas levels remained unchanged after food deprivation. In contrast, ghrelin expression did not exhibit significant changes in any condition.. Our data indicate that a ghrelin system exists in the acinar cells of the exocrine pancreas. This system is subject to regulation by physiological and pathophysiological stimuli and may thus regulate exocrine functions by paracrine and/or autocrine mechanisms.

Topics: Achlorhydria; Acute Disease; Animals; Autocrine Communication; Ceruletide; Food Deprivation; Gastric Acid; Gene Expression Regulation; Ghrelin; Male; Omeprazole; Pancreas, Exocrine; Pancreatitis; Paracrine Communication; Peptide Hormones; Random Allocation; Rats; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; Receptors, Ghrelin; RNA, Messenger

We investigated the effect of Adrenomedullin (AM) on cerulein-induced acute pancreatitis in rats. AM treatment (100 ng/kg per rat, subcutaneous) after one hour of cerulein injection reduced the plasma amylase levels, pancreatic weight, pancreatic malondialdehyde (MDA) levels, and the severity of the lesions microscopically. These data suggest that AM has a protective effect on cerulein-induced acute pancreatitis. These could be due to anti-inflammatory properties of AM, inhibition of proinflammatory cytokine secretion, reducing the endothelial permeability increased by reactive oxygen species, endotoxins or cytokines.

Topics: Acute Disease; Adrenomedullin; Amylases; Animals; Ceruletide; Female; Injections, Subcutaneous; Lipid Peroxidation; Male; Malondialdehyde; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

To observe the apoptosis and oncosis of pancreatic acinar cells and secondary inflammatory reaction in pancreatic tissue from rats with acute pancreatitis (AP), and the influences of artemisinin on them.. AP was induced by 4 intraperitoneal injections of caerulein at 1 h intervals. To induce apoptosis, solution of artemisinin (50 mg/kg) was given intraperitoneally 1, 12, 24 and 36 h after the last caerulein injection. Histological examination of impairment of pancreatic tissue and detection of serum amylase were performed to evaluate the severity of acute pancreatitis. Apoptosis and oncosis were detected with acridine orange (AO) and ethylene dibromide (EB) staining. Caspase-3 and myeloperoxidase (MPO) activity were measured by colorimetric assay. Nuclear factor-kappa B (NF-kappaB) activation was detected by flow cytometry. Macrophage inflammatory protein-1alpha (MIP-1alpha) protein was measured by Western blot. Interleukin-1beta (IL-1beta) mRNA was detected by RT-PCR.. Addition of artemisinin increased the number of apoptotic cells (11.7% +/- 1.4% vs 6.3% +/- 0.7%, P < 0.05), while reduced the number of oncotic cells (13.0% +/- 2.4% vs 17.5% +/- 2.2%, P < 0.05). The activity of caspase-3 speeded up (1.52 +/- 0.21 vs 1.03 +/- 0.08, P < 0.05), the pancreas pathological impairment was relieved (3.0 +/- 0.5 vs 4.0 +/- 0.5, P < 0.05) and the level of serum amylase decreased (5642 +/- 721 U/dL vs 7821 +/- 653 U/dL, P < 0.05). The activation of NF-kB (29% +/- 4.1% vs 42% +/- 5.8%), MIP-1alpha protein (3.7 +/- 0.5 vs 5.8 +/- 0.7), MPO (0.52 +/- 0.06 U/g vs 0.68 +/- 0.09 U/g), IL-1beta mRNA (1.7 +/- 0.3 vs 2.4 +/- 0.4) in the apoptosis inducing group was obviously decreased (P < 0.05).. Inducing apoptosis can relieve pathological impairment and inflammatory reaction in AP rats.

Topics: Acute Disease; Animals; Apoptosis; Artemisinins; Caspase 3; Ceruletide; Male; NF-kappa B; Pancreatitis; Peroxidase; Rats; Rats, Wistar

The aim of the present study was to elucidate the pathogenic role of endogenous 5-HT in pancreatitis. Injections of cerulein at hourly intervals caused edematous pancreatitis in mice characterized by hyperenzymemia and histological alterations. While the cerulein-induced hyperenzymemia was attenuated in mice pretreated with p-CPA, a 5-HT depletor, it was exaggerated by the preferential 5-HT2A agonist (DOI), but not by the preferential 5-HT2B agonist (BW723C86) or the preferential 5-HT2C agonist (mCPP). Selective 5-HT2A antagonists (risperidone, spiperone, ketanserin, AMI-193, and MDL 11,939) dose-dependently attenuated the hyperenzymemia; and their potency order, excepting that of ketanserin which has considerable affinity at the 5-HT2C receptor as well, paralleled their reported pKi values at the 5-HT2A receptor. Selective 5-HT2B (SB204741) and 5-HT2C (SB242084) antagonists hardly affected the hyperenzymemia. Although the non-selective 5-HT2A/2B/2C antagonists (metergoline, ritanserin, and methysergide) dose-dependently attenuated the hyperenzymemia, they were relatively less potent compared to their high pKi values at the 5-HT2A receptor. In another set of experiments, risperidone, but not SB204741 and SB242084, dose-dependently reversed the cerulein-induced histological alteration of the pancreas (inflammatory cell infiltration). These results suggest that endogenously released 5-HT activates 5-HT2A receptors to aggravate cerulein-induced pancreatitis. We propose that selective 5-HT2A antagonists may provide a new therapy for acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Female; Mice; Mice, Inbred ICR; Pancreas; Pancreatitis; Receptor, Serotonin, 5-HT2A; Receptors, Cholecystokinin; Serotonin; Serotonin Antagonists; Serotonin Receptor Agonists; Spiro Compounds

Glycogen synthase kinase (GSK)-3 is a ubiquitous serine-threonine protein kinase that participates in a multitude of cellular processes and signal transduction pathways. It also plays an important role in the pathophysiology of a number of diseases characterized by an enhanced or unregulated inflammatory response. Here we investigate the effects of GSK-3beta inhibition on the development of experimental acute pancreatitis induced by cerulein in mice.. Prospective, randomized study.. University-based research laboratory.. One-hundred and sixty anesthetized male CD mice.. Pancreatitis was induced by intraperitoneal injection of cerulein (hourly x5, 50 microg/kg). In the treatment group, the potent and selective GSK-3beta inhibitor 4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8) was administered 1 hr and 6 hrs after the first injection of cerulein (10 mg/kg, intraperitoneally). Sham groups were treated with vehicle (0.1 mL of 0.9% NaCl, intraperitoneally) and TDZD-8. In another set of experiments, mice were monitored for 24 days to determine their mortality rate.. The injection of cerulein resulted in acute necrotizing pancreatitis. TDZD-8 significantly reduced the degree of pancreas injury, amylase, and lipase serum levels (p < .01); nuclear factor-kappaB activation (p < .01); the production of tumor necrosis factor-alpha and interleukin-1beta (p < .01); the expression of adhesion molecules and neutrophil accumulation (p < .01); the formation of oxygen and nitrogen-derived radicals (p < .01); the degree of lipid peroxidation (p < .01); the expression of transforming growth factor-beta and vascular endothelial growth factor (p < .01); and-ultimately-the mortality rate (p < .01).. Inhibition of GSK-3beta reduces the degree of cerulein-induced acute pancreatitis and the associated mortality rate in mice. Blocking protein kinase activity may be a novel approach to treatment of this inflammatory condition.

Topics: Acute Disease; Animals; Cell Adhesion Molecules; Ceruletide; Cytokines; Enzyme Inhibitors; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Male; Mice; Mice, Inbred Strains; Neutrophil Activation; NF-kappa B; Oxidative Stress; Pancreatitis; Prospective Studies; Random Allocation; Survival Analysis; Thiadiazoles

To test the hypothesis that disruption of acinar cell membranes is the earliest event that takes place after the onset of acute pancreatitis.. Cerulein and taurocholate pancreatitis were induced in rats. Furthermore, stimulation with different doses of bombesin, pilocarpine, and cerulein was performed. Five to 180 minutes after initiation of treatment, animals were killed. Disruption of cell membranes was detected by the penetration of the experimental animal's own albumin or immunoglobulin G (IgG) into acinar cells by immunocytological localization. Tissue was further analyzed by electron microscopy and electron microscopic immunostaining.. Animals with pancreatitis displayed significantly greater antialbumin and anti-IgG immunostaining in the cytoplasm of acinar cells and in vacuoles in comparison with controls, confirming membrane disruption. This was not detectable after stimulation with bombesin, pilocarpine, and nonsupramaximal doses of cerulein. The first changes were seen after 5 minutes of induction of pancreatitis. Results were verified by electron microscopy and electron microscopic immunohistochemistry.. The penetration of albumin and IgG into acinar cells indicates that wounding of their plasma membrane occurs at the onset of acute pancreatitis. Disruption of the membranes could be expected to allow the influx of calcium ions, causing massive intracellular alterations, and exit of molecules, such as enzymes from acinar cells.

Topics: Acute Disease; Animals; Capillary Permeability; Cell Membrane; Ceruletide; Cytoplasm; Disease Models, Animal; Edema; Immunoglobulin G; Male; Microscopy, Electron, Transmission; Microscopy, Immunoelectron; Pancreas, Exocrine; Pancreatitis; Rats; Rats, Sprague-Dawley; Serum Albumin; Taurocholic Acid; Time Factors; Vacuoles

To study if the course of cerulein-induced pancreatitis in rats changes in a state of triglyceride-rich lipoprotein metabolism alteration.. Two groups of rats received control diet during a 90-day period (A) and sucrose-rich diet to induce endogenous hypertriglyceridemia (B). Subgroups A2 and B2 received i.p. 45 microg cerulein/kg body weight (to induce acute pancreatitis). Histological examination of pancreas tissue, serum pancreatic lipase, lipoprotein profile and VLDL chemical composition were assessed. Then, pancreatic lipase hydrolytic activity on VLDL-triglycerides was evaluated in vitro.. Cellular vacuolization was observed in all of the cerulein-injected rats, but only in subgroup B2 fat necrosis was present. Serum triglycerides were higher in subgroup B1 than in subgroup A1 (mean +/- SEM, mg/dl 123,77 +/- 25.7 vs. 65.8 +/- 7, p < 0.01). Triglycerides from rats fed with sucrose-rich diet, decreased after cerulein-induced pancreatitis (80.38 +/- 11.3 vs. 123,77 +/- 25.7, p < 0.02). Moreover, the endogenous hypertriglyceridemic rats showed an increment of VLDL triglyceride content, which decreased when rats were injected with cerulein. A negative correlation was found between VLDL-triglyceride content and serum pancreatic lipase activity (r = 0.58, p < 0.02). The in vitro assay showed a decrease in VLDL-triglyceride content post incubation with pancreatic lipase enriched serum (mean +/- SD: 59.2 +/- 27.7%, p < 0.01).. The endogenous hypertriglyceridemia intensifies the course of cerulein-induced pancreatitis and it could be related to the decrease in VLDL-triglycerides as a consequence of pancreatic lipase hydrolytic activity.

Topics: Acute Disease; Animals; Ceruletide; Cholesterol, VLDL; Hypertriglyceridemia; Lipase; Lipoproteins, VLDL; Male; Pancreatitis; Random Allocation; Rats; Rats, Wistar; Triglycerides

The role of oxidative stress has been evaluated in experimental models of acute pancreatitis (AP). The aim of this study is to investigate the effect of melatonin on the ultrastructural changes in cerulein-induced AP in rats. Acute pancreatitis was induced by two i.p. injections of cerulein at 2-hr intervals (50 microg/kg BW). One group received additionally melatonin (20 mg/kg BW) i.p. before each injection of cerulein. The rats were sacrificed 12 hr after the last injection. Pancreatic oxidative stress markers were evaluated by changes in the amount of lipid peroxides and changes in the antioxidant enzyme levels, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and total glutathione (GSH) levels. Ultrastructural examination was performed using a transmission electron microscope. Formation of numerous, large autophagosomes, mitochondrial damage, dilatation of rough endoplasmic reticulum (RER) and Golgi apparatus, margination and clumping of nuclear chromatin were the major ultrastructural alterations observed in the AP group. Melatonin administration prevented mitochondrial and nuclear changes and dilatation of RER and Golgi apparatus. Rare, small autophagosomes were present within the cytoplasm of some of the acinar cells. Pancreatic damage was accompanied by a significant increase in tissue MDA levels (P < 0.05) and a significant decrease in CAT, SOD, GPx activities and GSH levels (P < 0.005). Melatonin administration significantly reduced MDA levels but increased CAT, SOD, GPx activities and GSH levels (P < 0.005). Melatonin also reduced serum amylase and lipase activities, which were significantly elevated in AP (P < 0.05 and P < 0.005 respectively). These results suggest that oxidative injury is important in the pathogenesis of AP. Melatonin is potentially capable of limiting pancreatic damage produced during AP by protecting the fine structure of acinar cells and tissue antioxidant enzyme activities.

Topics: Acute Disease; Animals; Apoptosis; Autophagy; Catalase; Ceruletide; Female; Glutathione; Glutathione Peroxidase; Lipid Peroxidation; Melatonin; Microscopy, Electron; Oxidative Stress; Pancreas; Pancreatitis; Phagosomes; Rats; Rats, Sprague-Dawley; Superoxide Dismutase

To investigate the role of oxidative injury in pancreatitis-induced hepatic damage and the effect of antioxidant agents such as melatonin, ascorbic acid and N-acetyl cysteine on caerulein-induced pancreatitis and associated liver injury in rats.. Thirty-eight female Wistar rats were used. Acute pancreatitis (AP) was induced by two i.p. injections of caerulein at 2-h intervals (at a total dose of 100 microg/kg b.wt). The other two groups received additional melatonin (20 mg/kg b.wt) or an antioxidant mixture containing L(+)-ascorbic acid (14.3 mg/kb.wt.) and N-acetyl cysteine (181 mg/kg b.wt.) i.p. shortly before each injection of caerulein. The rats were sacrificed by decapitation 12 h after the last injection of caerulein. Pancreatic and hepatic oxidative stress markers were evaluated by changes in the amount of lipid peroxides measured as malondialdehyde (MDA) and changes in tissue antioxidant enzyme levels, catalase (CAT) and glutathione peroxidase (GPx). Histopathological examination was performed using scoring systems.. The degree of hepatic cell degeneration, intracellular vacuolization, vascular congestion, sinusoidal dilatation and inflammatory infiltration showed a significant difference between caerulein and caerulein + melatonin (P = 0.001), and careulein and caerulein + L(+)-ascorbic acid + N-acetyl cysteine groups (P = 0.002). The degree of aciner cell degeneration, pancreatic edema, intracellular vacuolization and inflammatory infiltration showed a significant difference between caerulein and caerulein + melatonin (P = 0.004), and careulein and caerulein + L(+)-ascorbic acid + N-acetyl cysteine groups (P = 0.002). Caerulein-induced pancreatic and liver damage was accompanied with a significant increase in tissue MDA levels (P = 0.01, P = 0.003, respectively) whereas a significant decrease in CAT (P = 0.002, P = 0.003, respectively) and GPx activities (P = 0.002, P = 0.03, respectively). Melatonin and L(+)-ascorbic acid+N-acetyl cysteine administration significantly decreased MDA levels in pancreas (P=0.03, P=0.002, respectively) and liver (P = 0.007, P = 0.01, respectively). Administration of these agents increased pancreatic and hepatic CAT and GPx activities. Melatonin significantly increased pancreatic and hepatic CAT (P = 0.002, P = 0.001, respectively) and GPx activities (P = 0.002, P = 0.001). Additionally, L(+)-ascorbic acid + N-acetyl cysteine significantly increased pancreatic GPx (P = 0.002) and hepatic CAT and GPx activities (P = 0.001, P = 0.007, respectively).. Oxidative injury plays an important role not only in the pathogenesis of AP but also in pancreatitis-induced hepatic damage. Antioxidant agents such as melatonin and ascorbic acid + N-acetyl cysteine, are capable of limiting pancreatic and hepatic damage produced during AP via restoring tissue antioxidant enzyme activities.

Topics: Acetylcysteine; Acute Disease; Animals; Antioxidants; Ascorbic Acid; Ceruletide; Female; Lipid Peroxidation; Liver; Melatonin; Pancreatitis; Rats; Rats, Wistar; Reactive Oxygen Species

A considerable body of recent evidence suggests that oxidative stress and exaggerated production of reactive oxygen species play a major role in several aspects of inflammation and shock. Hypericum perforatum is a medicinal plant species containing many polyphenolic compounds, namely flavonoids and phenolic acids. Because polyphenolic compounds have high antioxidant potential, in this study we evaluated the effect of Hypericum perforatum extract on acute pancreatitis induced by cerulein administration in male CD mice. Intraperitoneal injection of cerulein in mice resulted in a severe, acute pancreatitis, which was characterized by edema, neutrophil infiltration, tissue hemorrhage, and cell necrosis as well as increases in the serum levels of amylase and/or lipase in comparison to sham-treated mice. The infiltration of the pancreatic tissue of these animals with neutrophils (measured as increase in myeloperoxidase activity) was associated with expression of the adhesion molecule ICAM-1. Immunohistochemical examination demonstrated a marked increase in the staining (immunoreactivity) for nitrotyrosine and poly(ADP-ribose) (PAR) in the pancreas of cerulein-treated mice in comparison to sham-treated mice. In contrast, the degree of (a) pancreatic inflammation and tissue injury (histological score), (b) expression of ICAM-1, (c) the staining for nitrotyrosine and PAR, and (d) myeloperoxidase activity was markedly reduced in pancreatic tissue sections obtained from cerulein-treated mice administered Hypericum perforatum extract (30 mg/kg, suspended in 0.2 mL of saline solution, o.s.). Moreover, the treatment with Hypericum perforatum extract significantly reduced the mortality rate at 5 days after cerulein administration. Taken together, our results indicate that Hypericum perforatum extract reduces the development of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Flavonoids; Hypericum; Inflammation; Male; Mice; Pancreatitis; Phenols; Phytotherapy; Plant Extracts; Polyphenols

To determine whether neutrophil depletion and Kupffer cell inhibition might combine their protective effects to decrease the severity of acute pancreatitis.. Mice had cerulein administration to induce acute pancreatitis and were pretreated with either anti-mouse neutrophil serum or gadolinium chloride (GdCl3) to prevent Kupffer cell activation, or both treatments. Injury was assessed in pancreas and lungs. Myeloperoxidases (MPO) assessed neutrophil infiltration. Interleukin-6 (IL-6) and IL-10 were measured in serum, pancreas, lungs and liver.. In mice with acute pancreatitis, neutrophil depletion reduced the severity of pancreatitis and pancreatitis-associated lung injury. Kupffer cell inactivation by GdCl3 had less protective effect, although IL-6 and IL-10 concentrations were significantly decreased. The protective treatment brought by neutrophil depletion was not enhanced by Kupffer cell inactivation and both treatments did not combine their protective effects.. Our results confirm the role of activated neutrophils in aggravating organ injury in acute pancreatitis while the role of Kupffer cell activation is less obvious.

Topics: Acute Disease; Amylases; Animals; Cell Count; Ceruletide; Gadolinium; Interleukins; Kupffer Cells; Liver; Lung; Macrophage Activation; Male; Mice; Mice, Inbred C57BL; Neutrophil Activation; Neutrophils; Pancreas; Pancreatitis; Peroxidase; Random Allocation; Severity of Illness Index

Acute pancreatitis accompanied by a subsequent infectious attack can often lead to multisystem organ dysfunction, including acute lung injury (ALI), but the molecular mechanisms are poorly defined. In this study, we explored the role of the priming insult by induction of cerulein pancreatitis, which was followed by the second attack due to endotoxemia, in the development of ALI in mice. Experiments revealed that LPS injection in mice with acute pancreatitis caused the development of ALI, as indicated by blood-gas derangements, pulmonary vascular hyperpermeability, increased inflammatory cell counts in bronchoalveolar lavage, and histologic lung damage. This was associated with the pancreatitis-induced increase in expression of macrophage migration inhibitory factor (MIF) in the lungs, together with elevated expression of Toll-like receptor (TLR)-4, both of which were inhibited by administration of anti-protease-activated receptor (PAR)-2 antibody. Furthermore, anti-MIF antibody treatment suppressed the pancreatitis-induced elevation of TLR-4 pulmonary expression. Genetic removal of MIF from mice resulted in less development of ALI in the setting of acute pancreatitis complicated by endotoxemia. These findings demonstrate that activation of protease-activated receptor-2 with trypsin, which can be released after pancreatitis induction, positively regulates the transcript level of MIF, and increased MIF results in exaggerated pulmonary expression of TLR-4, leading to the development of ALI with a subsequent infectious attack. We thus suggest that interventions designed to modulate MIF may have therapeutic advantages in treating ALI in patients with acute pancreatitis complicated by bacterial infection.

Topics: Acute Disease; Animals; Bronchoalveolar Lavage Fluid; Ceruletide; Endotoxemia; Lipopolysaccharides; Lung; Lung Injury; Macrophage Migration-Inhibitory Factors; Male; Mice; Mice, Inbred BALB C; Pancreatitis; Receptors, Proteinase-Activated; Toll-Like Receptor 4

Bacterial infection of the pancreas aggravates the course of acute pancreatitis. Since bacterial translocation from the gut is likely to be an early event, in an animal model of pancreatitis, we investigated the effect of early bacterial supra-infection of the pancreas on the course of the disease.. Six hours after the induction of acute pancreatitis in male Wistar rats (n = 180) by supramaximal stimulation with cerulein (or placebo in a control group), the animals were operated and a suspension of Helicobacter pylori, Escherichia coli or saline were introduced either in the pancreatic duct or interstitium (12 groups of 15 rats each); after 24 h, animals were killed and the following parameters analysed: macroscopic and histologic appearance of the pancreas (score), wet-to-dry weight ratio, pancreas trypsinogen activation peptide level, serum amylase, interleukin-6 and phospholipase A2 activity.. All parameters were increased in rats with cerulein-induced pancreatitis in comparison to placebo. Interstitial and intraductal application of bacteria increased the pancreatic damage. This effect was more evident with the application of E. coli in both cerulein and placebo groups. Application of E. coli but not of H. pylori determined pancreatic activation of trypsinogen, increased mortality and induced the production of interleukin-6.. Bacterial invasion of the pancreas worsens the histologic and clinical picture of disease and induces a systemic inflammatory response.

Topics: Acute Disease; Amylases; Animals; Bacterial Infections; Ceruletide; Disease Models, Animal; Interleukin-6; Male; Organ Size; Pancreas; Pancreatitis; Pancreatitis, Acute Necrotizing; Phospholipases A; Phospholipases A2; Rats; Rats, Wistar; Systemic Inflammatory Response Syndrome

To investigate the effects of hyperlipidemia on acute pancreatitis (AP) and the possible mechanisms.. Rat models of hyperlipidemia and AP were established by Triton WR1339 and cerulein respectively. Human albumin was used to treat AP complicated by hyperlipidemia. In each group, we compared the histological score, volume of ascites, ratio of pancreatic wet/dry weight, serum amylase (AMY) and pancreatic acinar cell apoptosis. The level of protein kinase C (PKC) membrane translocation in pancreatic tissue was detected by Western blot.. In the hyperlipidemia model established by Triton WR1339, triglyceride (TG) increased remarkably and reached its peak 6 h after injection, and most rats developed mild acute pancreatitis. Histological score, volume of ascites, ratio of wet/dry weight and serum AMY in AP animals with hyperlipidemia were obviously higher than those in AP animals (P < 0.05) and decreased after albumin therapy but not significantly (P > 0.05). Apoptotic cells detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) increased in AP animals with hyperlipidemia and did not change distinctly after albumin therapy. PKC membrane translocation level increased in AP animals with hyperlipidemia and decreased remarkably after albumin therapy (P < 0.05).. Hyperlipidemia may induce AP or intensify pancreatic injury. Albumin therapy can not alleviate pancreatic lesion effectively. PKC activation may be one mechanism by which AP is intensified by hyperlipidemia.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Ascites; Ceruletide; Enzyme Activation; Hyperlipidemias; In Situ Nick-End Labeling; Lipids; Male; Organ Size; Pancreas; Pancreatitis; Polyethylene Glycols; Protein Kinase C; Rats; Rats, Sprague-Dawley

The peroxisome proliferator-activated receptor-alpha (PPAR-alpha) is a member of the nuclear receptor superfamily of ligand-dependent transcription factors related to retinoid, steroid and thyroid hormone receptors. The aim of the present study was to examine the effects of endogenous PPAR-alpha ligand on the development of acute pancreatitis caused by cerulein in mice. Intraperitoneal injection of cerulein into PPAR-alpha wild-type (WT) mice resulted in severe, acute pancreatitis characterized by oedema, neutrophil infiltration and necrosis and by elevated serum levels of amylase and lipase. Infiltration of pancreatic and lung tissue with neutrophils (measured as an increase in myeloperoxidase activity) was associated with enhanced expression of the adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and P-selectin. Immunohistochemical examination demonstrated a marked increase in the staining (immunoreactivity) for transforming growth factor-beta (TGF-beta) and vascular endothelial growth factor (VEGF) in the pancreas of cerulein-treated PPAR-alpha wild-type (WT) mice in comparison to sham-treated mice. Acute pancreatitis in PPAR-alphaWT mice was also associated with a significant mortality (20% survival at 5 days after cerulein administration). In contrast, the degree of pancreatic inflammation and tissue injury (histological score), up-regulation/formation of ICAM-1 and P-selectin, infiltration of neutrophils, and the expression of TGF-beta and VEGF was markedly enhanced in pancreatic tissue obtained from cerulein-treated PPAR-alpha knockout (KO) mice. Thus, endogenous PPAR-alpha ligands reduce the degree of pancreas injury caused by acute pancreatitis induced by cerulein administration.

Topics: Acute Disease; Amylases; Animals; Biomarkers; Blotting, Western; Ceruletide; Intercellular Adhesion Molecule-1; Islets of Langerhans; Lipase; Mice; Mice, Knockout; Neutrophil Infiltration; P-Selectin; Pancreatitis; Peroxidase; PPAR alpha; Random Allocation; Survival Rate; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

The recruitment of inflammatory cells contributes significantly to tissue injury in acute pancreatitis. This process implies several molecular interactions between circulating and endothelial cells. The adhesion molecule junctional adhesion molecule C (JAM-C) is involved in leukocyte transendothelial migration and it can form homophilic (JAM-C/JAM-C) and heterophilic interactions with the leukocyte integrin alpha(M)beta(2). In this study, the effect of early administration of monoclonal antibodies directed against JAM-C in cerulein-induced acute pancreatitis was assessed. This reagent significantly blocked influx of leukocytes, release of serum amylase, secretion of inflammatory cytokines, and acinar cell necrosis. These effects were rapid and protected against tissue injury throughout the duration of the model. Conversely, cerulein-induced acute pancreatitis was more severe in transgenic mice overexpressing JAM-C on endothelial cells under the control of the Tie2 promoter. It is proposed that JAM-C expressed by endothelial cells contributes to the pathophysiology of acute pancreatitis and could be considered a target for clinical applications.

Topics: Acute Disease; Amylases; Animals; Antibodies, Monoclonal; Blotting, Western; Cell Adhesion Molecules; Ceruletide; Chemotaxis, Leukocyte; Edema; Endothelial Cells; Immunoglobulins; Immunohistochemistry; Interleukin-6; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Models, Animal; Necrosis; Pancreas; Pancreatitis

Primary sensory neurons of the C and Adelta subtypes express the vanilloid capsaicin receptor TRPV1 and contain proinflammatory peptides such as substance P (SP) that mediate neurogenic inflammation. Pancreatic injury stimulates these neurons causing the release of SP in the pancreas resulting in pancreatic edema and neutrophil infiltration that contributes to pancreatitis. Axons of primary sensory neurons innervating the pancreas course through the celiac ganglion. We hypothesized that disruption of the celiac ganglion by surgical excision or inhibition of C and Adelta fibers through blockade of TRPV1 would reduce the severity of experimental pancreatitis by inhibiting neurogenic inflammation. Resiniferatoxin (RTX) is a specific TRPV1 agonist that, in high doses, selectively destroys C and Adelta fibers. Sprague-Dawley rats underwent surgical ganglionectomy or application of 10 microg RTX (vs. vehicle alone) to the celiac ganglion. One week later, pancreatitis was induced by six hourly intraperitoneal injections of caerulein (50 microg/kg). The severity of pancreatitis was assessed by serum amylase, pancreatic edema, and pancreatic myeloperoxidase (MPO) activity. SP receptor (neurokinin-1 receptor, NK-1R) internalization in acinar cells, used as an index of endogenous SP release, was assessed by immunocytochemical quantification of NK-1R endocytosis. Caerulein administration caused significant increases in pancreatic edema, serum amylase, MPO activity, and NK-1R internalization. RTX treatment and ganglionectomy significantly reduced pancreatic edema by 46% (P < 0.001) and NK-1R internalization by 80% and 51% (P < 0.001 and P < 0.05, respectively). RTX administration also significantly reduced MPO activity by 47% (P < 0.05). Neither treatment affected serum amylase, consistent with a direct effect of caerulein. These results demonstrate that disruption of or local application of RTX to the celiac ganglion inhibits SP release in the pancreas and reduces the severity of acute secretagogue-induced pancreatitis. It is possible that selectively disrupting TRPV1-bearing neurons could be used to reduce pancreatitis severity.

Topics: Acute Disease; Animals; Ceruletide; Denervation; Ganglia, Sympathetic; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Substance P

Acute pancreatitis is a disease involving pro-inflammatory mediators. Two complex and multifactorial pathogenetic ways lead to edematous or necrotizing pancreatitis. The course of the disease is thought to be the consequence of an acute inflammatory response.. The authors examined the impact of Escherichia coli LPS on the acute cerulein pancreatitis in rats.. The study was performed on rats using the ceruleine pancreatitis model. The activation status of polymorphonuclear cells, blood IL-6 concentration, oxidative stress parameters, pancreatic enzymes concentration and microscopic alterations were determined at 5th and 9th h observations.. In acute pancreatitis and acute pancreatitis with LPS groups, the peripheral polymorphonuclear cells activity was lower than in control one. Authors noticed the same neutrophil activation in acute pancreatitis after lipopolysaccharide administration although the peripheral blood polymorphonuclear cells count was significantly higher at the 9th h observation. LPS neither changed the oxidative stress within pancreatic gland, nor amylase or serum lipase activity. LPS given to acute pancreatitis animals resulted in significant increase of serum IL-6 concentration at 5th observation turning normal after 9th h.. Collected data supports thesis of early polymorphonuclear cells involvement in acute pancreatitis and oxidative stress evidence in pancreatic parenchyma. However, results did not reveal that administration of LPS amplified inflammatory response during the course of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Interleukin-6; Leukocyte Count; Lipase; Lipopolysaccharides; Male; Neutrophil Activation; Neutrophils; Oxidative Stress; Pancreas; Pancreatitis; Rats; Rats, Wistar; Shock, Septic

Acute pancreatitis (AP) is an inflammatory disease involving the production of different cytokines and chemokines and is characterized by leukocyte infiltration. Because the chemokine receptor CCR5 and its ligands [the CC chemokines CCL3/MIP-1alpha, CCL4/MIP-1beta, and CCL5/regulated upon activation, normal T cell expressed and secreted (RANTES)] regulate leukocyte chemotaxis and activation, we investigated the expression of CCR5 ligands and the role of CCR5 and its ligands in experimental AP in mice. AP was induced by hourly intraperitoneal injections of cerulein in CCR5-deficient (CCR5(-/-)) or wild-type (WT) mice. Induction of AP by cerulein resulted in an early increase of pancreatic CCL2, CCL3, and CCL4 mRNA expression, whereas CCL5 mRNA expression occurred later. CCR5(-/-) mice developed a more severe pancreatic injury than WT mice during cerulein-induced AP, as assessed by a more pronounced increase in serum amylase and lipase levels and by more severe pancreatic edema, inflammatory infiltrates (mainly neutrophils), and necrosis. CCR5(-/-) mice also exhibited increased production of CCL2/MCP-1, CCL3/MIP-1alpha, and CCL4/MIP-1beta during the course of cerulein-induced AP. In vivo simultaneous neutralization of CC chemokines with monoclonal antibodies in CCR5(-/-) mice reduced the severity of cerulein-induced AP, indicating a role of CC chemokines in exacerbating the course of AP in the absence of CCR5. Moreover, simultaneous neutralization of CCR5 ligands in WT mice also reduced the severity of cerulein-induced AP. In conclusion, lack of the chemokine receptor CCR5 exacerbates experimental cerulein-induced AP and leads to increased levels of CC chemokines and a more pronounced pancreatic inflammatory infiltrate, suggesting that CCR5 expression can modulate severity of AP.

Topics: Acute Disease; Animals; Ceruletide; Female; Mice; Mice, Knockout; Pancreatitis; Receptors, CCR5

Poly (ADP-ribose) polymerase (PARP), a nuclear enzyme activated by strand breaks in DNA, plays an important role in the colon injury associated with experimental colitis. The aim of the present study was to examine the effects of 3-aminobenzamide (3-AB), an inhibitor of PARP activity, in the development of acute pancreatitis caused by cerulein in mice. Intraperitoneal injection of cerulein in mice resulted in severe, acute pancreatitis characterized by oedema, neutrophil infiltration and necrosis and elevated serum levels of amylase and lipase. Infiltration of pancreatic and lung tissue with neutrophils (measured as increase in myeloperoxidase activity) was associated with enhanced expression of the intercellular adhesion molecule-1 (ICAM-1) and P-selectin. Immunohistochemical examination demonstrated a marked increase in the staining (immunoreactivity) for transforming growth factor-beta (TGF-beta) and vascular endothelial growth factor (VEGF) in the pancreas of cerulein-treated mice in comparison to sham-treated mice. Acute pancreatitis in vehicle-treated mice was also associated with a significant mortality (40% survival at 5 days after cerulein administration). In contrast, (1) the degree of pancreatic inflammation and tissue injury (histological score), (2) upregulation/formation of ICAM-1 and P-selectin, (4) neutrophils infiltration and (5) the expression of TGF-beta and VEGF was markedly reduced in pancreatic tissue obtained from cerulein-treated mice which have been treated with 3-AB. These findings provide the evidence that PARP inhibition reduce the degree of pancreas injury caused by acute pancreatitis induced by cerulein administration.

Topics: Acute Disease; Animals; Benzamides; Ceruletide; Disease Models, Animal; Enzyme Inhibitors; Immunohistochemistry; Intercellular Adhesion Molecule-1; Male; Mice; Neutrophil Infiltration; P-Selectin; Pancreas; Pancreatitis; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Survival Rate; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

Recent studies have shown that stimulation of cannabinoid 1 (CB1) receptor reduces the area of ischemic myocardial necrosis and affects activity of the digestive tract. The aim of the present study was to check whether the administration of CB1 receptor agonist or antagonist affects the stress-induced gastric ulceration and development of edematous pancreatitis.. Experiments were performed on rats. Gastric lesions were induced by water immersion and restrain stress (WRS). Acute pancreatitis was induced by cerulein. Prior to WRS or before and during cerulein administration, a natural endogenous ligand for CB1 receptor, anandamide was administered intraperitoneally at the dose of 0.8, 1.5 or 3.0 micromol/kg. A synthetic CB1 receptor antagonist, AM 251 (ALEXIS(R) Biochemicals) was administrated at the dose of 4 micromol/kg i.p. alone or in combination with anandamide at the dose of 1.5 micromol/kg.. Administration of anandamide reduced gastric lesions and this effect was associated with am increase in gastric mucosal blood flow and mucosal DNA synthesis; whereas serum level of pro-inflammatory interleukin-1 beta was reduced. Treatment with AM 251 aggravated gastric damage and reversed protective effect of anandamide administration. Opposite effect was observed in the pancreas. Administration of anandamide increased dose-dependently the severity of pancreatitis. In histological examination, we observed an increase in pancreatic edema and inflammatory infiltration. Also, treatment with anandamide augmented the pancreatitis-induced increase in serum level of lipase, amylase, poly-C ribonuclease, and pro-inflammatory interleukin-1 beta; whereas pancreatic DNA synthesis was reduced. Treatment with AM 251 reduced histological and biochemical signs of pancreatic damage and reversed deleterious effect of anandamide in cerulein-induced acute pancreatitis.. Activation of CB1 receptors evokes opposite effects in the stomach and pancreas: in the stomach, exhibits protective effect against stress-induced gastric mucosal lesions; whereas in the pancreas, increases the severity of cerulein-induced pancreatitis.

Topics: Acute Disease; Animals; Arachidonic Acids; Cannabinoids; Ceruletide; DNA; Endocannabinoids; Gastric Mucosa; Interleukin-1beta; Male; Pancreas; Pancreatitis; Piperidines; Polyunsaturated Alkamides; Pyrazoles; Rats; Rats, Wistar; Receptor, Cannabinoid, CB1; Restraint, Physical; Stomach Ulcer; Stress, Physiological

Microcirculatory disturbances are important early pathophysiological events in various organs during acute pancreatitis (AP). The aim of the study was to investigate an influence of L-arginine (nitric oxide substrate) and N(G)-nitro-L-arginine (L-NNA, nitric oxide synthase inhibitor) on organ microcirculation in experimental acute pancreatitis induced by four consecutive intraperitoneal cerulein injections (15 microg/kg/h). The microcirculation of pancreas, liver, kidney, stomach, colon and skeletal muscle was measured by laser Doppler flowmeter. Serum interleukin 6 and hematocrit levels were analyzed. AP resulted in a significant drop of microperfusion in all examined organ. L-arginine administration (2 x 100 mg/kg) improved the microcirculation in the pancreas, liver, kidney, colon and skeletal muscle, and lowered hematocrit levels. L-NNA treatment (2 x 25 mg/kg) caused aggravation of edematous AP to the necrotizing situation, and increased IL-6 and hematocrit levels. A further reduction of blood perfusion was noted in the stomach only. It is concluded that L-arginine administration has a positive influence on organ microcirculatory disturbances accompanying experimental cerulein-induced AP. NO inhibition aggravates the course of pancreatitis.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Enzyme Inhibitors; Interleukin-6; Male; Microcirculation; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pancreas; Pancreatitis; Rats; Rats, Wistar; Regional Blood Flow

Decreased levels of expression of opsonin receptors (CD11b and CD32/16) on peritoneal exudate neutrophils may lead to susceptibility to infection. Granulocyte colony-stimulating factor (G-CSF) increases the expression levels of CD11b on neutrophils and prolongs neutrophil survival. The effects of G-CSF on neutrophils and opsonin receptor expressions of neutrophils were investigated in cerulein-induced acute pancreatitis.. Forty-two mice were randomly assigned to each group (n = 6). Mice received subcutaneous G-CSF (120 microg/kg body weight) before the induction of acute pancreatitis with cerulein. Saline was used for instead of G-CSF or cerulein solution in control groups. CD11b and CD32/16 expression levels on circulatory and peritoneal exudate neutrophils were investigated 6 and 24 hours after the induction of acute pancreatitis.. Treatment with G-CSF did not aggravate the inflammation of pancreatic tissue evaluated by plasma amylase, acinar necrosis. However, it significantly increased the number of peritoneal exudate neutrophils (P < 0.05) and the CD11b- (P < 0.05) and CD32/16-positive (P < 0.05) peritoneal exudate neutrophils in mice with cerulein-induced acute pancreatitis. The means of fluorescence intensity for CD11b and CD32/16 expressions on circulatory and peritoneal exudate neutrophils were also elevated in the G-CSF groups.. G-CSF administration increases the numbers of neutrophils and improves expression levels of opsonin receptors on neutrophils in mice with cerulein-induced acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; CD11b Antigen; Ceruletide; Female; Granulocyte Colony-Stimulating Factor; Leukocyte Count; Mice; Mice, Inbred BALB C; Neutrophils; Pancreatitis; Receptors, IgG; Receptors, Immunologic

VIP receptor has been clarified to exist on immune cells, indicating its possible involvement in immunity and inflammatory response. Therefore, we investigated the effects of VIP and selective agonists for 2 subtypes of VIP receptor (VPAC1-R and VPAC2-R agonist) on acute pancreatitis.. Acute pancreatitis was induced in mice by 4 intraperitoneal injections of cerulein and an injection of LPS. VIP, VPAC1-R agonist, VPAC2-R agonist, or secretin (5 nmol/body) was administered 30 minutes before and after the administration of LPS. Serum amylase and cytokine levels were determined, and histologic changes were evaluated. In vitro, IL-6 and TNF-alpha production by monocytes from the spleen was determined under the stimulation of LPS with VIP, VPAC1-R agonist, or VPAC2-R agonist, and the expression of VPAC1-R and VPAC2-R mRNA in monocytes was examined.. VPAC1-R agonist significantly decreased serum amylase, IL-6, and TNF-alpha, whereas VPAC2-R agonist markedly increased serum amylase. Histologically, VIP and VPAC1-R agonist attenuated the severity of pancreatitis, although VPAC2-R agonist or secretin showed no significant effect. In vitro, VPAC1-R and VPAC2-R mRNA were obviously expressed in monocytes. Under the stimulation with LPS, VIP presented a biphasic pattern that once decreased IL-6 production from monocytes and then enhanced at high concentration. VPAC1-R agonist reduced IL-6 levels, whereas VPAC2-R agonist increased IL-6 dose-dependently. VPAC1-R agonist reduced TNF-alpha levels in a dose-dependent manner.. VIP attenuated the experimental acute pancreatitis enzymatically and morphologically by inhibiting proinflammatory cytokine production from monocytes mainly through the VPAC1-R.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cytokines; Interleukin-10; Interleukin-6; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Monocytes; Pancreas; Pancreatitis; Receptors, Vasoactive Intestinal Peptide, Type II; Receptors, Vasoactive Intestinal Polypeptide, Type I; RNA, Messenger; Severity of Illness Index; Spleen; Tumor Necrosis Factor-alpha; Vasoactive Intestinal Peptide

Pituitary adenylate cyclase activating-peptide (PACAP) is a late member of the secretin/glucagon/vasoactive intestinal peptide (VIP) family of brain-gut peptides. It is unknown whether PACAP takes part in the development of acute pancreatitis and whether PACAP or its antagonists can be used to suppress the progression of acute pancreatitis. We investigated the actions of PACAP and its receptor antagonists in acute pancreatitis on rats.. Acute pancreatitis was induced in rats with caerulein or 3.5% sodium taurocholate. The rats were continuously infused with 5-30 microg/kg PACAP via jugular vein within the first 90 min, while 10-100 microg/kg PACAP6-27 and (4-Cl-D-Phe6, Leu17) VIP (PACAP receptor antagonists) were intravenously infused for 1 h. Biochemical and histopathological assessments were made at 4 h after infusion. Pancreatic and duodenal PACAP concentrations were determined by enzyme-linked immunosorbent assay (ELISA). Chinese ink-perfused pancreas was fixed, sectioned and cleared for counting the functional capillary density.. PACAP augmented caerulein-induced pancreatitis and failed to ameliorate sodium taurocholate-induced pancreatitis. ELISA revealed that relative concentrations of PACAP in pancreas and duodenum were significantly increased in both sodium taurocholate- and caerulein-induced pancreatitis compared with those in normal controls. Unexpectedly, PACAP6-27 and (4-Cl-D-Phe6, Leu17) VIP could induce mild acute pancreatitis and aggravate caerulein-induced pancreatitis with characteristic manifestations of acute hemorrhagic/necrotizing pancreatitis. Functional capillary density of pancreas was interpreted in the context of pancreatic edema, and calibrated functional capillary density (calibrated FCD), which combined measurement of functional capillary density with dry weight/wet weight ratio, was introduced. Hyperemia or congestion, rather than ischemia, characterized pancreatic microcirculatory changes in acute pancreatitis.. PACAP may take part in the pathogenesis of acute pancreatitis in rats. The two PACAP receptor antagonsits might act as partial agonists. Calibrated functional capillary density can reflect pancreatic microcirculatory changes in acute pancreatitis.

Topics: Acute Disease; Animals; Capillaries; Ceruletide; Cholagogues and Choleretics; Disease Models, Animal; Duodenum; Hormone Antagonists; Male; Nerve Growth Factors; Neuropeptides; Neurotransmitter Agents; Pancreas, Exocrine; Pancreatitis; Peptide Fragments; Pituitary Adenylate Cyclase-Activating Polypeptide; Rats; Rats, Wistar; Receptors, Cell Surface; Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; Taurocholic Acid; Vasoactive Intestinal Peptide

To investigate the changes of platelet endothelial cell adhesion molecule-1 (PECAM-1) expression on polymorphonuclear leukocytes (PMNs) in peripheral circulation and pancreatic microcirculation in cerulein-induced acute edematous pancreatitis (AEP).. Fifty Wistar rats were randomly divided into control group (n = 10) and AEP group (n = 40). A model of AEP was established by subcutaneous injection of cerulein 5.5 and 7.5 mug/kg at 0 and 1 h after the beginning of experiment respectively. PECAM-1 expression on PMNs from splenic vein and inferior vena cava was determined by RT-PCR at mRNA level and determined by flow cytometry at protein level.. In experimental rats, an increased PECAM-1 mRNA expression was seen from 4 to 8 h of AEP in peripheral circulation (0.77+/-0.25%, 0.76+/-0.28%, 0.89+/-0.30%, 1.00+/-0.21%), while in pancreatic microcirculation, expression decreased from 2 h and reached the lowest level at 6 h of AEP (0.78+/-0.29%, 0.75+/-0.26%, 0.62+/-0.28%, 0.66+/-0.20%). There were significant differences at 8-h time point of AEP between peripheral circulation and pancreatic microcirculation (1.00+/-0.21% vs 0.66+/-0.20%, P<0.05). Meanwhile, the difference at protein level was also found.. A reverse expression of PECAM-1 on PMNs was found between peripheral circulation and pancreatic microcirculation, suggesting that inhibition of PECAM-1 expression may improve the pathological change of AEP.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Edema; Flow Cytometry; Gene Expression; Male; Microcirculation; Neutrophils; Organ Size; Pancreas; Pancreatitis; Platelet Endothelial Cell Adhesion Molecule-1; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction

Chemokines are believed to play a key role in the pathogenesis of acute pancreatitis. We have earlier shown that pancreatic acinar cells produce the chemokine monocyte chemotactic protein (MCP)-1 in response to caerulein hyperstimulation, demonstrating that acinar-derived MCP-1 is an early mediator of inflammation in acute pancreatitis. Blocking chemokine production or action is a major target for pharmacological intervention in a variety of inflammatory diseases, such as acute pancreatitis. 2-Methyl-2-[[1-(phenylmethyl)-1H-indazol-3yl]methoxy]propanoic acid (bindarit) has been shown to preferentially inhibit MCP-1 production in vitro in monocytes and in vivo without affecting the production of the cytokines IL-1, IL-6, or the chemokines IL-8, protein macrophage inflammatory-1alpha, and RANTES. The present study aimed to define the role of MCP-1 in acute pancreatitis with the use of bindarit. In a model of acute pancreatitis induced by caerulein hyperstimulation, prophylactic as well as therapeutic treatment with bindarit significantly reduced MCP-1 levels in the pancreas. Also, this treatment significantly protected mice against acute pancreatitis as evident by attenuated hyperamylasemia neutrophil sequestration in the pancreas (pancreatic MPO activity), and pancreatic acinar cell injury/necrosis on histological examination of pancreas sections.

Topics: Acute Disease; Animals; Ceruletide; Chemokine CCL2; Disease Models, Animal; Indazoles; Inflammation; Mice; Necrosis; Pancreas; Pancreatitis; Propionates

To assess the effect of non-selective ET(A/B) (LU 302872) and selective ET(A) (LU 302146) antagonist on pancreatic histology and ultrastructure of acinar cells in connection with trypsinogen activation in early caerulein-induced AP.. Male Wistar rats with caerulein-induced AP, lasting 4 h, were treated i.p. with 10 and 20 mg/kg b.w. of each antagonist. Edema, inflammatory infiltration, necrosis and vacuolization of acinar cells in the pancreas were scored at 0-3 scale. Free active trypsin (FAT), total potential trypsin (TPT) after activation with enterokinase, and index of trypsinogen activation (%FAT/TPT) were assayed in pancreatic homogenates.. In untreated AP, the edema, inflammatory infiltration, necrosis and vacuolization increased as compared to control healthy rats (P<0.01). None of the treatment exerted any meaningful effect on the edema and inflammatory infiltration. The selective antagonist increased slightly the necrosis score to 0.82+/-0.06 at higher dose (P<0.05) vs 0.58+/-0.06 in untreated AP. The non-selective antagonist increased slightly the vacuolization score to 2.41+/-0.07 at higher dose (P<0.01) vs 1.88+/-0.08 in untreated AP. The decrease in the number of zymogen granules, disorganization of endoplasmic reticulum, autophagosomes and cytoplasmic vacuoles were more prominent in treated AP than in untreated AP groups. %FAT/TPT in untreated AP increased about four times (18.4+/-3.8 vs 4.8+/-1.3 in control group without AP, P<0.001). Treatment of AP with both antagonists did not affect significantly augmented trypsinogen activation.. The treatment with endothelin-1 receptors (non-selective ET(A/B) and selective ET(A)) antagonists has essential effect neither on the edema and inflammatory infiltration nor on trypsinogen activation observed in the early course of caerulein-induced AP. Nevertheless a slight increase of the necrosis and vacuolization score and some of the ultrastructural data could suggest the possibility of their undesired effects in caerulein-induced AP at investigated doses.

Topics: Acute Disease; Animals; Benzhydryl Compounds; Ceruletide; Endothelin A Receptor Antagonists; Male; Microscopy, Electron; Pancreas; Pancreatitis; Propionates; Pyrimidines; Rats; Rats, Wistar; Trypsinogen

Uncoupling-protein 2 (UCP2) is a mitochondrial protein that appears to be involved in cellular oxidant defense and in the regulation of oncotic cell death, both of which are important features of acute pancreatitis. However, UCP2 expression in acute pancreatitis has not been previously reported. In the current experiments, pancreatic gene expression was studied by real-time reverse-transcription/polymerase chain reaction and Northern blots. Two models of acute experimental pancreatitis were investigated: cerulein-induced pancreatitis in mice at two different time points and taurocholate-induced pancreatitis in rats at two degrees of severity. After cerulein administration, acinar injury and leukocyte infiltration was significantly higher at 24 h compared with 12 h after the first injection of cerulein (P<0.05, P<0.005, respectively). UCP2 mRNA was unchanged at 12 h but was nearly 12-fold greater than control levels after 24 h (P<0.001). UCP2 gene expression correlated with acinar injury (r=0.69; P<0.001). By 72 h after taurocholate administration, the severe group had more necrosis than the mild group (P<0.005). Pancreatic UCP2 mRNA was increased fourfold in the severe group compared with controls (P<0.01). UCP2 expression correlated with parenchymal necrosis (r=0.61; P<0.01). Thus, pancreatic UCP2 mRNA increased in two models of acute pancreatitis. The increase in UCP2 gene expression was correlated with the severity of the disease. Up-regulation of UCP2 in the pancreas may be a protective response to oxidative stress, but this increase may also have a negative influence on cellular energy metabolism. Therefore, acinar UCP2 may be an important modifier of the severity of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Female; Gene Expression Regulation; Ion Channels; Male; Membrane Transport Proteins; Mice; Mice, Inbred C57BL; Mitochondrial Proteins; Necrosis; Pancreatitis; Rats; Rats, Zucker; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Taurocholic Acid; Time Factors; Uncoupling Protein 2; Up-Regulation

To detect Toll-like receptor 4 (TLR4) expression and distribution in rat pancreas and the change of TLR4 expression in cerulein-induced pancreatitis (CIP).. Acute pancreatitis was induced by subcutaneous injections of cerulein at a total dose of 20 microg/kg. Immunohistochemistry (IHC) was used to detect and localize TLR4 in rat pancreas, and real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to quantitatively determine the expression of TLR4 mRNA in CIP.. IHC showed the presence of TLR4 in rat pancreas, and its distribution was specifically localized to pancreatic ductal epithelium, vascular endothelium, and islet. No TLR4 staining was detected in exocrine acinar cells. Real-time RT-PCR results revealed low-level TLR4 mRNA expression in the rat pancreas, and the change of TLR4 in CIP only developed within the first 4 hours, which is a rapid up-regulation process that peaks at the first hour. TLR4 mRNA was sustained at baseline level from 4 to 24 hours.. TLR4 protein was expressed in pancreas and localized to epithelial (pancreatic duct) or endothelial (vessels) tissues; TLR4 responded favorably to the inflammatory process, and the change of expression was characterized as a rapid up-regulation in the early stage of CIP.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cytokines; Endothelium; Female; Gene Expression; Immunohistochemistry; Lipase; Male; Pancreas, Exocrine; Pancreatic Ducts; Pancreatitis; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Toll-Like Receptor 4; Up-Regulation

Acute pancreatitis is an auto-digestive disease resulting in inflammation. At the cellular level, acute pancreatitis disrupts posttranslational protein processing and traffic in the secretory pathway, and zymogens become activated in the acinar cell. To better understand the disruption of the secretory pathway in pancreatitis, pulse-chase [(35)S]met/cys analysis was used to study the effects of supramaximal cerulein stimulation on posttranslational modification in the secretory pathway of the major sulfated glycoprotein of the mouse pancreas, pro-Muclin, and the lysosomal membrane protein LAMP1. Maximal cerulein or high concentration bombesin stimulation had little effect on glycoprotein processing. By contrast, supramaximal cerulein stimulation strongly inhibited pro-Muclin processing as measured by the failure of Muclin to attain its normal mature size of 300 kDa and to become highly sulfated and decreased proteolytic cleavage of pro-Muclin to produce apactin. Digestion of immunoprecipitated [35S]met/cys-labeled Muclin and LAMP1 with endoglycosidase H demonstrated that the supramaximal cerulein-induced block in processing occurred before the medial Golgi compartment. With supramaximal cerulein stimulation, vacuoles formed which contained Muclin, amylase, and LAMP1. Earlier autoradiographic studies showed that newly synthesized proteins end up in pancreatitis-associated vacuoles, so it is likely that glycoproteins with incomplete posttranslational processing are also present in vacuoles. Because glycoproteins are believed to protect the membranes of lysosomes and zymogen granules, when they are not correctly processed, their defensive mechanisms may be impaired, and this could contribute to vacuole fragility in pancreatitis.

Topics: Acute Disease; Animals; Bombesin; Ceruletide; Disease Models, Animal; Glycoproteins; Golgi Apparatus; Lysosomal Membrane Proteins; Membrane Glycoproteins; Mice; Mucins; Pancreas; Pancreatitis; Protein Processing, Post-Translational; Time Factors

To observe the expressions of early growth response factor-1 (Egr-1) and tissue factor (TF) in rats with cerulein-induced acute pancreatitis and to explore its significance.. A large dose of cerulein was used to create the experimental acute pancreatitis model in rats. The changes of Egr-1 mRNA and protein in rats were observed during 30 min to 4 h after the treatment and immunohistochemical method was used to observe the localized expression of Egr-1 in tissues. In addition to the mRNA expression of Egr-1 target gene, TF was also observed. A blank control group, and a bombesin-administered group were used for comparison.. After the stimulation of a large dose of cerulein, the rats showed typical inflammatory changes of acute pancreatitis. Thirty minutes after the stimulation, the mRNA expression of Egr-1 in the pancreatic tissue reached its peak and then declined, while the expression of Egr-1 protein reached its peak 2 h after the stimulation. Histologically, 2 h after the stimulation, almost all pancreatic acinar cells had the expression of Egr-1 protein, which was focused in the nuclei. The mRNA expression of TF occurred 1 h after the stimulation and gradually increased within 4 h. However, a large dose of bombesin only stimulated the pancreatic tissue to produce a little mRNA expression of Egr-1 and no mRNA expression of Egr-1 protein and TF.. Egr-1 as a pro-inflammatory transcription factor may play an important role in the pathogenesis of acute pancreatitis by modulating the expression of TF.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; DNA-Binding Proteins; Early Growth Response Protein 1; Gene Expression; Immediate-Early Proteins; Male; Pancreatitis; Rats; Rats, Wistar; Thromboplastin; Transcription Factors

The severity of acute pancreatitis results from the transmigration and activation of leukocytes within the pancreas and the local synthesis and release of proinflammatory-soluble mediators that transform a local injury into a systemic inflammatory response. Poly(ADP-ribose)polymerase-1 (PARP-1) is a nuclear DNA-binding protein that has been shown to play a relevant role in cell necrosis and organ failure in various diseases associated with inflammation. Therefore, we set out to investigate whether the genetic deletion of PARP-1 or PARP-2 (a new member of the PARP family) genes, or pharmacological inhibition of PARP activity might affect the development and severity of acute pancreatitis and pancreatitis-associated lung injury. Secretagogue-induced acute pancreatitis was achieved by 12 hourly intraperitoneal injections of cerulein in mice deficient in PARP-1 or PARP-2 genes, and wild-type (WT) littermate mice untreated or treated with PARP activity inhibitors. The severity of pancreatitis was assessed by measurements of serum amylase, lipase, interleukin-1beta and IL-6, pancreatic water content, histologic grading and pancreas myeloperoxidase (MPO) activity. Lung injury was evaluated by quantifying MPO activity and morphological changes. We found that the severity of acute pancreatitis and pancreatitis-associated lung injury was significantly attenuated in mice lacking PARP-1, but not PARP-2, compared with WT mice. Interestingly, administration of PARP inhibitors, 3-aminobenzamide or PJ34 (N-(6-oxo-5,6-dihydro-phenanthridin-2-yl)-N,N-dimethyacetamide HCl), in WT mice markedly decreased acute pancreatitis severity and pulmonary-associated injury in a larger extension than genetic deletion of PARP-1. Our results support the potential therapeutic application of PARP inhibitors in the development and severity of acute pancreatitis and associated lung injury.

Topics: Acute Disease; Amylases; Animals; Benzamides; Ceruletide; Interleukin-1; Interleukin-6; Lipase; Lung Diseases; Mice; Mice, Knockout; Neutrophils; Pancreatitis; Peroxidase; Phenanthrenes; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases

The effects of R-102444 ((2R, 4R)-4-lauroyloxy-2-[2-[2-[2-(3-methoxy)phenyl]ethyl]phenoxy]ethyl-1-methylpyrrolidine hydrochloride) and its active metabolite R-96544 ((2R, 4R)-2-[2-[2-[2-(3-methoxy)phenyl]ethyl]phenoxy]ethyl-4-hydroxy-1-methylpyrrolidine hydrochloride), potent and selective 5-hydroxytryptamine 2A (5-HT2A) receptor antagonists, on development of pancreatitis were investigated in experimental models of acute and chronic pancreatitis. Rat acute pancreatitis was induced by caerulein (20 microg/kg) intraperitoneal injection and by pancreatic duct ligation. In both the models, serum amylase and lipase activities were markedly increased. R-102444 dose-dependently reduced these enzyme activities at a dose range of 10 to 100 mg/kg (p.o.) for the caerulein model and 0.3 to 10 mg/kg (p.o.) for the ligation model. In a mouse model of acute pancreatitis induced by a choline-deficient, ethionine (0.5%)-supplemented diet, subcutaneous administration of R-96544 (10-100 mg/kg, bid) reduced serum amylase activity. Histological analysis showed that R-96544 dose-dependently attenuated pancreatic necrosis, inflammation and vacuolization. The effect of R-102444 was further examined in male Wistar Bonn/Kobori rats (4-9 months of age) which spontaneously show pancreatic fibrosis and parenchymal destruction compatible with human chronic pancreatitis. In Wistar Bonn/Kobori rats (from 3 to 9 months of age) fed a diet containing 0.017% and 0.17% of R-102444, pancreatic weight, pancreatic protein and amylase content were higher compared to those in non-treated pancreatitis control rats. Histological analysis showed that R-102444 suppressed parenchymal destruction and replacement with adipose tissue, indicating inhibition of pancreatic atrophy. These results clearly indicate that R-102444 and R-96544 inhibit the progression of acute and chronic pancreatitis and support the contention of possible involvement of 5-HT2A receptors in the progression of experimental pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Choline; Chronic Disease; Dietary Supplements; Ethionine; Injections, Intraperitoneal; Ligation; Lipase; Male; Organ Size; Pancreas; Pancreatic Ducts; Pancreatitis; Pyrrolidines; Rats; Rats, Inbred Strains; Rats, Sprague-Dawley; Rats, Wistar; Receptor, Serotonin, 5-HT2A; Serotonin 5-HT2 Receptor Antagonists; Serotonin Antagonists; Time Factors

The aim of this study was to test the effect of gut manipulation by either novel synbiotics or by metronidazole on either endotoxemia or the severity of liver damage in the course of acute pancreatitis from alcohol ingestion.. Sprague-Dawley rats were fed for 1 week through an intragastric tube a liquid diet with either: (i) 1 mL t.i.d. of a mixture of synbiotics (Lactobacillus acidophilus, Lactobacillus helveticus and Bifidobacterium in an enriched medium); (ii) 20 mg/kg t.i.d. metronidazole; or (iii) standard diet. Then, acute pancreatitis was induced by caerulein and when the disease was full-blown, rats were fed an alcohol-rich diet. Synbiotic and metronidazole treatment was given for a further 2 weeks. Transaminase and endotoxemia levels were measured before treatment, after 6 h, after 24 h and 2 weeks later, at the time the rats were killed. Liver samples were obtained for histological analysis.. Synbiotics but not metronidazole improved the acute pancreatitis-induced increase in endotoxemia and transaminase levels. The addition of alcohol worsened these variables to a limited extent in the synbiotic-treated group, while metronidazole had a negative effect on liver damage.. Gut flora pretreatment with synbiotics was able to effectively protect against endotoxin/bacterial translocation, as well as liver damage in the course of acute pancreatitis and concomitant heavy alcohol consumption. The beneficial effect of synbiotics on liver histology seems to be correlated with endotoxemia. Metronidazole did not produce such a beneficial effect; in fact, it further worsened liver damage when alcohol was added to the background of ongoing acute pancreatic inflammation.

Topics: Acute Disease; Animals; Bacterial Translocation; Bifidobacterium; Ceruletide; Disease Models, Animal; Endotoxemia; Endotoxins; Ethanol; Gastrointestinal Tract; Lactobacillus acidophilus; Lactobacillus helveticus; Liver Diseases, Alcoholic; Metronidazole; Pancreatitis; Probiotics; Protective Agents; Rats; Rats, Sprague-Dawley; Transaminases

Effects of dexamethasone and N(G)-nitro-L-arginine methyl ester (L-NAME), the nitric oxide (NO) synthase inhibitor, on caerulein-induced acute pancreatitis were examined in rats. Acute pancreatitis was induced by caerulein (20 mug/kg, s.c.) given repeatedly 2 or 4 times every hour, and serum amylase levels, pancreas weight and myeloperoxidase (MPO) activity were measured 6 h after the first injection of caerulein. Dexamethasone (3 mg/kg) and L-NAME (30 mg/kg) were administered p.o. 30 min before the first injection of caerulein. Caerulein caused moderate or severe pancreatitis, depending on the times of injections, resulting in different degrees of increase in serum amylase levels and pancreas weight, and the marked elevation of MPO activity was observed only after injections of caerulein given 4 times per hour. Both dexamethasone and L-NAME suppressed the severity of pancreatits, yet the effect of L-NAME as compared with dexamethasone was more potent against mild pancreatitis but less potent against severe pancreatitis. These results suggest that caerulein-induced acute pancreatitis shows different responsiveness to L-NAME and dexamethasone, depending on the severity; the former is more effective against pancreatitis with less inflammation, while the latter is more effective against pancreatitis with severe inflammation. It is assumed that endogenous NO may be involved in oedema formation as the early event in the development of acute pancreatitis.

Topics: Acute Disease; Administration, Oral; Amylases; Animals; Anti-Inflammatory Agents; Ceruletide; Dexamethasone; Disease Models, Animal; Enzyme Inhibitors; Injections, Subcutaneous; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Organ Size; Pancreas; Pancreatitis; Peroxidase; Rats; Rats, Wistar; Severity of Illness Index

To assess effects of NF-kappaB activation inhibitor (pyrrolidine dithiocarbamate--PDTC) alone or with endothelins (ET-1, ET-2, ET-3) in early course of cerulein-induced acute pancreatitis (AP) in rats.. After 4 h of AP in Wistar rats, treated with PDTC 10 or 40 mg/kg or with PDTC 10 mg/kg and ET-1, ET-2 or ET-3, 0.5 or 1.0 nmol/kg twice i.p. in 1 h interval, free active trypsin (FAT), total potential trypsin (TPT) and lipase in 12000 x g supernatants of pancreatic homogenates, plasma alpha-amylase and histological changes were assayed. %FAT/TPT was an index of trypsinogen activation.. %FAT/TPT significantly increased to 12.42 +/- 2.14%, lipase to 5.51 +/- 0.84 U/mg protein and alpha-amylase to 28.5 +/- 5.61 U/mL in AP vs 1.96 +/- 0.31%, 1.29 +/- 0.11 U/mg and 5.80 +/- 1.38 U/ml in healthy control. Higher dose PDTC attenuated trypsinogen activation to 3.01 +/- 0.53% and alpha-amylase to 15.3 +/- 1.38. PDTC and ET-1 attenuated %FAT/TPT to 2.55 +/- 0.18% with lower and 2.34 +/- 0.44% with higher dose. ET-3 was less effective than ET-1: 6.76 +/- 0.46% with lower dose. Lower doses of ET-1 and ET-2 with PDTC, diminished lipase activity to 2.60 +/- 0.36 and 2.94 +/- 0.33.. Cumulative attenuation of trypsinogen activation after lower dose of PDTC and ET-1 approximated the effect of higher dose of PDTC. Additional effect of ET-3 was weaker than ET-1, and ET-2 was ineffective in this respect. The combination of this NF-kappaB activation inhibitor and ET-1 could be beneficial in early course of edematous AP by attenuating of trypsinogen activation. However, it should be treated with caution because of some unfavorable effects on histological scores of pancreatic injury.

Topics: Acute Disease; alpha-Amylases; Animals; Antioxidants; Ceruletide; Endothelin-1; Endothelin-2; Endothelin-3; Enzyme Activation; Lipase; Male; NF-kappa B; Pancreatitis; Pyrrolidines; Rats; Rats, Wistar; Thiocarbamates; Trypsinogen

Acute pancreatitis (AP) is a complex disease that may be linked to acinar cell apoptosis and inadequate acinar cell replacement. Differentiation of acinar cells is regulated by p48, a DNA binding subunit of the transcription factor PTF1, and the Notch signaling pathway. Acinar cell apoptosis is triggered by oxygen deprivation, ie, hypoxia, by activation of hypoxia inducible factor-1alpha (HIF-1alpha). The aim of this study was to characterize by Northern blot analyses expression of HIF-1alpha, HIF-1alpha-inducible genes (GLUT-1, VEGF, p53), p48, and genes involved in the Notch signaling pathway (Notch-1, Dll1, RBP-Jk, HES-1) during cerulein-induced AP in mice. Maximal expression of HIF-1alpha, HIF-1alpha-inducible genes, p48, and Notch signaling genes occurred 8-12 hours after induction of AP. Maximal expression of p53 occurred 12-48 hours after induction of AP. These findings demonstrate that multiple pancreatic genes are activated acutely during AP that support pancreatic cell replenishment, regulation of expression of acinar cell-specific genes, and apoptosis.

Topics: Acute Disease; Animals; Basic Helix-Loop-Helix Transcription Factors; Blotting, Northern; Ceruletide; DNA-Binding Proteins; Female; Gene Expression Regulation; Homeodomain Proteins; Hypoxia-Inducible Factor 1, alpha Subunit; Immunoglobulin J Recombination Signal Sequence-Binding Protein; Membrane Proteins; Mice; Nuclear Proteins; Pancreatitis; Proteins; Receptor, Notch1; Receptors, Cell Surface; Receptors, Notch; RNA, Messenger; Signal Transduction; Time Factors; Transcription Factor HES-1; Transcription Factors

Tumor necrosis factor alpha (TNF-alpha) has a central role in the pathogenesis of acute pancreatitis and related systemic complications. The aim of this study is to investigate the therapeutic effectiveness of monoclonal TNF antibody (infliximab) in acute edematous and severe necrotizing pancreatitis models in rats.. One hundred rats were randomly divided into 10 groups. Acute edematous pancreatitis (AEP) was induced by injection of cerulein 20 microg/kg 4 times subcutaneously at hourly intervals. Severe necrotizing pancreatitis (SNP) was induced by retrograde injection of 3% taurocholate into the common biliopancreatic duct. Infliximab 8 mg/kg was given via intravenous infusion. Serum amylase activity, pancreatic histopathology, myeloperoxidase enzyme activity (MPO), and pulmonary changes were assessed.. Infliximab treatment significantly decreased serum amylase activity (11939 +/- 1914 U/L versus 3458 +/- 915 U/L, P < 0.001) and histopathologic score (4.1 +/- 0.5 versus 1.5 +/- 0.3, P < 0.001) in AEP. It also suppressed neutrophil infiltration and MPO activity of the pancreatic tissue. In SNP, infliximab treatment was found to decrease pathologic score (9.4 +/- 1.2 versus 3.6 +/- 0.8, P < 0.001) and serum amylase activity (20442 +/- 2375 versus 8990 +/- 1730, P < 0.01). It ameliorated both parenchymal and fatty tissue necrosis of the pancreas. Infliximab also alleviated alveolar edema and acute respiratory distress syndrome like pulmonary complications, but the difference was not significant.. Chimeric TNF antibody, infliximab, should be evaluated for treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Antibodies, Monoclonal; Ceruletide; Edema; Infliximab; Male; Necrosis; Pancreas; Pancreatitis; Pancreatitis, Acute Necrotizing; Peroxidase; Pulmonary Edema; Rats; Rats, Wistar; Severity of Illness Index; Taurocholic Acid; Treatment Outcome; Tumor Necrosis Factor-alpha

Oxidative stress involvement in damage to the pancreas in acute pancreatitis (AP) is well documented. However, little is known about oxidative damage occurring in the different subcellular fractions of pancreatic cells. The aim of this study was to ascertain the main targets of oxidative damage inside cells after AP and the role of endogenous nitric oxide (NO) in it.. A model of cerulein-induced AP in rats was used and N-nitro-l-arginine methyl ester (l-NAME) was administered as an NO production inhibitor. After pancreatitis induction, indicative parameters of lipid peroxidation and protein oxidation together with some enzymatic and nonenzymatic endogenous free radical scavengers were assessed in serum and pancreatic subcellular fractions.. In pancreatitic rats, malondialdehyde and protein carbonyl group concentrations were significantly increased (P < 0.05) in serum and some fractions. The increases were higher in l-NAME-treated rats (P < 0.05). Superoxide dismutase and catalase activities were also increased (P < 0.05) but were decreased (P < 0.05) with l-NAME. The alpha-tocopherol concentration diminished (P < 0.05) in serum and all the studied subcellular fractions and the decrease was stronger in l-NAME-treated rats. Our data suggest that microsomes followed by lysosomal + mitochondrial are the fractions most susceptible to oxidative damage in AP. Endogenous NO plays a protective role against oxidative damage to subcellular fractions.

Topics: Acute Disease; alpha-Tocopherol; Animals; Catalase; Ceruletide; Enzyme Inhibitors; Lipid Peroxides; Male; Malondialdehyde; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Pancreas; Pancreatitis; Proteins; Random Allocation; Rats; Rats, Wistar; Subcellular Fractions; Superoxide Dismutase

Supramaximal dosage of the cholecystokinin analog caerulein leads to edematous pancreatitis with subsequent acinar cell destruction predominantly by apoptosis. We have used immunohistochemistry to reveal the expression of the anti-apoptotic protein galectin-3 in pancreatic acinar cells. Galectin-3, which occurs only in duct cells under physiological conditions, is expressed in a subset of acinar cells after the end of a 12-h caerulein infusion, giving rise to a "patchy" staining pattern. During the subsequent period of inflammation and regeneration, galectin-3 expression increases in those acinar cells that undergo apoptosis. By 48 h after the end of caerulein infusion, morphologically normal cells do not contain galectin-3 and participate in regeneration by proliferation. Tubular complexes, being transient structures from degenerative acini, accumulate galectin-3 in the remnants of the epithelium cells. Stimulation with supramaximal dosages of caerulein of the cell line AR4-2J, which is derived from rat pancreatic acinar cells, also results in a marked increase of galectin-3, confirming the in vivo results. We postulate that the high expression of the anti-apoptotic protein galectin-3 regulates the time course of the apoptotic process in pancreatic acinar cells.

Topics: Acute Disease; Amino Acid Sequence; Animals; Apoptosis; Cell Line, Tumor; Ceruletide; Disease Models, Animal; Edema; Fluorescent Antibody Technique, Indirect; Galectin 3; Humans; Male; Molecular Sequence Data; Pancreas; Pancreatic Ducts; Pancreatitis; Rats; Rats, Wistar; Regeneration

It is well established that damage to the outer membrane of cells is a common phenomenon allowing abnormal transmission of substances into the cytosol. Penetration of albumin into acinar cells has been detected in experimental acute pancreatitis, raising the possibility that membrane damage is a very early event, potentially representing the first changes leading to pancreatitis. To determine if direct damage to the cell membrane is a key factor during induction of acute pancreatitis, thus altering the balance of extra- and intracellular substances, fluorescein-dextran was administered with supramaximal doses of caerulein via the jugular vein or by injection directly into the pancreas. This tracer rapidly penetrates into cells. Two patterns of tracer penetration are observed: cytosolic and vesicular/vacuolar. Fluorescein-dextran administered intravenously with caerulein penetrates into the cytosol of acinar cells within 10 min. Strong cytoplasmic fluorescence occurs within 5 min after direct injection. It may be concluded that supramaximal caerulein, administered in vivo, damages the cell membrane of acinar cells, allowing large molecules to enter the cytosol. Thus Ca(2+) and other substances may enter the cells in abnormally high concentrations, initiating the cellular changes characteristic of pancreatitis. The results raise the question whether membrane wounding may play a role in the initiation of human pancreatitis.

Topics: Acute Disease; Animals; Cell Membrane; Cell Membrane Permeability; Ceruletide; Dextrans; Ferritins; Fluoresceins; Fluorescent Dyes; Male; Molecular Weight; Pancreatitis; Rats; Rats, Sprague-Dawley

Recent studies have shown that inosine, a purine nucleoside produced during the breakdown of adenosine, has immunomodulatory and anti-inflammatory properties. The aim of this study was to examine the effects of inosine on the course of acute pancreatitis.. Edematous pancreatitis was induced by the intraperitoneal injection of caerulein (50 micro g/kg), seven times, at 1-h intervals, in male Wistar rats (caerulein pancreatitis). Inosine (100 mg/kg) was administered 30 min before or 1 h after the first injection of caerulein. The effects of inosine on the severity of pancreatitis were assessed by serum amylase, pancreatic edema (wet/dry ratio), myeloperoxidase activity, cytokine-induced neutrophil chemoattractant-1 concentrations, and histological changes.. Prophylactic administration of inosine significantly decreased the elevation of serum amylase, myeloperoxidase activity, and cytokine-induced neutrophil chemoattractant-1 concentrations in the pancreas and the lung. Inosine did not significantly affect edema formation. Histologically, vacuole formation in pancreatic acinar cells, infiltration of inflammatory cells in the pancreas and the lung, and alveolar wall thickening in the lung were reduced. Inosine improved the histological findings and reduced myeloperoxidase activity even if it was administered 1 h after the first injection of caerulein.. Inosine reduced the severity of acute pancreatitis, suggesting a possible application of this compound in the treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Chemokines, CXC; Enzyme-Linked Immunosorbent Assay; Inosine; Intercellular Signaling Peptides and Proteins; Lung; Lung Diseases; Male; Pancreas; Pancreatitis; Peroxidase; Rats; Rats, Wistar

The effect of inhibiting nitric oxide (NO) synthase (NOS) or enhancing NO on the course of acute pancreatitis (AP) is controversial, in part because three NOS isoforms exist: neuronal (nNOS), endothelial (eNOS), and inducible (iNOS). We investigated whether inhibition or selective gene deletion of NOS isoforms modified the initiation phase of caerulein-induced AP in mice and explored whether this affected pancreatic microvascular blood flow (PMBF). We investigated the effects of nonspecific NOS inhibition with N(omega)-nitro-l-arginine (l-NNA; 10 mg/kg ip) or targeted deletion of eNOS, nNOS, or iNOS genes on the initiation phase of caerulein-induced AP in mice using in vivo and in vitro models. Western blot analysis was performed to assess eNOS phosphorylation status, an indicator of enzyme activity, and microsphere studies were used to measure PMBF. l-NNA and eNOS deletion, but not nNOS or iNOS deletion, increased pancreatic trypsin activity and serum lipase during the initiation phase of in vivo caerulein-induced AP. l-NNA and eNOS did not affect trypsin activity in caerulein-hyperstimulated isolated acini, suggesting that nonacinar events mediate the effect of NOS blockade in vivo. The initiation phase of AP in wild-type mice was associated with eNOS Thr(495) residue dephosphorylation, which accompanies eNOS activation, and a 178% increase in PMBF; these effects were absent in eNOS-deleted mice. Thus eNOS is the main isoform influencing the initiation of caerulein-induced AP. eNOS-derived NO exerts a protective effect through actions on nonacinar cell types, most likely endothelial cells, to produce greater PMBF.

Topics: Acute Disease; Animals; Ceruletide; Cytoprotection; Enzyme Inhibitors; Mice; Mice, Inbred C57BL; Mice, Knockout; Microcirculation; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Nitroarginine; Pancreas; Pancreatitis; Phosphorylation; Regional Blood Flow; Trypsin

In the present study, we investigated the effects of rosiglitazone (10 mg/kg, i.p.), a PPAR-gamma agonist, on the development of acute pancreatitis.. Intraperitoneal injection of cerulein in mice induced an acute pancreatitis characterized by edema, neutrophil infiltration elevated serum levels of amylase and lipase. This experimental model was performed to test the anti-inflammatory activity of rosiglitazone. SETTING. University research laboratory.. Male CD mice (20-22 g) were allocated into four groups (n=10 for each group): (a) Cerulein+vehicle group. Mice were treated hourly (x 5) with cerulein (50 microg/kg, in saline solution, i.p.); (b) Rosiglitazone group (same as the Cerulein+vehicle group but were administered rosiglitazone, 10 mg/kg bolus, 30 min prior to cerulein); (c) Sham+saline group. Mice were treated with saline instead of cerulein; (d) Sham+Rosiglitazone. Identical to Rosiglitazone group except that the saline was administered instead of cerulein. Mice were killed at 6 h after the induction of pancreatitis. Blood samples, pancreas, and lungs were collected.. Infiltration of pancreatic and lung tissue with neutrophils was associated with enhanced lipid peroxidation. Immunohistochemical examination demonstrated a marked increase in immunoreactivity for nitrotyrosine and for ICAM-1 in the pancreas of cerulein-treated mice. In contrast, the degree of (a) pancreatic inflammation and tissue injury, (b) upregulation/formation of ICAM-1 and nitrotyrosine, and (c) neutrophils infiltration was markedly reduced in pancreatic tissue obtained from rosiglitazone-treated mice.. These findings support the view that rosiglitazone and other potent PPAR-gamma agonists may be useful in the therapy of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Cell Adhesion Molecules; Ceruletide; Lipase; Lipid Peroxidation; Male; Mice; Pancreatitis; Receptors, Cytoplasmic and Nuclear; Rosiglitazone; Thiazolidinediones; Transcription Factors

Caerulein-induced pancreatitis is a widely used experimental model for studies on acute pancreatitis, however, the molecular mechanisms underlying pancreatitis in response to caerulein hyperstimulation are incompletely understood. We therefore studied early effects of caerulein on tight junctional integrity. Mice were injected with the cholecystokinin analogue caerulein (50microg/kg BW/h) to induce pancreatitis. In pancreatic tissue occludin, claudin 1, zonula occludens protein 1 (ZO-1) were stained immunohistochemically and F-actin was visualized with phalloidin-TRITC. Stained sections and isolated acini were studied by confocal laser scanning microscopy. Under control conditions occludin, claudin1, ZO-1, and F-actin showed a linear staining pattern delineating the apical membranes of intralobular duct cells and of acinar cells. While in vitro caerulein hyperstimulation induced within 10 minutes disassembly of both occludin and ZO-1, in vivo caerulein hyperstimulation induced disassembly of occludin and claudin1 but not of ZO-1 from the tight junctions. Subsequent progressive disruption of ZO-1 was detected in a time dependent manner. Disruption of the transmembrane tight junction proteins occludin and claudin1 is an early event of caerulein hyperstimulation and may allow evasion of noxious luminal content into the interstitium, which may augment edema formation in acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Blotting, Western; Ceruletide; Claudin-1; Immunohistochemistry; Lipase; Male; Membrane Proteins; Mice; Occludin; Pancreas, Exocrine; Pancreatitis; Phosphoproteins; Tight Junctions; Zonula Occludens-1 Protein

To investigate the effects of rhubarb on severe acute pancreatitis (SAP) in rats.. Severe acute pancreatitis was induced by two intraperitoneal injections of cerulein (40 microg/kg body weight) plus 5-h restraint water-immersion stress. Rhubarb (75-150 mg/kg) was orally fed before the first cerulein injection. The degree of pancreatic edema, serum amylase level, local pancreatic blood flow (PBF), and histological alterations were investigated. The effects of rhubarb on pancreatic exocrine secretion in this model were evaluated by comparing with those of somatostatin.. In the Cerulein+Stress group, severe edema and diffuse hemorrhage in the pancreas were observed, the pancreatic wet weight (11.60+/-0.61 g/Kg) and serum amylase (458 490+/-43 100 U/L) were markedly increased (P<0.01 vs control). In the rhubarb (150 mg/kg) treated rats, necrosis and polymorphonuclear neutrophil (PMN) infiltration in the pancreas were significantly reduced (P<0.01), and a marked decrease (50%) in serum amylase levels was also observed (P<0.01). PBF dropped to 38% (93+/-5 mL/min per 100 g) of the control in the Cerulein+Stress group and partly recovered in the Cerulein+Stress+Rhubarb 150 mg group (135+/-12 mL/min per 100 g) (P<0.01). The pancreatic exocrine function was impaired in the SAP rats. The amylase levels of pancreatic juice were reduced in the rats treated with rhubarb or somatostatin, comparing with that of untreated SAP group. The bicarbonate concentration of pancreatic juice was markedly elevated only in the rhubarb-treated group (P<0.01).. Rhubarb can exert protective effects on SAP, probably by inhibiting the inflammation of pancreas, improving pancreatic microcirculation, and altering exocrine secretion.

Topics: Acute Disease; Animals; Ceruletide; Cytoprotection; Male; Pancreatitis; Phytotherapy; Rats; Rats, Sprague-Dawley; Rheum; Severity of Illness Index

Melatonin, produced from L-tryptophan, protects the pancreas against acute damage by improving the antioxidative status of tissue. Melatonin receptors have been detected in the brain, but the contribution of these receptors to the pancreatic protection is unknown. The aim of our study was to compare the effects of melatonin precursor; L-tryptophan given intracerebroventricularly (i.c.v.) or intraperitoneally (i.p.) on the course of acute pancreatitis. Acute pancreatitis was induced by subcutaneous infusion of caerulein (5 microg/kg-h x 5 h). L-tryptophan was given i.p. (2.5, 25 or 250 mg/kg) or administered into right cerebral ventricle (0.02, 0.2 or 2.0 mg/rat) 30 min prior to the start of caerulein infusion. Plasma amylase, lipase and TNF alpha activities were measured to determine the severity of caerulein-induced pancreatitis (CIP). The lipid peroxidation products: malonylodialdehyde and 4-hydroksynonenal (MDA + 4-HNE) and activity of superoxide dismutase (SOD) were measured in the pancreas of intact or CIP rats with or without L-tryptophan pretreatment. Melatonin blood level was measured by RIA. CIP was confirmed by histological examination and manifested as an edema and rises of plasma levels of amylase, lipase and TNF alpha (by 550%, 1000% and 600%). MDA + 4-HNE was increased by 600%, whereas SOD activity was reduced by 75% in the pancreas of CIP rats. All manifestations of CIP were significantly reduced by pretreatment of the rats with L-tryptophan given i.c.v. at doses of 0.2 or 2.0 mg/rat, or by peripheral administration of this amino acid used at dose of 250 mg/kg i.p. In control rats plasma level of melatonin averaged about 40 +/- 2 pg/ml and was not significantly affected by CIP, by central application of L-tryptophan (0.02, 0.2 or 2.0 mg/rat) or by peripheral administration of this melatonin precursor used at doses of 2.5 or 25 mg/kg i.p. Plasma melatonin level was markedly increased by pretreatment of the rats with L-tryptophan given i.p. at dose of 250 mg/kg. We conclude that central administration of melatonin precursor; L-tryptophan, as well as peripheral application of high dose of this melatonin precursor prevented the pancreatic damage produced by CIP. The favorable effect of peripherally administered L-tryptophan could be related to the rise of melatonin plasma level and to pancreatoprotective action of this indoleamine. The beneficial effect of centrally administered L-tryptophan could be mediated through activation of central recep

Topics: Acute Disease; Aldehydes; Amylases; Animals; Ceruletide; Dose-Response Relationship, Drug; Drug Administration Schedule; Infusions, Parenteral; Injections, Intraperitoneal; Injections, Intraventricular; Lipase; Male; Malondialdehyde; Melatonin; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Reactive Oxygen Species; Superoxide Dismutase; Tryptophan; Tumor Necrosis Factor-alpha

There have been various studies of exocrine pancreatic function after acute pancreatitis, but few have examined the relationship between this function and the etiology of the pancreatitis. The aim of this work was to study pancreatic function in patients who had had acute alcoholic or acute biliary pancreatitis.. Seventy-five patients who had had a single attack of acute pancreatitis were studied. The etiology was alcohol in 36 and cholelithiasis in 39. Pancreatic function was studied between 4 and 18 months after pancreatitis by duodenal intubation in 18 patients (8 alcohol, 10 lithiasis) and by the amino acid consumption test (AACT) in the remaining 57 (28 alcohol, 29 lithiasis). For those who underwent AACT, the test was repeated 1 year after the first examination.. Among the 36 patients with alcoholic pancreatitis, most had impaired pancreatic function at both duodenal intubation (8/8, 100%) and at AACT (22/28, 78.6%); at the second test, the AACT remained pathological (18/23, 82.1%). Of the 39 patients with biliary pancreatitis, only 4 of the 10 (40%) who underwent duodenal intubation and only 5 of the 29 (17.2%) who performed AACT had pancreatic insufficiency; at the second test, only 4 of the 26 (15.4%) who repeated the AACT were pathological. The differences in the frequency and degree of pancreatic insufficiency between patients with alcoholic and those with biliary pancreatitis were statistically significant.. The results show that after alcoholic acute pancreatitis, the pancreatic insufficiency was significantly more frequent and more severe than after biliary pancreatitis. These findings together with the fact that the insufficiency was also more persistent suggest that acute alcoholic pancreatitis may occur in a pancreas that already has chronic lesions.

Topics: Acute Disease; Adolescent; Adult; Aged; Amino Acids; Bicarbonates; Ceruletide; Cholelithiasis; Chymotrypsin; Exocrine Pancreatic Insufficiency; Female; Humans; Lipase; Male; Middle Aged; Pancreas; Pancreatitis; Pancreatitis, Alcoholic

The pancreas contains a local renin-angiotensin system (RAS), which is subject to activation by experimental pancreatitis. In the exocrine pancreas, angiotensin II receptor subtypes AT1 and AT2 have been localized in the pancreatic ducts, blood vessels and acinar cells. We hypothesize that local RAS activities may have a potential role in regulating pancreatic acinar digestive enzyme secretion. The present study was designed to elucidate firstly the existence of RAS components in pancreatic acinar cells and their regulation by acute pancreatitis. Secondly, the differential roles of AT1 and AT2 receptors in controlling digestive enzyme secretion from dispersed functional pancreatic acini were also investigated. The mRNA levels of RAS components were assessed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). Acinar secretions were assayed by the measurement of alpha-amylase and lipase activities. Induction of acute pancreatitis was achieved by hyperstimulation of two intraperitoneal (i.p.) injections of cerulein (50 microg/kg/h). Results from RT-PCR showed that the mRNA levels of the major RAS components (angiotensinogen, AT1 and AT2 receptors) were expressed in isolated rat pancreatic acinar cells, and they were upregulated during pancreatitis. Exogenous addition of angiotensin II could stimulate a dose-dependent release of digestive enzymes from the acinar cells. Administration of the selective AT1 receptor antagonist losartan significantly inhibited the acinar digestion enzyme secretion in both normal and pancreatitis-induced acini. However, a specific AT2 receptor blocker PD123319 did not exhibit such a suppressive effect. These data indicate the existence of an acinar RAS in the pancreas of potential importance in the physiological regulation of digestive enzyme secretion. The differential actions of AT1 and AT2 receptors and their upregulation may have clinical relevance to the pathogenesis and management of acute pancreatitis.

Topics: Acute Disease; Angiotensin II Type 1 Receptor Blockers; Animals; Ceruletide; Enzymes; Gene Expression Regulation, Enzymologic; Imidazoles; Losartan; Pancreas; Pancreatitis; Pyridines; Rats; Rats, Sprague-Dawley; Receptor, Angiotensin, Type 1; Receptor, Angiotensin, Type 2; Renin-Angiotensin System; Vasoconstrictor Agents

Calpain, a calcium-dependent cytosolic cysteine protease, is implicated in a multitude of cellular functions but also plays a role in cell death. Recently, we have shown that two ubiquitous isoforms, termed micro-calpain and m-calpain, are expressed in rat pancreatic acinar cells and that calcium ionophore-induced calpain activation leads to acinar cell injury. On the basis of these observations, we have now investigated the role of both calpain forms and the endogenous calpain inhibitor calpastatin in acute pancreatitis. After treatment of rats either without or with calpain inhibitor Z-Val-Phe methyl ester (ZVP; 60 mg/kg i.p.), pancreatitis was induced by cerulein injections (10 microg/kg i.p.; 5 times at hourly intervals). Calpain activation and calpastatin expression in the pancreatic tissue were studied by Western blot analysis. Pancreatic injury was assessed by plasma amylase activity, pancreatic wet/dry weight ratio (edema), histological and electron-microscopic analyses, as well as fluorescence labeling of actin filaments. Cerulein caused an activation of both micro-calpain and m-calpain, accompanied by degradation of calpastatin. Prophylactic administration of ZVP reduced the cerulein-induced calpain activation but had no effect on calpastatin alterations. In correlation to the diminished calpain activity, the severity of pancreatitis decreased as indicated by a decline in amylase activity (P < 0.01), pancreatic edema formation (P < 0.05), histological score for eight parameters (P < 0.01), and actin filament alterations. Our findings support the hypothesis that dysregulation of the calpain-calpastatin system may play a role in the onset of acute pancreatitis.

Topics: Acute Disease; Animals; Calcium-Binding Proteins; Calpain; Ceruletide; Dipeptides; Enzyme Activation; Female; Pancreas; Pancreatitis; Rats; Rats, Inbred Lew; Severity of Illness Index

Traditional Chinese medicine is a potent agent in the management of clinical and experimental acute pancreatitis (AP), but the molecular mechanism of its therapeutic action is unclear. Numerous experimental and clinical studies have shown that platelet endothelial cell adhesion molecule-1 (PECAM-1) is pivotal to leukocyte recruitment, which results in microcirculatory injury during inflammation, but its role in acute pancreatitis is poorly understood. We investigated the effects of a compound of traditional Chinese medicine pancreatitis-1 (TCMP-1) on the changes of platelet endothelial cell adhesion molecule-1 (PECAM-1) expression on polymorphonuclear leukocytes (PMNs) in acute edematous pancreatitis (AEP).. The model of acute pancreatitis was established by subcutaneous injection of caerulein, and TCMP-1 treated groups were given TCMP-1 by catheterization from mouth to stomach (20 ml/kg) immediately after first time subcutaneous injection of caerulein. The changes of expression of PECAM-1 on leukocytes from the blood of the splenic vein and inferior vena cava were determined by flow cytometry.. In the AEP group, expression of PECAM-1 on PMNs was not significantly different between pancreatic microcirculation and systemic circulation at AEP2h and AEP4h time point. Then from AEP4h time point to AEP8h time point, expression of PECAM-1 was up-regulated in systemic circulation while it was down-regulated in pancreatic microcirculation and was significantly different between pancreatic microcirculation and systemic circulation at AEP8h time point (P<0.05). In the TCMP-1 treated group, compared with the AEP group, expression of PECAM-1 on PMNs decreased in different levels between pancreatic microcirculation and systemic circulation and was of significant difference at AEP8h time point (P<0.05).. Inhibition of PECAM-1 expression on PMNs may prevent PMNs from transmigration through the endothelium and may be one of the treatment mechanisms of TCMP-1 decoction on AEP.

Topics: Acute Disease; Animals; Ceruletide; Drugs, Chinese Herbal; Female; Gastrointestinal Agents; Leukocytes; Male; Models, Animal; Pancreatitis; Platelet Endothelial Cell Adhesion Molecule-1; Rats; Rats, Wistar

To investigate the role of inducible cyclooxygenase (COX-2) mRNA expression in local microvessels in rats with acute interstitial pancreatitis (AIP) induced by caerulein injection.. The reverse transcription polymerase chain reaction (RT-PCR) was used to detect COX-2 gene expression in pancreatic tissue. Parameters of acute pancreatitis, such as serum amylase (AMS) and plasma myeloperoxidase (MPO) activities, were assayed using spectrophotometry. Intravital fluorescence microscopy with fluorescein isothiocyanate-labelled erythrocytes was used to study the pancreatic microvessels of rats with AIP and normal control rats.. Highly significant increases in COX-2 expression and AMS and MPO activity were seen in rats with AIP compared with controls. After caerulein injection, pancreatic capillary blood flow was decreased (4 hours, p > 0.05; 8 hours, p < 0.001), functional capillary density was reduced (4 hours, p > 0.05; 8 hours, p < 0.001), and there was irregular and intermittent capillary perfusion at 8 hours. There was also a positive correlation between the level of COX-2 expression and MPO activity (plasma, r = 0.5449, p < 0.05; tissue, r = 0.5698, p < 0.05).. The correlations between increased COX-2 expression and decreased capillary perfusion and blood flow and increased oedema following AIP may show that COX-2 expression can induce neutrophil sequestration to the pancreas, which may be one of the cascading inflammatory factors in the development of AIP.

Topics: Acute Disease; Animals; Ceruletide; Cyclooxygenase 2; Gastrointestinal Agents; Gene Expression; Isoenzymes; Male; Microcirculation; Models, Animal; Pancreas; Pancreatitis; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Wistar

Nuclear factor-kappaB (NF-kappaB) is a transcription factor which plays a pivotal role in the induction of genes involved in the response to injury and inflammation. Calpain I inhibitor is a potent antioxidant which is an effective inhibitor of NF-kappaB. This study examined whether the postulate that calpain I inhibitor attenuates experimental acute pancreatitis.. In a murine model we measured NF-kappaB activation, expression of intercellular adhesion molecule (ICAM) 1, nitrotyrosine, inducible nitric oxide synthase (iNOS), nuclear enzyme poly(ADP-ribose) synthetase (PARS), myeloperoxidase, malondialdehyde, amylase and lipase and determined histological evidence of lung and pancreas injury in four groups: control (saline only), cerulein, calpain I inhibitor plus cerulein and calpain I inhibitor plus saline.. Intraperitoneal injection of cerulein in mice resulted in severe, acute pancreatitis characterised by oedema, neutrophil infiltration, tissue haemorrhage and necrosis and elevated serum levels of amylase and lipase. Infiltration of pancreatic and lung tissue with neutrophils (measured as increase in myeloperoxidase activity) was associated with enhanced lipid peroxidation (increased tissue levels of malondialdehyde). Immunohistochemical examination demonstrated a marked increase in immunoreactivity for nitrotyrosine, iNOS and PARS in the pancreas and lung of cerulein-treated mice. In contrast, pre-treatment with calpain I inhibitor markedly reduced: the degree of pancreas and lung injury; upregulation/expression of ICAM-1; staining for iNOS, nitrotyrosine and PARS; and lipid peroxidation. Additionally, calpain I inhibitor treatment significantly prevented the activation of NF-kappaB as suggested by the inhibition of IkappaB-alpha; degradation in the pancreas tissues after cerulein administration.. Taken together, our results clearly demonstrate that prevention of the activation of NF-kappaB by calpain I inhibitor ameliorates experimental murine acute pancreatitis.

Topics: Acute Disease; Analysis of Variance; Animals; Blotting, Western; Calpain; Ceruletide; Disease Models, Animal; Immunohistochemistry; Intercellular Adhesion Molecule-1; Lipid Peroxidation; Male; Mice; NF-kappa B; Nitric Oxide Synthase; Pancreatitis; Poly(ADP-ribose) Polymerases; Random Allocation; Respiratory Distress Syndrome; Tyrosine

Protease-activated receptor 2 can be stimulated by interstitially released trypsin during acute inflammation of the pancreas. In this study, we investigated the roles of pancreatic and circulatory protease-activated receptor 2 in the pathogenesis of acute pancreatitis by using in vitro and in vivo model systems.. Physiological and pathologic effects of protease-activated receptor 2 activation were measured in isolated pancreatic cells and in rats with experimental pancreatitis. Consequences of protease-activated receptor 2 activation on the systemic and inflammatory responses were measured after treatments with trypsin or protease-activated receptor 2-activating peptide.. Stimulation of protease-activated receptor 2 in rat pancreatic acinar cells activated short-lasting (Ca(2+) signaling) and long-lasting (extracellular signal-related kinase) signaling pathways and protected the cells against bile-induced cell damage. More importantly, protease-activated receptor 2 activation ameliorated the pathologic effects observed in the in vivo model of cerulein-induced pancreatitis. Trypsin in the circulation of rats with taurocholate-induced severe acute pancreatitis reached levels sufficient to activate endothelial and immune cells to stimulate nitric oxide and interleukin-8 production, respectively. Most notably, activation of systemic protease-activated receptor 2 by circulating protease-activated receptor 2 agonists induced a hemodynamic response pattern similar to that observed in rats with severe acute pancreatitis. The effects of protease-activated receptor 2 agonists and acute pancreatitis were not additive.. These findings suggest that protease-activated receptor 2 may have a dual role in acute pancreatitis: protecting acinar and duct cells against pancreatitis-induced cell damage while mediating or aggravating the systemic complications of acute pancreatitis, which are the major cause of mortality in the early phase of necrotizing pancreatitis.

Topics: Acute Disease; Animals; Blood Pressure; Cell Survival; Cells, Cultured; Ceruletide; Endopeptidases; Gene Expression; In Vitro Techniques; Male; Monocytes; Pancreas; Pancreatic Ducts; Pancreatitis; Rats; Rats, Sprague-Dawley; Receptor, PAR-2; Trypsin

Heat shock protein (Hsp) 27 regulates actin cytoskeletal dynamics, and overexpression of Hsp27 in fibroblasts protects against stress in a phosphorylation-dependent manner. Induction of Hsps occurs in acute pancreatitis, but Hsp27 has not been ascribed a specific role. To examine whether Hsp27 would ameliorate acute pancreatitis, we generated transgenic mice overexpressing human Hsp27 (huHsp27) or Hsp27 with the phosphorylatable residues Ser(15,78,82) mutated to aspartic acid (huHsp27-3D) to mimic phosphorylation or to alanine (huHsp27-3A), which is nonphosphorylatable.. huHsp27 was expressed at high levels in the exocrine pancreas by use of a cytomegalovirus promoter. Protein expression was analyzed by Western blotting and immunofluorescence. Acute pancreatitis was induced with 6 or 12 hourly cerulein injections (50 microg/kg intraperitoneally) and its severity assessed by measuring serum amylase and lipase levels, pancreatic trypsin activity, edema, and morphologic changes by quantitative scoring of multiple histologic sections and visualization of filamentous actin. Systemic inflammatory effects were monitored by measuring lung myeloperoxidase activity (a marker of neutrophil infiltration).. huHsp27 protein was overexpressed in the pancreas and localized to pancreatic acini. Acute pancreatitis was ameliorated by overexpression of huHsp27 and the huHsp27-3D mutant, which were associated with suppression of pancreatic trypsin activity and acinar cell injury and preservation of the actin cytoskeleton. In contrast, these changes were unaffected by overexpression of the nonphosphorylatable huHsp27-3A mutant.. Pancreatic overexpression of huHsp27 protects against cerulein-induced acute pancreatitis in a specific phosphorylation-dependent manner and is associated with preservation of the actin cytoskeleton.

Topics: Actins; Acute Disease; Animals; Ceruletide; Cytoskeleton; Female; Gastrointestinal Agents; Heat-Shock Proteins; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Models, Animal; Molecular Chaperones; Neoplasm Proteins; Pancreatitis; Phosphorylation; Severity of Illness Index

Infection of pancreatic necrosis is a severe complication of acute pancreatitis. Because Toll-like receptor 4 (TLR4) has been identified as a receptor necessary to transduct the signal of bacteria-derived lipopolysaccharide into cells, we investigated the role of TLR4 on pancreatic and pulmonary injury in acute pancreatitis and acute pancreatitis associated with endotoxemia in wild-type and TLR4-deficient mice.. Laboratory investigation.. University laboratory.. Heterozygous TLR4 mice.. Mice were injected intraperitoneally with a supramaximal dose of cerulein each hour for 10 hrs. To mimic infection, additional groups of mice were injected with lipopolysaccharide in the presence or absence of cerulein injections.. The severity of acute pancreatitis was assessed by serum amylase activity, pancreatic edema, acinar cell necrosis, and pancreas myeloperoxidase activity. Lung injury was quantitated by lung microvascular permeability and lung myeloperoxidase activity. Injections of cerulein induced an edematous pancreatitis that was of similar severity in wild-type and TLR4-deficient mice. Lipopolysaccharide alone had no toxic effect on pancreas and lungs and did not worsen the pancreatic injury induced by cerulein in wild-type and TRL4-deficient mice. In contrast, lipopolysaccharide worsened pancreatitis-associated lung injury, and the deficiency in TLR4 fully prevented this aggravation.. TLR4 may not play a role in the pancreatitis-associated lung injury but participates in the pulmonary injury mediated by endotoxemia.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Lipopolysaccharides; Lung Diseases; Membrane Glycoproteins; Mice; Pancreas; Pancreatitis; Receptors, Cell Surface; Toll-Like Receptor 4; Toll-Like Receptors

Microcirculatory disturbances are important early pathophysiological events in various organs during acute pancreatitis. The aim of the study was to evaluate changes in microperfusion of the pancreas, liver, kidney, stomach, colon, skeletal muscle, and to investigate the influence of heparin on the organ microcirculation in caerulein-induced experimental acute pancreatitis.. Acute pancreatitis was induced by 4 intraperitoneal injections of caerulein (Cn) (15 microg/kg). The organ microcirculation was measured by laser Doppler flowmetry. Serum interleukin 6 and hematocrit levels were analysed.. Acute pancreatitis resulted in a significant drop of microperfusion in all examined organs. Heparin administration (2 x 2.5 mg/kg) improved the microcirculation in pancreas (36.9 +/- 4% vs 75.9 +/- 10%), liver (56.6 +/- 6% vs 75.2 +/- 16%), kidney (45.1 +/- 6% vs 79.3 +/- 5%), stomach (65.2 +/- 8% vs 78.1 +/- 19%), colon (69.8 +/- 6% vs 102.5 +/- 19%), and skeletal muscle (59.2 +/- 6% vs 77.9 +/- 13%). Heparin treatment lowered IL-6 (359.0 +/- 66 U/mL vs 288.5 +/- 58 U/mL) and hematocrit level (53 +/- 4% vs 46 +/- 3%).. Heparin administration has a positive influence on organ microcirculatory disturbances accompanying experimental Cn-induced acute pancreatitis.

Topics: Acute Disease; Animals; Anticoagulants; Ceruletide; Hematocrit; Heparin; Male; Microcirculation; Pancreas; Pancreatitis; Rats; Rats, Wistar; Splanchnic Circulation

Oxidative stress plays an important role in the early stage of acute pancreatitis, as well as in the associated multiple organ injury. This study tests the hypothesis that M40401, a new superoxide dismutase mimetic, attenuates experimental acute pancreatitis. Intraperitoneal injection of cerulein in mice resulted in a severe, acute pancreatitis that was characterized by edema, neutrophil infiltration, tissue hemorrhage, and cell necrosis, as well as increases in the serum levels of amylase and/or lipase. The infiltration of the pancreatic tissue of these animals with neutrophils (measured as an increase in myeloperoxidase activity) was associated with expression of intercellular adhesion molecule-1, as well as signs of enhanced lipid peroxidation (e.g., increased tissue levels of malondialdehyde). Immunohistochemical examination demonstrated a marked increase in the staining (immunoreactivity) for nitrotyrosine and poly (ADP-ribose) polymerase in the pancreas of cerulein-treated mice. In contrast, the degree of pancreatic inflammation and tissue injury (histological score), the expression of intercellular adhesion molecule-1, the staining for nitrotyrosine and poly (ADP-ribose) polymerase, and lipid peroxidation were markedly reduced in pancreatic tissue sections obtained from cerulein-treated mice administered with M40401. These results confirm our hypothesis that superoxide anions play an important role in cerulein-mediated acute pancreatitis and support the possible clinical use of low-molecular-weight synthetic superoxide dismutase mimetics in those conditions that are associated with overproduction of superoxide.

Topics: Acute Disease; Animals; Ceruletide; Male; Manganese; Mice; Mice, Inbred Strains; Organometallic Compounds; Pancreatitis; Peroxidase; Poly(ADP-ribose) Polymerases; Superoxide Dismutase

To investigate the fluid shear stress induced changes of (Ca(2+))i in neutrophils in pancreatic microcirculation of experimental acute pancreatitis (AP).. Wistar rats (n = 36) were randomized into three groups. A model of AP was established by subcutaneous injection of caerulein. Low-shear 30 viscometer was used to provide steady fluid shear stress on separated neutrophils. The mean fluorescent intensity tested by flow cytometry was used as the indication of [Ca(2+)]i quantity.. Under steady shear, cytosolic [Ca(2+)]i showed biphasic changes. The shear rate changed from low to high, [(Ca(2+)]i in different groups decreased slightly and then increased gradually to a high level (P<0.05). A close correlation was observed between the cytosolic [Ca(2+)]i level and the alteration of fluid shear stress in regional microcirculation of AP.. The increase of [Ca(2+)]i is highly related to the activation of neutrophils, which contributes to neutrophil adhesion to endothelium in the early phase of AP. The effect of fluid shear stress on [Ca(2+)]i may play a crucial role in pancreatic microcirculatory failure of AP.

Topics: Acute Disease; Animals; Calcium; Ceruletide; Cytosol; Male; Microcirculation; Neutrophils; Pancreas; Pancreatitis; Rats; Rats, Wistar; Stress, Mechanical

Nosocomial pneumonia is a feared complication in the critically ill patient. Serious acute pancreatitis is frequently complicated by infections. The objectives of this study were to determine the influence of acute pancreatitis on host defense against Pseudomonas pneumonia and to determine the influence of Pseudomonas pneumonia on the severity of concurrent pancreatitis.. A controlled, in vivo laboratory study.. Research laboratory of a health sciences university.. Female C57Bl/6 mice.. Pancreatitis was induced by 12 hourly intraperitoneal injections of cerulein (pancreatitis) or saline (sham) immediately followed by intranasal administration of Pseudomonas aeruginosa (to induce pneumonia) or saline (controls). Mice were killed 24 hrs later. Hence, four groups were studied: sham/control, pancreatitis/control, sham/pneumonia, and pancreatitis/pneumonia mice.. When compared with sham/pneumonia mice, pancreatitis/pneumonia mice demonstrated exaggerated lung inflammation, higher bacterial counts in lungs and pancreas, and enhanced dissemination of the infection. Concurrently, pneumonia prolonged the course of pancreatitis, as reflected by histopathology and higher plasma amylase and relative pancreas weights (all p < .05 for the difference between pancreatitis/pneumonia and pancreatitis/control mice), which was associated with the localization of Pseudomonas in the pancreas.. Acute pancreatitis impairs host defense against Pseudomonas pneumonia, whereas pneumonia prolongs the course of pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Female; Gastrointestinal Agents; Mice; Mice, Inbred C57BL; Models, Animal; Pancreatitis; Pneumonia, Bacterial; Pseudomonas Infections; Severity of Illness Index

Matrix metalloproteinases, in particular gelatinase B/MMP-9, are key mediators in autoimmune diseases like multiple sclerosis and rheumatoid arthritis, but their pathogenic roles in diabetes are not well established. Gelatinase B has previously been shown to be upregulated in pancreas tissue from patients with acute and chronic pancreatitis and was suggested to exacerbate diabetes by cleaving insulin. In this study, the role of gelatinase B in diabetes was investigated using two streptozotocin-induced animal models of type I diabetes. In both a hyperacute and a subacute model, gelatinase B upregulation was found to be associated with disease activity. However, gelatinase B deficiency did not significantly protect against diabetes development, and wild-type and gelatinase B-deficient animals behaved similarly in terms of beta-cell apoptosis or necrosis. The fact that gelatinase B was found almost exclusively as the inactive pro-enzyme in most of the streptozotocin-induced diabetic animals may explain the lack of a gelatinase B effect. On the contrary, gelatinase B was completely activated in a minority (15%) of wild-type animals. This coincided with exocrine pancreatic inflammation, as revealed by the presence of active trypsin. The discovery of in vivo activation of progelatinase B by trypsin in acute pancreatitis is extended in a model of caerulein-induced pancreatitis. In the latter model, trypsinogen activation is systematically achieved and gelatinase B is found in its active form. In conclusion, gelatinase B itself is not a causative factor but, when activated by endogenous trypsin, is a permissive factor for insulin degradation and diabetes.

Topics: Acute Disease; Animals; Apoptosis; Blood Glucose; Ceruletide; Diabetes Mellitus, Experimental; Enzyme Activation; Islets of Langerhans; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Mice, Knockout; Necrosis; Pancreas, Exocrine; Pancreatitis; Trypsin; Up-Regulation

In pancreatic acini, the G-protein-activated phosphoinositide 3-kinase-gamma (PI3K gamma) regulates several key pathological responses to cholecystokinin hyperstimulation in vitro. Thus, using mice lacking PI3K gamma, we studied the function of this enzyme in vivo in two different models of acute pancreatitis. The disease was induced by supramaximal concentrations of cerulein and by feeding mice a choline-deficient/ethionine-supplemented diet. Although the secretive function of isolated pancreatic acini was identical in mutant and control samples, in both models, genetic ablation of PI3K gamma significantly reduced the extent of acinar cell injury/necrosis. In agreement with a protective role of apoptosis in pancreatitis, PI3K gamma-deficient pancreata showed an increased number of apoptotic acinar cells, as determined by terminal dUTP nick-end labeling and caspase-3 activity. In addition, neutrophil infiltration within the pancreatic tissue was also reduced, suggesting a dual action of PI3K gamma, both in the triggering events within acinar cells and in the subsequent neutrophil recruitment and activation. Finally, the lethality of the choline-deficient/ethionine-supplemented diet-induced pancreatitis was significantly reduced in mice lacking PI3K gamma. Our results thus suggest that inhibition of PI3K gamma may be of therapeutic value in acute pancreatitis.

Topics: Acute Disease; Animals; Apoptosis; Caspase 3; Caspases; Ceruletide; Choline Deficiency; Class Ib Phosphatidylinositol 3-Kinase; Diet; Dietary Supplements; Ethionine; In Situ Nick-End Labeling; Isoenzymes; Mice; Mice, Knockout; Necrosis; Neutrophils; Pancreatitis; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Survival Rate

To assess the effect of endothelins: ET-1, ET-2 and ET-3 on trypsinogen activation, lipase activity and histological changes in the pancreas in early (4 hrs) cerulein acute pancreatitis (AP) in rats.. In 45 Wistar rats with cerulein induced AP (2 x 40 microg/kg i.p. at 1 hour interval, the effect of endothelins at the dose 2 x 0.5 or 2 x 1.0 nmol/kg i.p. was assessed vs untreated AP; 6 healthy rats were control (C). Free active trypsin (FAT), total potential trypsin after activation with enterokinase (TPT), lipase in 12000 xg supernatants of pancreatic homogenates and the plasma alpha-amylase were assayed. The %FAT/TPT was an index of trypsinogen activation.. %FAT/TPT increased from 3.0 +/- 0.6 in C to 16.2 +/- 3.1 in AP (p < 0.01). ET-1 decreased this index to 4.8 +/- 1.1 after higher dose (p < 0.01); the effect of lower dose was insignificant. Attenuating effect of ET-2 was significant: 7.3 +/- 1.7 after higher dose (p < 0.05) and 6.1 +/- 0.9 after lower dose (p < 0.01). ET-3 diminished this index to 4.5 +/- 1.5 (p < 0.01) and to 6.3 +/- 2.2 (p < 0.05) respectively. Lipase activity in supernatant increased from 4.1 +/- 0.6 in C to 6.3 +/- 0.7 U/mg protein in untreated AP (p < 0.05) and plasma alpha-amylase from 7.0 +/- 0.6 in C to 25.9 +/- 4.3 U/ml in AP (p < 0.001), without essential changes in treated groups vs untreated AP. Higher doses of endothelins decreased inflammatory cell infiltration score in AP.. The exogenous endothelins, especially ET-2 and ET-3 and to lesser extent ET-1 exerted some protective effect in early, edematous acute pancreatitis by the attenuation of trypsinogen activation and inflammatory cell infiltration in the pancreas.

Topics: Acute Disease; alpha-Amylases; Animals; Ceruletide; Endothelin-1; Endothelin-2; Endothelin-3; Endothelins; Enzyme Activation; Lipase; Male; Pancreatitis; Rats; Rats, Wistar; Time Factors; Trypsinogen

Proinflammatory cytokines act as mediators of the local and systemic manifestations of acute pancreatitis (AP).. To investigate whether moderate hypothermia (MH) (32 degrees C) can reduce the severity of AP by inhibiting cytokine production in a rat model of caerulein-induced pancreatitis.. Rats were divided into three groups: control rats (Group I), AP rats treated with normothermia (38 degrees C) (Group II), and AP rats treated with MH (Group III). AP was induced by intramuscular injection of caerulein and intraperitoneal infusion of lipopolysaccharide. MH was induced 4 hours after the first caerulein injection. Serum interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, IL-6, amylase, and lipase levels were determined 8 hours after the first injection. The pancreas and lung were examined histologically.. MH in comparison with normothermia significantly reduced serum levels of IL-1beta, TNF-alpha, IL-6, amylase, and lipase. Histologically, the MH group showed less vacuolization of the acinar cells and cellular infiltration into the interacinar areas of the pancreas than were shown in the normothermia group, but these effects were not evident in the lung.. Our results suggest that MH may be clinically applicable for reducing the severity of AP.

Topics: Acute Disease; Amylases; Animals; Blood Gas Analysis; Blood Pressure; Ceruletide; Cytokines; Hypothermia, Induced; Interleukin-1; Interleukin-6; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Tumor Necrosis Factor-alpha

The early events leading to acinar cell injury during acute pancreatitis are poorly characterized. Signaling through gap junction channels contributes to the homeostasis of the exocrine pancreas by coordinating acinar cell activity within an acinus. To explore the role of gap junctional communication in acinar cell response to injury, we analyzed the course of acute pancreatitis induced by injection of cerulein in mice deficient for Cx32, the major gap junction protein expressed in the exocrine pancreas.. The severity of pancreatitis was evidenced by measuring serum amylase activity, pancreatic edema, acinar cell necrosis, pancreatic tumor necrosis factor alpha concentration, and myeloperoxidase activity. Acinar cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL), caspase-3 activity, and Bax/Bcl-2 expression. Expression and function of connexin were evaluated by immunofluorescence and dye coupling.. Cx32-deficient mice exhibited a deleterious course of acute pancreatitis with increased necrosis, edema, and inflammation of the exocrine pancreas. In addition, the exocrine pancreas of Cx32-deficient mice showed a decreased number of TUNEL-positive acinar cells and decreased caspase-3 activity but no change in Bax or Bcl-2 pancreatic expression. Interestingly, chemicals known to induce apoptosis in vivo had no effect on Cx32-deficient pancreatic acinar cells.. Deficiency of a pancreatic connexin converts a mild reversible form of acute pancreatitis into a severe disease and decreases the sensitivity of acinar cells to apoptotic stimuli. The results show that acinar cell-to-cell communication plays a key role in the modulation of severity of acute pancreatitis.

Topics: Acute Disease; Animals; Apoptosis; bcl-2-Associated X Protein; Ceruletide; Connexins; Gap Junction beta-1 Protein; Glutathione; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Knockout; Pancreas; Pancreatitis; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Severity of Illness Index

Topics: Acute Disease; Animals; Bacterial Proteins; Catecholamines; Ceruletide; Chaperonin 60; Chaperonins; Pancreatitis

Acute pancreatitis is an inflammatory disease characterized by pancreatic tissue edema, acinar cell necrosis, hemorrhage and inflammation of the damaged gland. It is believed that acinar cell injury is initiated by the activation of digestive zymogens inside the acinar cells, leading finally to the autodigestion of the pancreas. Previous study in our laboratory demonstrated that cerulein-induced acute pancreatitis was associated with an up-regulation of local renin-angiotensin system (RAS) in rat pancreas. Therefore, the utilization of RAS inhibitors may provide a novel and alternative treatment for acute pancreatitis. By means of a rat model of cerulein-induced acute pancreatitis, results from the present study showed that an intravenous injection of saralasin, an antagonist for angiotensin II receptors, at a dose of 40 microg/kg 30 min before the induction of acute pancreatitis significantly attenuated pancreatic edema. Results from the biochemical measurements showed that pretreatment with saralasin at a dose of 20 microg/kg markedly reduced pancreatic injury, as evidenced by the decreased activities of alpha-amylase and lipase in plasma. However, the same recipe of ramiprilat, a specific inhibitor for angiotensin-converting enzyme, at a dose of 20 microg/kg did not provide any protective effect against acute pancreatitis. On the contrary, pretreatment with ramiprilat at a dose 40 microg/kg enhanced cerulein-induced pancreatic injury. Results from histopathological analysis of these RAS inhibitors further confirmed with those results as obtained from biochemical analysis. These data indicate that administration of saralasin but not ramiprilat could be protective against acute pancreatitis and that activation of pancreatic RAS in acute pancreatitis may play a role in pancreatic tissue injury.

Topics: Acute Disease; alpha-Amylases; Angiotensin Receptor Antagonists; Animals; Ceruletide; Disease Models, Animal; Edema; Injections, Intravenous; Lipase; Necrosis; Pancreatitis; Ramipril; Rats; Rats, Sprague-Dawley; Renin-Angiotensin System; Saralasin

To establish a non-traumatic, easy to induce and reproducible mouse model of severe acute pancreatitis (SAP) induced with caerulein and lipopolyasccharide (LPS).. Thirty-two healthy mature NIH female mice were selected and divided at random into four groups (each of 8 mice), i.e., the control group (NS group), the caerulein group (Cn group), the lipopolysaccharide group (LPS group), and the caerulein+LPS group (Cn+LPS group). Mice were injected intraperitoneally with caerulein only, or LPS only, and caerulein and LPS in combination. All the animals were then killed by neck dislocation three hours after the last intraperitoneal injection. The pancreas and exo-pancreatic organs were then carefully removed for microscopic examination. And the pancreatic acinus was further observed under transmission electron microscope (TEM). Pancreatic weight, serum amylase, serum nitric oxide (NO) concentration, superoxide dismutase (SOD) and malondialdehyde (MDA) concentration of the pancreas were assayed respectively.. (1) NS animals displayed normal pancreatic structure both in the exocrine and endocrine. In the LPS group, the pancreas was slightly edematous, with the infiltration of a few inflammatory cells and the necrosis of the adjacent fat tissues. All the animals of the Cn group showed distinct signs of a mild edematous pancreatitis characterized by interstitial edema, infiltration of neutrophil and mononuclear cells, but without obvious parenchyma necrosis and hemorrhage. In contrast, the Cn+LPS group showed more diffuse focal areas of nonviable pancreatic and hemorrhage as well as systemic organ dysfunction. According to Schmidt's criteria, the pancreatic histologic score showed that there existed significant difference in the Cn+LPS group in the interstitial edema, inflammatory infiltration, parenchyma necrosis and parenchyma homorrhage in comparison with those of the Cn group, LPS group and NS group (P<0.01 or P<0.05). (2) The ultrasturcture of acinar cells was seriously damaged in the Cn+LPS group. Chromatin margination of nuclei was present, the number and volume of vacuoles greatly increased. Zymogen granules (ZGs) were greatly decreased in number and endoplasmic reticulum exhibited whorls. The swollen mitochondria appeared, the crista of which was decreased in number or disappeared. (3) Pancreatic weight and serum amylase levels in the Cn +LPS was significantly higher than those of the NS group and the LPS group respectively (P<0.01 or P<0.05). However, the pancreatic wet weight and serum amylase concentration showed no significant difference between the Cn+LPS group and the Cn group. (4) NO concentration in the Cn+LPS group was significantly higher than that of NS group, LPS group and Cn group(P<0.05 or P<0.01). (5) The SOD and MDA concentration of the pancreas in the Cn+LPS group were significantly higher than those of NS, LPS and Cn groups (P<0.05 or P<0.01).. The mouse model of severe acute pancreatitis could be induced with caerulein and LPS, which could be non-traumatic and easy to induce, reproducible with the same pathological characteristics as those of SAP in human, and could be used in the research on the mechanism of human SAP.

Topics: Acute Disease; Animals; Ceruletide; Female; Lipopolysaccharides; Mice; Mice, Inbred Strains; Pancreatitis; Severity of Illness Index

Free radical-mediated pancreatic injury is believed to play a key role in the pathogenesis of acute pancreatitis. Most of these studies have focused on the effects of antioxidant enzymes and free radical scavengers on improving the pancreatic injury. Recent findings showed that cerulein-induced acute pancreatitis was associated with an upregulation of a local pancreatic renin-angiotensin system in the pancreas. In the current study we hypothesized that inhibition of this renin-angiotensin system by saralasin, a nonspecific antagonist for angiotensin II receptor, could attenuate the severity of cerulein-induced pancreatitis.. The effects of saralasin on oxidative stress and tissue injury in cerulein-induced pancreatitis were assessed by histopathologic analysis and on the basis of biochemical changes of plasma alpha-amylase level, pancreatic glutathione status, oxidative modification of protein, and lipid peroxidation.. Data from the biochemical analysis showed that intravenous injections of saralasin at doses of 10 microg/kg to 50 microg/kg 30 minutes before the induction of acute pancreatitis significantly reduced pancreatic injury, as indicated by a decrease in plasma alpha-amylase activity in comparison with the cerulein-treated control. The effect of saralasin was further manifested by significant suppressions of glutathione depletion, oxidative modification of proteins, and lipid peroxidation in cerulein-treated rat pancreas. Histopathologic examination findings were in agreement with the biochemical data.. These data suggest that prophylactic administration of saralasin can ameliorate the oxidative stress and tissue injury in cerulein-induced pancreatitis. Such a protective effect may provide new insight into the potential value of angiotensin II receptor antagonists in the clinical therapy for acute pancreatitis.

Topics: Acute Disease; alpha-Amylases; Angiotensin Receptor Antagonists; Animals; Ceruletide; Glutathione; Lipid Peroxidation; Oxidative Stress; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Saralasin

Impaired lung function in severe acute pancreatitis is the primary cause of morbidity and mortality in this condition. Preprotachykinin-A (PPT-A) gene products substance P and neurokinin (NK)-A have been shown to play important roles in neurogenic inflammation. Substance P acts primarily (but not exclusively) via the NK1 receptor. NKA acts primarily via the NK2 receptor. Earlier work has shown that knockout mice deficient in NK1 receptors are protected against acute pancreatitis and associated lung injury. NK1 receptors, however, bind other peptides in addition to substance P, not all of which are derived from the PPT-A gene. To examine the role of PPT-A gene products in acute pancreatitis, the effect of PPT-A gene deletion on the severity of acute pancreatitis and the associated lung injury was investigated. Deletion of PPT-A almost completely protected against acute pancreatitis-associated lung injury, with a partial protection against local pancreatic damage. These results show that PPT-A gene products are critical proinflammatory mediators in acute pancreatitis and the associated lung injury.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Dose-Response Relationship, Drug; Gene Deletion; Lung; Lung Diseases; Mice; Mice, Knockout; Pancreas; Pancreatitis; Protein Precursors; Tachykinins

Acute pancreatitis is an inflammatory process of variable severity, and leukocytes are thought to play a key role in the development of pancreatitis and pancreatitis-associated lung injury. The effects of mediators released by these inflammatory cells may induce tissue damage. The aim of our study was to evaluate the role of the chemokine, macrophage inflammatory protein-2 (MIP-2), in the pathogenesis of cerulein-induced pancreatitis and pancreatitis-associated lung injury. The severity of pancreatitis was measured by serum amylase, pancreatic edema, acinar cell necrosis, and myeloperoxidase activity. Lung injury was quantitated by evaluating lung microvascular permeability and lung myeloperoxidase activity. To determine the role of MIP-2 in the pathophysiology of the disease, anti-MIP-2 antibody was administered either 1 hour before or 2 hours after the start of cerulein administration. MIP-2 concentrations increased in serum, pancreas, and lung tissues in mice treated with cerulein. Anti-MIP-2 antibody administrated either before or after cerulein partially protected against pancreas and lung injury. These results show that MIP-2 plays a key role in the pathophysiology of acute pancreatitis and that MIP-2 blockade may improve the outcome of the disease.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Chemokine CXCL2; Disease Models, Animal; Edema; Fluorescent Antibody Technique, Indirect; Gastrointestinal Agents; Injections, Intraperitoneal; Lung Diseases; Male; Mice; Mice, Inbred Strains; Monokines; Necrosis; Pancreas; Pancreatitis; Peroxidase

We hypothesized that genes expressed in pancreatic acinar cells during the initiation of acute pancreatitis determine the severity of the disease. Therefore, we utilized microarrays to identify those genes commonly induced in rat pancreatic acinar cells within 1-4 h in two in vivo models, caerulein and taurocholate administration. This strategy yielded 51 known genes representing a complex array of molecules, including those that are likely to either reduce or increase the severity of the disease. Novel genes identified in the current study included ATF3, BRF1, C/EBPbeta, CGRP, EGR-1, ephrinA1, villin2, ferredoxin, latexin, lipocalin, MKP-1, NGFI-B, RhoA, tissue factor (TF), and syndecan. To validate these microarray results, the role of EGR-1 was further investigated using quantitative RT-PCR, Western blotting, and immunocytochemistry. EGR-1 expression occurred within acinar cells and correlated with the development of caerulein-induced acute pancreatitis in rats. Furthermore, the levels of the inflammation-related genes MCP-1, PAI, TF, IL-6, and ICAM-1 and the extent of lung inflammation were reduced during the initiation of caerulein-induced acute pancreatitis in EGR-1-deficient mice. Thus this study identified EGR-1 and several other novel genes likely to be important in the development and severity of acute pancreatitis.

Topics: Acute Disease; Animals; Cells, Cultured; Ceruletide; DNA-Binding Proteins; Early Growth Response Protein 1; Gene Expression Profiling; Gene Expression Regulation; Immediate-Early Proteins; Inflammation; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; Taurocholic Acid; Transcription Factors

In isolated rat pancreatic acini, protein expression of RhoA and Rho-associated kinase, ROCK-II, and the formation of immunocomplex of RhoA with ROCK-II were enhanced by CCK-8, carbachol, and the phorbol ester TPA. The ROCK-specific inhibitor, Y-27632, did not alter basal amylase secretion, whereas it potentiated CCK-stimulated pancreatic enzyme secretion in vitro. During caerulein-induced pancreatitis occurring in mice in vivo, Y-27632 enhanced serum amylase levels and the formation of interstitial edema and vacuolization at 12-18h after the first injection of caerulein. Y-27632 in turn inhibited the recovery of protein expression of ROCK-II at 18h after the first caerulein injection. These results suggest that RhoA and ROCK-II assemble normal CCK-stimulated pancreatic enzyme secretion and prevent caerulein-induced acute pancreatitis.

Topics: Acute Disease; Amides; Amylases; Animals; Cell Polarity; Cells, Cultured; Ceruletide; Enzyme Inhibitors; Intracellular Signaling Peptides and Proteins; Male; Mice; Pancreas; Pancreatitis; Protein Serine-Threonine Kinases; Pyridines; Rats; Rats, Sprague-Dawley; rho-Associated Kinases; rhoA GTP-Binding Protein

Severity of systemic lesions and mortality of experimental acute pancreatitis (AP) are reduced after pancreatic enzyme content reduction induced by cerulein administration. Octreotide has been used both prophylatically and therapeutically in AP. The possible effects of octreotide on pancreatic enzyme content and its influence on pulmonary lesions of experimental AP were assessed in this study.. Wistar male rats were divided in two branches: BRANCH I - Animals divided into three groups: Group Sa (n = 10) intravenous saline infusion; Group Ce (n = 10) intravenous cerulein infusion, (0.133 micro g/kg(-1).h(-1)) and Group Oc (n = 10) SC octreotide (10 micro g/kg(-1)). Trypsin, elastase and amylase pancreatic contents as well as serum amylase were determined thereafter in all three groups; BRANCH II - Rats treated as in branch I, were submitted to sodium taurocholate AP (Groups Sa+AP, Ce+AP and Oc+AP). Two hours thereafter amylase and TAP assays were performed in serum, ascites and pancreatic tissue in eight animals of each group. Pulmonary histology was studied by morphometry 24 h after AP in the remaining animals.. Increased serum amylase and pancreatic enzyme contents were observed in octreotide-treated animals when compared to animals receiving saline or cerulein. After AP increases of serum and ascitic fluid amylase and of pancreatic TAP were observed in octreotide pre-treated animals when compared to saline and cerulein groups. Pulmonary interstitial and alveolar edema after AP was significantly increased in rats receiving octreotide as compared to the cerulein group.. Octreotide administration acutely increases the enzymatic content of the pancreas and thus may have a potential deleterious influence in the evolution of AP.

Topics: Acute Disease; Amylases; Animals; Ascitic Fluid; Ceruletide; Disease Models, Animal; Drug Therapy, Combination; Gastrointestinal Agents; Injections, Intravenous; Male; Octreotide; Pancreas; Pancreatic Elastase; Pancreatitis; Pulmonary Edema; Rats; Rats, Wistar; Taurocholic Acid; Trypsin

1 Kinin B(2) receptor antagonists or tissue kallikrein (t-KK) inhibitors prevent oedema formation and associated sequelae in caerulein-induced pancreatitis in the rat. We have now further investigated the mechanism of kinin generation in the pancreas. 2 Kinins were elevated in the pancreatic tissue already before oedema formation became manifest. Peak values (421+/-59 pmol g(-1) dry wt) were reached at 45 min and remained elevated for at least 2 h; a second increase was observed at 24 h. Pretreatment with the B(2) receptor antagonist icatibant abolished kinin formation, while post-treatment was ineffective. 3 Total kininogen levels were very low in the pancreas of controls, but increased 75-fold during acute pancreatitis. This increase was absent in rats that were pretreated with icatibant. 4 During pancreatitis, t-KK-like and plasma kallikrein (p-KK)-like activity in the pancreas, as well as trypsinogen activation peptide (TAP) increased significantly. Icatibant pretreatment further augmented t-KK about 100-fold, while p-KK was significantly attenuated; TAP levels remained unaffected. 5 Endogenous protease inhibitors (alpha(1)-antitrypsin, alpha(2)-macroglobulin) were low in normal tissues, but increased 45- and four-fold, respectively, during pancreatitis. This increase was abolished when oedema formation was prevented by icatibant. 6 In summary, oedema formation is initiated by t-KK; the ensuing plasma protein extravasation supplies further kininogen and active p-KK to the tissue. Concomitantly, endogenous protease inhibitors in the oedema fluid inhibit up to 99% of active t-KK. Our data thus suggest a complex interaction between kinin action and kinin generation involving positive and negative feedback actions of the inflammatory oedema.

Topics: Acute Disease; alpha 1-Antitrypsin; alpha-Macroglobulins; Animals; Anti-Inflammatory Agents, Non-Steroidal; Bradykinin; Ceruletide; Edema; Enzyme Activation; Female; Kininogens; Kinins; Pancreas; Pancreatitis; Plasma Kallikrein; Rats; Rats, Sprague-Dawley; Receptors, Cell Surface; Serine Proteinase Inhibitors; Tissue Kallikreins; Trypsinogen

Acute pancreatitis (AP) has been shown in some studies to inhibit total protein synthesis in the pancreas, whereas in other studies, protein synthesis was not affected. Previous in vitro work has shown that high concentrations of cholecystokinin both inhibit protein synthesis and inhibit the activity of the guanine nucleotide exchange factor eukaryotic initiation factor (eIF)2B by increasing the phosphorylation of eIF2alpha. We therefore evaluated in C57BL/6 mice the effects of caerulein-induced AP on pancreatic protein synthesis, eIF2B activity and other protein translation regulatory mechanisms. Repetitive hourly injections of caerulein were administered at 50 microg/kg ip. Pancreatic protein synthesis was reduced 10 min after the initial caerulein administration and was further inhibited after three and five hourly injections. Caerulein inhibited the two major regulatory points of translation initiation: the activity of the guanine nucleotide exchange factor eIF2B (with an increase of eIF2alpha phosphorylation) and the formation of the eIF4F complex due, in part, to degradation of eIF4G. This inhibition was not accounted for by changes in the upstream stimulatory pathway, because caerulein activated Akt as well as phosphorylating the downstream effectors of mTOR, 4E-BP1, and ribosomal protein S6. Caerulein also decreased the phosphorylation of the eukaryotic elongation factor 2, implying that this translation factor was not inhibited in AP. Thus the inhibition of pancreatic protein synthesis in this model of AP most likely results from the inhibition of translation initiation as a result of increased eIF2alpha phosphorylation, reduction of eIF2B activity, and the inhibition of eIF4F complex formation.

Topics: Acute Disease; Animals; Ceruletide; Eukaryotic Initiation Factor-2B; Eukaryotic Initiation Factor-4E; Eukaryotic Initiation Factor-4F; Mice; Pancreatitis; Peptide Elongation Factor 2; Phosphorylation; Protein Biosynthesis; Protein Kinases; Protein Serine-Threonine Kinases; Proteins; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; TOR Serine-Threonine Kinases

Severe acute pancreatitis leads to a systemic inflammatory response characterized by widespread leucocyte activation and, as a consequence, distant lung injury. In CC chemokines the first two cysteine residues are adjacent to each other. The aim of this study was to evaluate the effect of Met-RANTES, a CC chemokine receptor antagonist, on pancreatic inflammation and lung injury in caerulein-induced acute pancreatitis in mice.. Acute pancreatitis was induced in mice by hourly intraperitoneal injection of caerulein. Met-RANTES was administered either 30 min before or 1 h after starting caerulein injections, and pancreatic inflammation and lung injury were assessed. There were five groups of eight mice each including controls.. Treatment with Met-RANTES had little effect on caerulein-induced pancreatic damage. Met-RANTES, however, reduced lung injury when given either before administration of caerulein (mean(s.e.m.) lung myeloperoxidase (MPO) 1.47(0.19) versus 3.70(0.86)-fold increase over control, P = 0.024; mean(s.e.m.) microvascular permeability 1.15(0.05) versus 3.57(0.63) lavage to plasma fluorescein isothiocyanate-labelled albumin fluorescence ratio (L/P) per cent, P = 0.002) or after caerulein administration (lung MPO 1.96(0.27) versus 3.65(0.63)-fold increase over control, P = 0.029; microvascular permeability 0.94(0.04) versus 2.85(0.34) L/P per cent, P < 0.001).. Treatment with Met-RANTES reduces lung damage associated with caerulein-induced pancreatitis in mice. Chemokine receptor antagonists may be of use for the treatment of the systemic complications of acute pancreatitis.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Ceruletide; Chemokine CCL5; Drug Evaluation, Preclinical; Lung Diseases; Mice; Microcirculation; Pancreatitis; Receptors, Chemokine; Systemic Inflammatory Response Syndrome

The prostaglandin D2 metabolite, 15d-PGJ2, a potent natural ligand for peroxisome proliferator-activated receptor gamma (PPARgamma), exerts antiinflammatory effects by inhibiting the induction of inflammatory response genes and NF-kappaB-dependent transcription. AIM To determine whether 15d-PGJ2 decreases the severity of secretagogue-induced acute pancreatitis (AP) and to assess cellular mechanisms contributing to these effects. METHODOLOGY Swiss Webster mice were injected with either saline or cerulein (50 microg/kg) hourly for 8 hours and received either 15d-PGJ2 (2 mg/kg) or vehicle 1 hour before and 4 hours after induction of AP. RESULTS Treatment with 15d-PGJ2 significantly attenuated AP, as determined by histologic assessment of edema, vacuolization, inflammation, and necrosis. This attenuation was associated with decreased cyclooxygenase-2 (COX-2) and intercellular adhesion molecule-1 (ICAM-1) expression and decreased serum and pancreatic IL-6 levels. Treatment with 15d-PGJ2 markedly inhibited NF-kappaB DNA-binding activity, and, moreover, this decreased activity was associated with a concomitant inhibition of IkappaB protein degradation. CONCLUSION Our findings demonstrate that 15d-PGJ2 attenuates the severity of AP most likely through the inhibition of COX-2 expression, IL-6 production, and NF-kappaB activation. Ligands specific for PPARgamma may represent novel and effective means of clinical therapy for AP.

Topics: Acute Disease; Animals; Blotting, Western; Cell Nucleus; Ceruletide; Cyclooxygenase 2; Electrophoretic Mobility Shift Assay; Female; Gene Expression Regulation; I-kappa B Proteins; Intercellular Adhesion Molecule-1; Interleukin-6; Isoenzymes; Ligands; Mice; NF-kappa B; Pancreatitis; Prostaglandin D2; Prostaglandin-Endoperoxide Synthases; Protein Transport; Receptors, Cytoplasmic and Nuclear; RNA, Messenger; Time Factors; Transcription Factors

To set up a nontraumatic and convenient mouse model of severe acute pancreatitis (SAP). Caerulein(Cn) was injected the mice intraperitonealy with lipopolysaccharide(LPS). Serum amylase and pancreas weight were measured in experiment. The pathological changes of pancreas and other organs were observed under light microscope. The ultrastructure of acini were observed under transmission electron microscope (TEM). Serum NO concentration were measured and the SOD and MDA in pancreas were examined. The results in Cn + LPS group were showed that serum amylase, NO concentration and pancreas weight were increased, SOD deduced and MDA increased. Severe edema, inflammation infiltration, necrosis and different extent of hemorrhage were showed. The acini were damaged severely. And the lesion of other organs were also happened. In Cn group, there were only pancreatic interstitial edema but no parenchmal necrosis or hemorrhage, and the other organs were normal. In LPS group, pancreas were almost normal and the organs besides pancreas were only showed light inflammation infiltration. The SAP mouse model induced by caerulein plus LPS has the same pathological characteristics of human SAP, which can be used in human SAP research. The unbalance of oxygen free radical release-elimination and oxidation-antioxidation mechanisms might be involved in the pathogenesis of mouse model of severe acute pancreatitis induced by intraperitoneal injection of caerulein plus LPS.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Female; Lipopolysaccharides; Mice; Mice, Inbred Strains; Pancreas; Pancreatitis

We demonstrated dynamic aspects of granulocyte activation in rat severe acute pancreatitis, which was induced by cerulein and aggravated following lipopolysaccharide (LPS) injection. Pancreatitis induced by cerulein increased intracellular elastase activity of granulocytes in the blood. However, significant systemic cytokinemia was not provoked under such conditions. After induction of severe pancreatitis by LPS, intracellular elastase activity of circulating granulocytes decreased markedly and immediately. This decrease occurred simultaneous to induction of systemic hypercytokinemia and granulocyte migration into the lung. Overall results imply that: (1) circulating granulocytes are activated by induction of mild pancreatitis; (2) activation of granulocytes is mediated by factors other than systemic cytokinemia, such as locally produced cytokines; (3) those priming granulocytes immediately and significantly migrate from the circulation into the extravascular space by induction of endotoxemia; and (4) migration of granulocytes, in turn, may be mediated by systemic cytokinemia.

Topics: Acute Disease; Animals; Ceruletide; Cytokines; Granulocytes; Interleukin-1; Leukocyte Count; Lipopolysaccharides; Lung; Male; Pancreatic Elastase; Pancreatitis; Peroxidase; Rats; Rats, Wistar; Time Factors; Tumor Necrosis Factor-alpha

The role of nitric oxide (NO) in bacterial translocation (BT) associated with acute pancreatitis is controversial. We investigated the effects of the NO synthase substrate, L-arginine, and the NO synthase inhibitor, N-nitro-L-arginine methyl ester (L-NAME), on BT in caerulein-induced acute pancreatitis in rats.. Acute pancreatitis was induced by subcutaneous injections of caerulein (12 microg/kg) at 6-hour intervals for 2 days. Subcutaneous injections of L-arginine (100 mg/kg) or L-NAME (10 mg/kg) were administeredonce daily for 2 days. At 48 h, pancreatic injury and BT to the mesenteric lymph nodes (MLN), liver, and peritoneum were assessed.. Compared with controls, rats that received caerulein injections alone had increased BT to the MLN and pancreatic inflammatory changes. L-Arginine significantly reduced the inflammation and BT caused by caerulein. L-NAME did not significantly alter pancreatic inflammation. Although caerulein + L-NAME-treated rats had increased BT to the peritoneum, MLN, and liver compared with controls, rates of BT did not significantly differ between caerulein alone- and caerulein + L-NAME-treated rats.. In acute edematous pancreatitis, BT is increased and is regulated by NO. NO substrates limit BT and pancreatic inflammation associated with acute pancreatitis, probably by their bactericidal actions and ability to improve pancreatic blood flow.

Topics: Acute Disease; Animals; Arginine; Bacterial Translocation; Ceruletide; Edema; Injections, Subcutaneous; Liver; Lymph Nodes; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Pancreas; Pancreatitis; Peritoneum; Rats; Rats, Wistar

Here we compare the degree of pancreatitis caused by cerulein in mice lacking 5-lipoxygenase (5-LO) and in the corresponding wild-type mice. Intraperitoneal injection of cerulein in mice resulted in severe, acute pancreatitis characterized by oedema, neutrophil infiltration and necrosis and elevated serum levels of amylase and lipase. Infiltration of pancreatic and lung tissue with neutrophils (measured as increase in myeloperoxidase activity) was associated with enhanced lipid peroxidation (increased tissue levels of malondialdehyde). Immunohistochemical examination demonstrated a marked increase in immunoreactivity for intracellular adhesion molecule-1 (ICAM-1), P-selectin and E-selectin in the pancreas and lung of cerulein-treated mice. In contrast, the degree of (1) pancreatic inflammation and tissue injury (histological score), (2) up-regulation/expression of P-selectin, E-selectin and ICAM-1, and (3) neutrophil infiltration was markedly reduced in pancreatic and lung tissue obtained from cerulein-treated 5-LO-deficient mice. These findings support the view that 5-LO plays an important, pro-inflammatory role in the acute pancreatitis caused by cerulein in mice.

Topics: Acute Disease; Animals; Arachidonate 5-Lipoxygenase; Ceruletide; E-Selectin; Intercellular Adhesion Molecule-1; Lipid Peroxidation; Liver Diseases; Mice; Mice, Knockout; Neutrophil Infiltration; P-Selectin; Pancreatitis; Respiratory Distress Syndrome

Peptide YY (PYY), a gastrointestinal regulatory peptide, improves survival and histologic parameters in animal models of acute pancreatitis. Its effects on pancreatic cell growth and acute pancreatitis in pancreatic acinar and ductal cells are unknown. We hypothesized that PYY would affect cell growth and attenuate acute pancreatitis in pancreatic acinar and ductal cells in vitro.. Rat pancreatic acinar and ductal cells were cultured in the presence of 1) cerulein, a synthetic cholecystokinin analog that induces pancreatitis, 2) PYY, or 3) a combination group pretreated with PYY prior to addition of cerulein. Cell survival was measured at 48 h using MTT assay. Amylase secretion, as marker for pancreatitis, was measured at 48 h using an amylase activity assay. Statistical significance was calculated using analysis of variance and the Student's t test.. Peptide yy significantly increased cell growth and decreased amylase secretion compared with control and cerulein groups. Pretreatment with PYY significantly protected against the pancreatitis effects of cerulein.. We have shown for the first time that PYY has a mitogenic effect on pancreatic acinar and ductal cells in vitro. In addition, it directly protects against cerulein-induced pancreatitis. Its potential therapeutic benefit in acute pancreatitis would therefore be twofold: amelioration of the inflammatory process, and augmenting growth of normal pancreas to replace necrotic or apoptotic cell loss.

Topics: Acute Disease; Amylases; Animals; Cell Culture Techniques; Cell Division; Cell Survival; Ceruletide; Gastrointestinal Agents; Mitogens; Pancreas; Pancreatitis; Peptide YY; Rats

Hepatic injury is considered one of the critical complications associated with acute pancreatitis. It was proposed that initial insults to the liver in the early phase of the attack have an important priming effect, and the subsequent infectious attack (e.g. infectious pancreatic necrosis, bacterial translocation episode) constitutes a second attack on the liver.. To evaluate the role of priming by induction of cerulein pancreatitis and a following second attack by endotoxemia.. Plasma clearance and biliary excretion of indocyanine green (a hepatophillic hydrophobic organic anion).. A model of acute pancreatitis in rats.. Four groups of rats: untreated control, cerulein pancreatitis, endotoxemia and endotoxemia following the induction of cerulein pancreatitis (pancreatitis + endotoxemia).. Biliary indocyanine green excretion was significantly disturbed only in the pancreatitis + endotoxemia group. Plasma clearance (a reflection of hepatic uptake) of indocyanine green from the blood was only slightly affected in endotoxemia group.. Biliary secretion is quite sensitive to this hepatic injury model. Both the preceding priming insult and the following second attack are important in the development of hepatic injury.

Topics: Acute Disease; Alanine Transaminase; Animals; Bile; Bile Ducts, Intrahepatic; Ceruletide; Disease Models, Animal; Endotoxemia; Indocyanine Green; Lipopolysaccharides; Liver Function Tests; Male; Pancreatitis; Rats; Rats, Wistar

Activation of zymogens within the pancreatic acinar cell is an early feature of acute pancreatitis. Supraphysiologic concentrations of cholecystokinin (CCK) cause intrapancreatic zymogen activation and pancreatitis. Supraphysiologic concentrations of CCK also cause zymogen activation in isolated pancreatic acini. This activation first occurs in a nonzymogen granule compartment that contains lysosomal markers. A low pH environment may also be needed for activation. To examine the ability of alcohols to sensitize the acinar cell to CCK, the conversion of zymogens to active enzymes in isolated acini was assayed. Alcohols, including 35 mmol/L ethanol, sensitized acini to CCK induced activation. The sensitization increased with chain length and was less in branched compared with unbranched alcohols. The relationship of alcohol's structure to sensitization may be related to the mechanism of sensitization.

Topics: Acute Disease; Alcohols; Animals; Ceruletide; Cholecystokinin; Chymotrypsin; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Precursors; Ethanol; Hydrogen-Ion Concentration; Oligopeptides; Organelles; Pancreas; Pancreatitis; Time Factors; Trypsin

Nuclear factor (NF)-kappaB plays a central role in acute pancreatitis. We studied cerulein (CER)-induced pancreatitis in NF-kappaB knockout (KO) mice. NF-kappaB KO mice and normal control littermate wild-type (WT) mice were given four hyperstimulating doses of cerulein every hour to elicit secreatagogue-induced pancreatitis. Malonildialdehyde activity, glutathione levels, myeloperoxidase activity, TNF-alpha, and NF-kappaB binding activity and its inhibitory protein IkappaBalpha were studied in the pancreas. Furthermore, we measured plasma lipase and amylase and the histological damage. KO mice had reduced malonildialdehyde levels (WT + CER = 4.083 +/- 0.95 micromol/g; KO + CER = 1.513 +/- 0.63 microol/g), decreased myeloperoxidase activity (WT + CER = 19.3 +/- 2.39 mU/g; KO + CER = 10.21 +/- 2.05 mU/g), increased glutathione levels (WT + CER 6.22 +/- 2.46 micromol/g; KO + CER = 15. 516 +/- 2.92 micromol/g), and reduced serum levels of amylase (WT + CER = 2519 +/- 656.9 U/L; KO + CER = 916 +/- 280.4 U/L) and lipase (WT + CER = 1420 +/- 170 U/L; KO + CER = 861 +/- 172. 3 U/L). KO mice showed reduced pancreatic NF-kappaB activation, decreased TNF-alpha tissue content, and reduced histologic alterations. Our data suggest that KO mice have an attenuated cerulein-induced pancreatitis and help to define the possible interaction between NF-kappaB activation and oxidative stress in this deleterious event.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Glutathione; Lipase; Lymphotoxin-alpha; Malondialdehyde; Mice; Mice, Knockout; NF-kappa B; Pancreas; Pancreatitis; Peroxidase; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Necrosis Factor-alpha; Up-Regulation

Ghrelin, a circulating growth hormone-releasing peptide isolated from human and rat stomach, stimulates growth hormone secretion, food intake and exhibits gastroprotective properties. Ghrelin is predominantly produced by a population of endocrine cells in the gastric mucosa, but its presence in bowel, pancreas, pituitary and hypothalamus has been reported. In human fetal pancreas, ghrelin is expressed in a prominent endocrine cell population. In adult pancreatic islets the population of these cell is reduced. The aim of present study was to investigate the influence of ghrelin administration on the development of acute pancreatitis.. Acute pancreatitis was induced in rat by caerulein injection. Ghrelin was administrated twice (30 min prior to the first caerulein or saline injection and 3 h later) at the doses: 2, 10 or 20 nmol/kg. Immediately after cessation of caerulein or saline injections the following parameters were measured: pancreatic blood flow, plasma lipase activity, plasma interleukin-1beta (IL-1beta) and interleukin 10 (IL-10) concentration, pancreatic DNA synthesis, and morphological signs of pancreatitis.. Administration of ghrelin without induction of pancreatitis did not affect significantly any parameter tested. Caerulein led to the development of acute edematous pancreatitis. Treatment with ghrelin at the dose 2 nmol/kg, during induction of pancreatitis, was without effect on pancreatic histology or biochemical and functional parameters. Treatment with ghrelin at the dose 10 and 20 nmol/kg attenuated the development of pancreatitis and the effects of both doses were similar. Administration of ghrelin (10 or 20 nmol/kg) reduced inflammatory infiltration of pancreatic tissue and vacuolization of acinar cells. Also, plasma lipase activity and plasma IL-1beta concentration were reduced, and caerulein-induced fall in pancreatic DNA synthesis was reversed. Administration of ghrelin at the dose 10 and 20 nmol/kg was without effect on caerulein-induced pancreatic edema and pancreatitis-related fall in pancreatic blood flow.. (1) Administration of ghrelin attenuates pancreatic damage in caerulein-induced pancreatitis; (2) Protective effect of ghrelin administration seems Background: Ghrelin, a circulating growth hormone-releasing peptide isolated from human and rat stomach, stimulates growth hormone secretion, food intake and exhibits gastroprotective properties. Ghrelin is predominantly produced by a population of endocrine cells in the gastric mucosa, but its presence in bowel, pancreas, pituitary and hypothalamus has been reported. In human fetal pancreas, ghrelin is expressed in a prominent endocrine cell population. In adult pancreatic islets the population of these cell is reduced. The aim of present study was to investigate the influence of ghrelin administration on the development of acute pancreatitis. Methods: Acute pancreatitis was induced in rat by caerulein injection. Ghrelin was administrated twice (30 min prior to the first caerulein or saline injection and 3 h later) at the doses: 2, 10 or 20 nmol/kg. Immediately after cessation of caerulein or saline injections the following parameters were measured: pancreatic blood flow, plasma lipase activity, plasma interleukin-1beta (IL-1beta) and interleukin 10 (IL-10) concentration, pancreatic DNA synthesis, and morphological signs of pancreatitis. Results: Administration of ghrelin without induction of pancreatitis did not affect significantly any parameter tested. Caerulein led to the development of acute edematous pancreatitis. Treatment with ghrelin at the dose 2 nmol/kg, during induction of pancreatitis, was without effect on pancreatic histology or biochemical and functional parameters. Treatment with ghrelin at the dose 10 and 20 nmol/kg attenuated the development of pancreatitis and the effects of both doses were similar. Administration of ghrelin (10 or 20 nmol/kg) reduced inflammatory infiltration of pancreatic tissue and vacuolization of acinar cells. Also, plasma lipase activity and plasma IL-1beta conc; concentration were reduced, and caerulein-induced fall in pancreatic DNA synthesis was reversed. Administration of ghrelin at the dose 10 and 20 nmol/kg was without effect on caerulein-induced pancreatic edema and pancreatitis-related fall in pancreatic blood flow. Conclusions: (1) Administration of ghrelin attenuates pancreatic damage in caerulein-induced pancreatitis; (2) Protective effect of ghrelin administration seems to be related the inhibition in inflammatory process and the reduction in liberation o

Topics: Acute Disease; Animals; Ceruletide; DNA; Dose-Response Relationship, Drug; Drug Administration Schedule; Ghrelin; Humans; Injections, Intraperitoneal; Interleukin-1; Interleukin-10; Lipase; Male; Pancreas; Pancreatitis; Peptide Hormones; Rats; Rats, Wistar; Regional Blood Flow; Vacuoles

The aim of our study was to evaluate the possible influence of adenosine receptors agonists and antagonists on pancreatic blood flow (PBF) and the development of acute pancreatitis (AP) in rats.. Ninety male Wistar rats were subdivided into ten equal groups, nine rats in each. The study was carried out in two stages. In the first one the first group (control) received i.v. saline infusion for 12 hours. The groups 2-5 (the first stage) received i.v. caerulein infusion as in the first group, but with pretreatment with: in the second group--DPCPX (A1 receptor antagonist), in the third group--CGS 21680 (A2 receptor agonist), in the fourth group--ZM 241385 (A2 receptor antagonist), in the fifth group--IB-MECA (A3 receptor agonist). In the second stage the first group received i.v. caerulein infusion at the dose of 5 micrograms/kg/h for 12 hours. The groups 2-5 (the second stage) received i.v. caerulein infusion as in the first group, but with pretreatment with: in the second group--DPCPX (A1 receptor antagonist), in the third group--CGS 21660 (A2 receptor agonist), in the fourth group--ZM 241385 (A2 receptor antagonist), in the fifth group--IB-MECA (A3 receptor agonist). Pancreatic blood flow was measured by laser Doppler flowmetry. Pancreatic inflammation was evaluated by serum alpha-amylase activity, pancreatic weight and histological changes in the pancreatic tissue.. We observed a significant attenuation of serum alpha-amylase activity increase (19.1 +/- 2.8 kIU/L vs 30.12 +/- 2.64 kIU/L), pancreatic weight (expressed as percentage of rat's body weight--0.85 +/- 0.16% vs 1.25 +/- 0.14%), and improvement of PBF (79.8 +/- 6.1% vs 60.1 +/- 3.6%), a reduced degree of pancreatic tissue damage (oedema, leukocyte infiltration, vacuolisation of acinar cells) in the third group (CGS 21680 + caerulein) compared with the first group in the second stage (only caerulein infusion). Neither agonists nor antagonists exerterd any appreciable effects on measured parameters in healthy rats.. Pretreatment with A2 receptors agonist seems to be protective against the damage to the pancreas during the course of caerulein-induced acute pancreatitis in rats. This effect could be due to improvement of pancreatic blood flow. This finding could have some therapeutic implications.

Topics: Acute Disease; Animals; Ceruletide; Gastrointestinal Agents; Male; Models, Animal; Pancreas; Pancreatitis; Rats; Rats, Wistar; Receptors, Purinergic P1

Increase in intracellular chymotrypsin activity was reported during acute pancreatitis. Beside chymotrypsin, there are at least two enzymes with chymotrypsin-like activity: proteasome and lysosomal cathepsin A. Until now it is not known whether and to what extent they contribute to increases in chymotrypsin activity in acute pancreatitis. Our aim was to study organ chymotrypsin-like activities during experimental acute pancreatitis.. Rat cerulein model of acute pancreatitis was used. The chymotrypsin-like activities were assessed in pancreas, liver, lung, heart, spleen and kidney using highly selective synthetic substrates of the proteasome and the cathepsin A, at neutral and acidic pH. Determinations after addition of selective inhibitor were also performed.. During acute pancreatitis we found in the pancreas an increase only in neutral chymotrypsin-like activity, as compared to the control animals. In other organs neutral chymotrypsin-like activity did not increase, and in kidney it even decreased. There were no changes in acidic chymotrypsin-like activity in any of organs studied. The studies using the inhibitor of the proteasome showed that the neutral chymotrypsin-like activity in the pancreas of the rats with acute pancreatitis should not be attributed to the proteasome activity, but rather to the chymotrypsin.. Our results did not confirm any significant contribution of proteasome or cathepsin A to increased chymotrypsin-like activity in acute pancreatitis. We showed a decrease in neutral chymotrypsin-like activity of proteasome in the kidney, but the significance of this finding remains to be established.

Topics: Acute Disease; Animals; Ceruletide; Chymotrypsin; Gastrointestinal Agents; Hydrogen-Ion Concentration; Models, Animal; Pancreatitis; Rats; Rats, Wistar

Elevated Ca(2+) concentrations within the pancreatic acinar cells represent a risk factor for the development of acute pancreatitis. Apoptosis is an important characteristic of pancreatitis, with induction of apoptotic genes and intraceullar increase of calcium, endonucleases, and protease. The present study, which aims to investigate whether (1) cerulein induces apoptotic gene expression (bax, bid, p53) in pancreatic acinar AR42J cells and (2) cerulein-induced gene expression is mediated by intracellular Ca(2+), monitored the gene expression profile in the cells treated with the Ca(2+) chelator BAPTA-AM. Results showed that cerulein (10(-7) M) evoked an initial peak Ca(2+) signal; a further Ca(2+) signal was induced with second treatment of cerulein. Cerulein-induced Ca(2+) signal could not be detected in the cells treated with the Ca(2+) chelator BAPTA-AM. Cerulein dose-dependently induced apoptosis, determined by DNA fragmentation and pro-apoptotic bid expression in AR42J cells. Cerulein induced bid, bax, and p53 mRNA expression, which was inhibited in the cells treated with cerulein and cultured in the presence of BAPTA-AM. The present results suggest that increase in the free cytosolic Ca(2+) may be the upstream event of apoptotic gene (bax, bid, p53) expression, which contribute to cerulein-induced apoptosis in pancreatic acinar cells.

Topics: Acute Disease; Apoptosis; Calcium; Calcium Signaling; Cell Line; Ceruletide; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Humans; Pancreatitis

experimental pancreatitis and explore the mechanism of bacterial translocation at the molecular level in mice with severe acute pancreatitis.. Thirty-six ICR mice were randomly divided into 6 groups with 6 mice in each group. The animals in the experimental groups (from A to E) received seven intraperitoneal injections of caerulein (50 microg/kg body weight) at hourly intervals over 6 hours, and were killed at 9, 18, 24, 48 and 72 hours after the first injection. The control group (F) received intraperitoneal injection of the same volume of saline, and the animals were killed at the 18th hour after the first injection. Blood and pancreatic tissue samples were obtained after the animals were killed. Amylase and pancreatic pathological alterations were observed. The ileum sequential segments were removed from each mouse. The amplification products of RT-PCR were electrophoresed and the images were analyzed by UVI software.. Acute necrotizing pancreatitis in ICR mice were induced by the intraperitoneal injection of caerulein at large doses. Markedly pathological lesions were observed at the 18th hour after the first injection of caerulein. And, intestinal cryptdin-4 mRNA expression was down regulated slightly at the 9th hour and most markedly at the 18th hour (P < 0.05). After 24 hours, the cryptdin-4 mRNA expression recovered to the normal level gradually.. Acquired cryptdin-4 deficiency may play an important role in the pathogenesis of bacterial tanslocation in acute necrotizing pancreatits.

Topics: Acute Disease; alpha-Defensins; Animals; Base Sequence; Ceruletide; Intestine, Small; Male; Mice; Mice, Inbred ICR; Molecular Sequence Data; Pancreatitis; Random Allocation; RNA, Messenger

Pancreatic stone protein (PSP/reg) is a constitutively secreted protein in pancreatic juice. Pancreatitis-associated protein (PAP) belongs to the same family of proteins. PAP is highly increased during acute pancreatitis, while no exact data exist regarding PSP/reg protein synthesis and secretion. Recently, an attempt to determine PSP/reg and PAP levels in sera of rats with acute pancreatitis showed a significant increase in PAP but failed to demonstrate changes in PSP/reg. Others reported that surgical manipulation of the pancreas, including sham controls, affected mRNA levels of PSP/reg. Neither report determined protein levels of PSP/reg.. Rats were treated intraperitoneally with a supramaximal dose of caerulein to induce pancreatitis, a physiological dose of caerulein, or a saline injection. Pancreata were analyzed for PAP and PSP/reg using ELISAs. RNA was extracted for Northern blot analysis of PAP I, II, and III and PSP/reg mRNA.. Experimental induction of acute pancreatitis caused a coordinate increase in both PSP/reg and PAP. PAP showed an acute response and returned to low levels within 48 h while PSP/reg exhibited a more sustained response. Intraperitoneal application of a physiological dose of caerulein and even a saline injection caused an increase in PSP/reg.. PSP/reg and PAP levels are increased through similar mechanisms by physiological and supramaximal doses of caerulein. However, PSP/reg regulation appears to sustain high levels while PAP levels are more transient. Since the regulation of this protein family is affected even under mild stress, we define them as secretory stress proteins.

Topics: Acute Disease; Acute-Phase Proteins; Animals; Antigens, Neoplasm; Biomarkers, Tumor; Calcium-Binding Proteins; Ceruletide; Lectins, C-Type; Lithostathine; Nerve Tissue Proteins; Pancreas; Pancreatitis; Pancreatitis-Associated Proteins; Protein Isoforms; Rats; RNA, Messenger; Tissue Distribution

Leptin is a pleiotropic hormone that is involved in the regulation of food intake and body weight. Recent findings demonstrated that leptin receptors are present in the pancreas but the involvement of leptin in pancreatitis remains unknown. The aim of the present study was: (1) to assess plasma leptin levels in rats with caerulein-induced pancreatitis (CIP) and humans with acute pancreatitis; and (2) to determine the effects of exogenous leptin on the course of acute CIP in rats.. CIP was produced in Wistar rats by s.c. infusion of 5 microg of caerulein for 5 h. Plasma leptin was measured by specific RIA and leptin expression in the pancreas was determined at the transcriptional and protein levels. In addition, the effects of exogenous leptin at the doses of 1 or 10 microg/kg i.p. on the course of CIP and the plasma levels and mRNA expression in pancreas of cytokines TNFalpha and IL-4 were studied. Furthermore, pancreatic cNOS and iNOS expression at mRNA level were measured in rats with CIP and pretreated with leptin. Parallel to these studies, the plasma levels of leptin were measured in 15 patients with acute edematous pancreatitis and in 30 healthy controls of comparable age and body mass index.. In rats, plasma leptin rose significantly from the median of 0.14 (0.03-0.3 ng/ml) in the control group to 0.56 (0.2-3.2 ng/ml) in rats with CIP. The CIP was associated with an upregulation of mRNA and protein for leptin in the pancreas. The administration of exogenous leptin significantly reduced the weight of pancreas, histological manifestations of pancreatitis, plasma TNFalpha and mRNA expression for iNOS in the pancreatic tissue. The assessment of leptin plasma level in humans demonstrated significantly higher median values of plasma leptin in patients with acute pancreatitis [7.5 (4.3-18.4 ng/ml)] than in healthy controls [2.1 (1.0-11.8 ng/ml)].. (1) Acute pancreatitis in rats and in humans is associated with a marked increase in the plasma level of leptin. (2) The transcriptional upregulation of leptin in the pancreas after induction of pancreatitis indicates that the inflamed pancreas could be the source of local production of leptin. (3) Exogenous leptin protects the pancreas against development of acute CIP in rats and one possible mechanism of action of leptin might be attributed to the activation of nitric oxide pathway.

Topics: Acute Disease; Animals; Ceruletide; Female; Humans; Interleukin-1; Interleukin-4; Leptin; Male; Middle Aged; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Pancreatitis; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Necrosis Factor-alpha

Severe pancreatitis is frequently associated with acute lung injury (ALI) and the respiratory distress syndrome. The role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in mediating the ALI associated with secretagogue-induced experimental pancreatitis was evaluated with GM-CSF knockout mice (GM-CSF -/-). Pancreatitis was induced by hourly (12x) intraperitoneal injection of a supramaximally stimulating dose of the cholecystokinin analog caerulein. The resulting pancreatitis was similar in GM-CSF-sufficient (GM-CSF +/+) control animals and GM-CSF -/- mice. Lung injury, quantitated by measuring lung myeloperoxidase activity (an indicator of neutrophil sequestration), alveolar-capillary permeability, and alveolar membrane thickness was less severe in GM-CSF -/- than in GM-CSF +/+ mice. In GM-CSF +/+ mice, pancreas, lung and serum GM-CSF levels increase during pancreatitis. Lung levels of macrophage inflammatory protein (MIP)-2 are also increased during pancreatitis, but, in this case, the rise is less profound in GM-CSF -/- mice than in GM-CSF +/+ controls. Administration of anti-MIP-2 antibodies was found to reduce the severity of pancreatitis-associated ALI. Our findings indicate that GM-CSF plays a critical role in coupling pancreatitis to ALI and suggest that GM-CSF may act indirectly by regulating the release of other proinflammatory factors including MIP-2.

Topics: Acute Disease; Animals; Antibodies; Ceruletide; Chemokine CXCL2; Chemokines; Granulocyte-Macrophage Colony-Stimulating Factor; Leukocyte Count; Lung; Lung Diseases; Mice; Mice, Knockout; Pancreas; Pancreatitis; Reference Values; Severity of Illness Index

A common pathway in the pathogenesis of acute pancreatitis is the generation of free oxygen radicals. The most important defense mechanisms are free radical scavengers, especially glutathione. This study evaluates the influence of the inhibition of glutathione synthesis with L-buthionine-(S,R)-sulfoximine (BSO) on the course of experimentally induced acute pancreatitis in rats and the effects on isolated pancreatic acini and their secretion pattern. Thus acute necrotizing pancreatitis was induced with intraductal infusion of low-dose glycodeoxycholic acid and subsequent hyperstimulation with cerulein with and without pretreatment with BSO. In vitro pancreatic acini were isolated and stimulated with different concentrations of cerulein with and without BSO. The BSO-treated group showed a significantly reduced survival, more necrosis, and a decreased secretion of amylase in vivo. No effect on secretion pattern in either groups was seen in vitro and BSO did not exert toxic effects. Based on the data presented, this study demonstrates deleterious effects of scavenger depletion on the course of experimental pancreatitis. This is due to the systemic effects of free oxygen radicals rather than to local effects.

Topics: Acute Disease; Amylases; Animals; Buthionine Sulfoximine; Ceruletide; Enzyme Inhibitors; Glutathione; Glycodeoxycholic Acid; In Vitro Techniques; Male; Pancreas; Pancreatitis, Acute Necrotizing; Rats; Rats, Wistar

Prognosis of acute pancreatitis is related mainly to systemic involvement. The establishment of this systemic inflammation is mediated by proinflammatory cytokines. Our aim is to study serum levels of some proinflammatory cytokines and the associated damage of the lung in a model of experimental acute pancreatitis.. Eighty seven male Wistar rats were divided into two groups: group A (control) with saline solution administration; group B with acute pancreatitis induced by intraperitoneal caerulein (50 mg/kg every hour, 4 doses). The animals were killed at 0, 2, 6 and 24 hours of the last dose of caerulein or saline solution. Pancreatic and pulmonary histology were examined, and serum levels of IL-1 beta, TNF-alpha and IL-6 were evaluated, as well as some laboratory parameters as indicators of systemic involvement.. The administration of caerulein induced an acute edematous pancreatitis without mortality and with a trend towards resolution in 24 hours. IL-1 beta in animals with acute pancreatitis showed significantly higher levels than in the control group at 6 hours. Serum transaminases, urea and creatinine were also significantly higher at 2 and 6 h. The group with acute pancreatitis showed histological lung damage all over the study.. In our model of acute pancreatitis we observed systemic involvement as judged by alterations of serum transaminases and parameters of renal function, as well as histological lung damage, that correlated with an increase in serum levels of IL-1b.

Topics: Acute Disease; Animals; Ceruletide; Cytokines; Gastrointestinal Agents; Lung; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar

We previously demonstrated that the immunostaining of the gap-junction protein, connexin 32 (Cx 32), in the pancreas was markedly reduced in caerulein (Cn)-induced acute pancreatitis. The expression of Cx 32 in the pancreas during the course of acute pancreatitis is unclear. To address this, we examined Cx 32 mRNA and protein expression in the pancreas.. Cx 32 mRNA and protein expression in the pancreas was examined by Northern blot analysis and Western blot analysis, respectively, 1, 4, 7, and 14 days after the induction of acute pancreatitis.. Cx 32 mRNA was identified in normal rat pancreas, and the value for the relative intensity against 18S rRNA was 0.57 +/- 0.15 (mean +/- SD). After the induction of acute pancreatitis by caerulein, the Cx 32 mRNA expression levels were increased on day 1, day 4, day 7, and day 14 compared with levels in the normal pancreas (1.63-fold, 1.61-fold, 1.49-fold, and 1.35-fold, respectively). A significant increase in Cx 32 protein expression was detected on day 1 and day 4 (1.67 +/- 0.15-fold and 1.72 +/- 0.2-fold, respectively), while Cx 32-positive spots, determined by immunohistochemical analysis, were markedly decreased on day 1 and had returned to normal by day 14.. These results show that the expression of Cx 32 increases early on after the induction of pancreatitis by Cn, and that the normalization of Cx 32-immunostained spots in Cn-induced acute pancreatitis occurs after the increase in Cx 32 mRNA and protein expression.

Topics: Acute Disease; Animals; Blotting, Northern; Blotting, Western; Ceruletide; Connexins; Gap Junction beta-1 Protein; Immunohistochemistry; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

Both cerulein and cholecystokinin activate mitogen-activated protein (MAP) kinase (ERK1/2) in vivo and in isolated pancreatic acini.. ERK1/2 in pancreas homogenates was activated in rats rendered pancreatitic by subcutaneous injections of cerulein (5 microg/kg per hour). To determine if blocking ERK1/2 activity might rescue cerulein-induced acute pancreatitis, the "MAP kinase kinase" (also known as MEK1/2) inhibitors PD98059 and U0126 were administered in vivo.. In rats pretreated with PD98059 (10 mg/kg per i.v. injection) or U0126 (5 mg/kg per i.v. injection) 30 minutes before and then together with hourly cerulein injections for 3 hours, pancreatitis was significantly attenuated on the basis of pancreatic wet weight and histology. Serum amylase concentration was significantly reduced when PD98059 was administered intraperitoneally (10 mg/kg per intraperitoneal injection). PD98059 also ameliorated pancreatitis over a 6-hour cerulein time course. The phosphorylation of pancreatic ERK1/2 was attenuated in PD98059- and U0126-treated animals at both 30 minutes and 3 hours after cerulein injection. Rats rendered neutropenic with vinblastine and pretreated with U0126 still showed attenuated manifestations of cerulein-induced acute pancreatitis, a finding suggesting that pancreatic ERK1/2 is mostly responsible for the effect, rather than infiltrating neutrophils.. Inhibition of pancreatic ERK1/2 in vivo affords significant protection against inflammatory sequelae following cerulein-induced acute pancreatitis.

Topics: Acute Disease; Animals; Butadienes; Ceruletide; Enzyme Inhibitors; Flavonoids; Male; MAP Kinase Kinase 1; MAP Kinase Kinase 2; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Nitriles; Pancreatitis; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Rats; Rats, Sprague-Dawley

Acute pancreatitis (AP) is associated with increased cytokine production, which can ultimately produce deleterious local and systemic effects. The transcription factor NF-kappaB is activated by degradation of its inhibitory factor, IkappaB, and can stimulate various cytokines. The purpose of this study was to determine whether the inhibition of NF-kappaB binding activity with a novel peptide that binds to the NF-kappaB essential modifier binding domain (NBD) could attenuate the severity of AP.. AP was induced in Swiss Webster mice by hourly injections of the cholecystokinin analogue cerulein (50 microg/kg). Mice were injected with either the wild-type or control (mutated) NBD peptide at the time of the first cerulein injection; they were then sacrificed over a time course, and pancreata and lungs were harvested for histologic analysis and scoring. Myeloperoxidase activity was measured to assess neutrophil sequestration as an indicator of inflammation. NF-kappaB binding activity and steady-state levels of IkappaB and NF-kappaB subunits were determined by gel shift and Western blot, respectively.. AP resulted in increased NF-kappaB DNA-binding activity and decreased steady-state levels of IkappaB. Treatment with NBD peptide decreased inflammation in the pancreas, decreased hemorrhage in the lungs, and decreased myeloperoxidase activity in both pancreas and lung.. The marked induction of NF-kappaB binding activity suggests a role for this transcription factor in the early inflammatory changes associated with AP. Treatment with the NBD peptide attenuated the severity of injury associated with AP. Novel compounds that selectively target NF-kappaB may prove to be useful treatment of AP and AP-associated lung injury.

Topics: Acute Disease; Animals; Ceruletide; Cholecystokinin; DNA; Female; I-kappa B Proteins; Mice; NF-kappa B; Pancreas; Pancreatitis; Peptides; Peroxidase; Protein Binding

Infections are frequent complications and determine clinical course and outcome in severe pancreatitis. A novel animal model was used to assess minimal transit time of bacterial translocation (BT) across the gut mucosa in vivo using green fluorescent protein-transfected Escherichia coli and intravital video microscopy.. Three hours after induction of acute pancreatitis by i.p. injection of 40 microg/kg cerulein, 0.5 ml of a suspension of green fluorescent protein-transfected E. coli were injected into the lumen of a small bowel reservoir formed by ligature in anesthetized Wistar rats. Translocation of E. coli was assessed by intravital microscopy. Animals were sacrificed 5 h after induction of pancreatitis.. BT across the mucosa and into the muscularis propria took a mean +/- SD of 36.4 +/- 8 min and 80.9 +/- 9.5 min, respectively, in sham animals. Pancreatitis resulted in a significantly shorter minimal transit time across the mucosa (16.4 +/- 4.9 min, p = 0.007) and into the muscularis propria (47.7 +/- 2.5 min, p = 0.001). E. coli were detected on frozen cross-sections and on bacteriological examination of pancreatic tissue in animals with acute pancreatitis but not in controls.. Intravital microscopy of fluorescent bacteria is a new approach towards studying BT in vivo. Minimal transit time of BT serves as a novel functional aspect of mucosal barrier function during acute pancreatitis. The observation of fluorescent bacteria translocating from the small bowel lumen into the pancreas provides substantial experimental proof for the gut-origin-hypothesis of infectious complications in pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Escherichia coli; Ileum; Intestinal Mucosa; Male; Microscopy, Video; Muscle, Smooth; Necrosis; Pancreatitis; Rats; Rats, Wistar

To study the feature of pancreatic microcirculatory impairment, especially the initial changes, in caerulein-induced experimental acute pancreatitis (AP).. The pancreatic microcirculation of caerulein-induced AP model was studied by intravital fluorescence microscopy with FITC-labeled erythrocytes (FITC-RBC), scanning electron microscopy of vascular corrosion casts, and light microscopy of Chinese ink-injected/cleared tissues.. Animals in caerulein-treated group showed hyperamylemia (X2), pancreatic oedema, infiltration of inflammatory cells in pancreas. Constrictions of intralobular arteriolar sphincters, presence of vacuoles in all layers of sphincter, and gross irregularity in capillary network of acini were found in the AP specimens. The decrease of pancreatic capillary blood flow (0.34+/-0.10 nl x min(-1) vs 0.91+/-0.06 nl x min(-1) of control, P<0.001), reduction of functional capillary density(277+/-13 cm(-1) vs 349+/-8 cm(-1) of control, P<0.001), and irregular intermittent perfusion were observed in caerulein-induced groups.. Impairment and constriction of pancreatic intralobular arteriolar sphincter are the initial microcirculatory lesions in the early phase of acute pancreatitis, and play a key role in the pancreatic ischaemia and pancreatic microvascular failure in acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Arterioles; Ceruletide; Disease Models, Animal; Edema; Erythrocytes; Fluorescein-5-isothiocyanate; Fluorescent Dyes; Male; Microscopy, Electron, Scanning; Microscopy, Fluorescence; Pancreas; Pancreatitis; Rats; Rats, Wistar; Vacuoles

Adenosine shows protective effects against cellular damage and dysfunction under several adverse conditions such as inflammation and ischemia. In the current study, we examined the effects of 3-[1-(6,7-diethoxy-2-morpholinoquinazolin-4-yl)piperidin-4-yl]-1,6-dimethyl-2,4(1,3 )-quinazolinedione hydrochloride (KF24345), an adenosine uptake inhibitor, on cerulein-induced acute pancreatitis in mice to investigate whether inhibition of adenosine uptake could ameliorate the severity of acute pancreatitis.. Acute pancreatitis was induced in mice with six intraperitoneal injections of cerulein (50 microg/kg each) at hourly intervals.. The cerulein injection increased activities of serum amylase and lipase and caused pathologic changes such as interstitial edema, polymorphonuclear cell infiltration, and acinar cell necrosis in the pancreas. KF24345 (10 mg/kg p.o.) ameliorated all these changes observed in mice with acute pancreatitis, and the suppressing effect of KF24345 on the elevation in serum amylase activity was abolished by the treatment with 8-(p-sulfophenyl)theophylline, an adenosine receptor antagonist. In addition, 2-(aminocarbonyl)- -(4-amino-2,6-dichlorophenyl)-4-[5,5-bis-(4-fluorophenyl)pentyl]-1-piperazineacetamide (R75231) and dipyridamole, other adenosine uptake inhibitors, also decreased the elevated serum amylase activity.. These are the first demonstrations that the adenosine uptake inhibitors ameliorate cerulein-induced acute pancreatitis in mice, and these data suggest that adenosine uptake inhibition could ameliorate the severity of acute pancreatitis in vivo.

Topics: Acute Disease; Adenosine; Amylases; Animals; Biological Transport; Ceruletide; Dipyridamole; Female; Lipase; Mice; Mice, Inbred BALB C; Models, Chemical; Pancreatitis; Piperazines; Pyrimidinones; Quinazolines; Theophylline

To explore the changes in hemorheology and expression of neutrophil adhesion molecules CD18 and CD62L in pancreatic microcirculation of Caerulein induced experimental acute pancreatitis (AP).. The Wistar rats (n = 21) were randomized into three groups. The model of AP was established by subcutaneous injection of Caerulein. The changes of apparent viscosity of whole blood were measured by Low- shear 30 rheometer. The expression of adhesion molecules on the surface of neutrophil in duced by shear stress was used with stationary control. CD18 expression was increased on neutrophils treated with shear rate, and andanalyzed using flow cytometry.. Rat treated with Caerulein showed hyperamyleimia (t = 69.029, t = 79.734, P < 0.05). Blood viscosity of two AP groups were significantly elevated (0.512 s(-1): t = 10.725, t = 16.945; 5.96 s(-1): t = 12.781, t = 11.992, P < 0.05). Compared with stationary control, CD18 expression was increased on neutrophil treated with shear rate, and significantly induced with shear rate >/= 94.5 s(-1) (94.5 s(-1): t = 7.403, t = 13.323, t = 16.655; 128.5 s(-1): t = 10.092, t = 28.531, t = 24.563, P < 0.05). The expression of CD62L was less sensitive to low shear rate, and began to be down-regulated significantly when the shear rate >/= 94.5 s(-1) (94.5 s(-1): t = 10.687, t = 19.376, t = 12.848; 128.5 s(-1): t = 26.152, t = 48.402, t = 56.814, P < 0.05).. The changes of apparent viscosity of whole blood, and the effect of fluid shear stress on the expression of neutrophil adhesion molecules CD18, CD62L may play an important role in the pancreatic microcirculatory failure of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Flow Cytometry; Hemorheology; Microcirculation; Neutrophils; Pancreatitis; Rats, Wistar

Many interrelationships exist between the thyroid gland and the gastrointestinal tract. Several past and recent studies have shown that the thyroid gland profoundly influences the structure and function of the exocrine pancreas in the rat. In the present study we investigated the effect of methimazole (METZ), an antithyroid drug, on cerulein induced acute pancreatitis (AP) in rats.. Rats were divided into 3 groups (10-12 weeks age, 200-250 g weight, n: 10). Group B was made hypothyroid with methimazole 5 mg/kg daily for 10 days and the others were untreated euthyroid groups. After 10 days, acute pancreatitis was induced with four doses of 20 microg/kg body weight of cerulein administered s.c at hourly intervals in group A and B while the control group C was given 4 doses of I ml saline. Pancreas wet weight (mg), plasma amylase activity (IU/l) and pancreatic histology were used as endpoints to quantify the severity of the AP.. Plasma tri-iodothyronine (T3) (ng/dl) and thyroxine (T4) (microg/dl) levels were significantly reduced after METZ treatment for 10 days (p < 0.01). METZ pretreatment reduced significantly the cerulein induced increase in pancreatic weight (1,205 +/- 12 mg in METZ treated AP group versus 1,617 +/- 14 mg in AP group, p < 0.05) and the rise in amylase activity (7,078 +/- 816 IU/l in METZ treated AP group versus 8,611 +/- 830 IU/l in AP group p < 0.05).. METZ reduces the severity of cerulein induced AP in rats. This effect might be through its antithyroid property.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Hypothyroidism; Male; Methimazole; Pancreatitis; Rats; Rats, Wistar; Thyroid Hormones

Acute pancreatitis leads to hypoxia caused by vasoconstriction and to activation of lysosomal and digestive enzymes resulting in pancreas autodigestion and damage. This causes activation of leucocytes and increased expression of adhesive molecules enabling margination and adhesion of activated leucocytes to the endothelium. Activated leucocytes are the source of proinflammatory cytokins and oxygen-free radicals which intensify the inflammatory response. The reports indicating that adenosine may prevent activation of the above-mentioned processes in ischaemia prompted us to undertake this study. The study was performed in two stages. The first stage was to evaluate the effects of agonists and antagonists of adenosine receptors on normal pancreas while the second one was to determine the influence of these substances on the development of caerulein-induced acute pancreatitis. During the first stage, the animals were injected intraperitoneally with the substances examined: the A1 receptor antagonist--DPCPX, the A2 receptor agonist--CGS 21680, the A2 receptor antagonist--ZM 241385 and the A3 receptor agonist--IB-MECA and then received intravenous saline. The control animals were subjected only to the 12 h intravenous infusion of 0.15 M NaCl. During the second stage, after the intraperitoneal administration of adenosine receptor agonists and antagonists (as in the first stage), acute pancreatitis was induced with the 12 h intravenous infusion of 5 micrograms/kg/h caerulein. Identical acute pancreatitis was induced in the control animals, however no other substances were administered. The pancreatic tissue samples were collected directly after intravenous infusion. The severity of inflammatory processes in the pancreas was evaluated on the basis of the plasma amylase activity, pancreatic weight and enhancement of histopathological changes observed in this organ. In the animals infused with saline alone, no effects of the substances examined on the pancreatic weight, plasma amylase activity and histopathological features were observed. The intravenous caerulein infusion induced acute pancreatitis expressed as bigger pancreatic weight, increased plasma amylase activity and tissue damage (oedema, cell vacuolization, leucocyte infiltration). The A2 receptor agonist administration preceding the induction of acute pancreatitis decreased the pancreas damage caused by caerulein. Lower weight of the pancreas and decreased plasma amylase activities were observed; on hist

Topics: Acute Disease; Adenosine; Animals; Ceruletide; Male; Pancreatitis; Rats; Rats, Wistar; Receptors, Purinergic P1

Prior thermal stress induces heat shock protein 70 (HSP70) expression in the pancreas and protects against secretagogue-induced pancreatitis, but it is not clear that this thermal stress-induced protection is actually mediated by HSP70 since thermal stress may have other, non-HSP related, effects.. In the present study, we have administered antisense (AS) oligonucleotides, which prevent pancreatic expression of HSP70 to rats, in vivo, to evaluate this issue. In a separate series of experiments, designed to examine the role of pancreatitis-induced HSP70 expression in modulating the severity of pancreatitis, rats not subjected to prior thermal stress were given AS-HSP70 before cerulein administration, and trypsinogen activation as well as the severity of pancreatitis were evaluated.. Hyperthermia induced HSP70 expression, prevented intrapancreatic trypsinogen activation, and protected against cerulein-induced pancreatitis. Administration of AS-HSP70 but not sense-HSP70 reduced the thermal stress-induced HSP70 expression, restored the ability of supramaximal cerulein stimulation to cause intrapancreatic trypsinogen activation, and abolished the protective effect of prior thermal stress against pancreatitis. In non-thermally stressed animals, pretreatment with AS-HSP70 before the induction of pancreatitis exacerbated all the parameters associated with pancreatitis.. These findings lead us to conclude that HSP70 induction, rather than some other thermal stress-related phenomenon, mediates the thermal stress-induced protection against pancreatitis and that it protects against pancreatitis by preventing intrapancreatic activation of trypsinogen. The worsening of pancreatitis, which occurs when non-thermally stressed animals are given AS-HSP70 before cerulein, suggests that cerulein-induced HSP70 expression in nontreated animals acts to limit the severity of pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Gene Expression Regulation; Heat Stress Disorders; HSP70 Heat-Shock Proteins; Oligonucleotides, Antisense; Pancreatitis; Rats; Rats, Wistar; Stress, Physiological; Trypsinogen

A premature and intracellular activation of digestive zymogens is thought to be responsible for the onset of pancreatitis. Because trypsin has a critical role in initiating the activation cascade of digestive enzymes in the gut, it has been assumed that trypsin also initiates intracellular zymogen activation in the pancreas. We have tested this hypothesis in isolated acini and lobules from rat pancreas. Intracellular trypsinogen activation was induced by supramaximal secretagogue stimulation and measured using either specific trypsin substrates or immunoreactivity of the trypsinogen activation peptide (TAP). To prevent a trypsin-induced trypsinogen activation, we used the cell-permeant, highly specific, and reversible inhibitor Nalpha-(2-naphthylsulfonyl)-3-amidinophenylalanine-carboxymethylpiperazide (S124), and to prevent cathepsin-induced trypsinogen activation, we used the cysteine protease inhibitor E-64d. Incubation of acini or lobules in the presence of S124 completely prevented the generation of trypsin activity in response to supramaximal caerulein but had no effect whatsoever on the generation of TAP. Conversely, when trypsin activity was recovered at the end of the experiment by either washout of S124 from acini or extensive dilution of lobule homogenates, it was up to 400% higher than after caerulein alone and corresponded, in molar terms, to the generation of TAP. Both trypsin activity and TAP release were inhibited in parallel by E-64d. We conclude that caerulein-induced trypsinogen activation in the pancreas is caused by an E-64d-inhibitable mechanism such as cathepsin-induced trypsinogen activation, and neither involves nor requires intracellular trypsin activity. Specific trypsin inhibition, on the other hand, prevents 80% of trypsin inactivation or autodegradation in the pancreas.

Topics: Acute Disease; Animals; Cathepsin B; Ceruletide; Enzyme Activation; Male; Naphthalenes; Pancreas; Pancreatitis; Piperazines; Rats; Rats, Wistar; Trypsin; Trypsin Inhibitors; Trypsinogen

Activation of zymogens within the pancreatic acinar cell is an early feature of acute pancreatitis. Supraphysiological concentrations of cholecystokinin (CCK) cause zymogen activation and pancreatitis. The effects of the CCK analog, caerulein, and alcohol on trypsin and chymotrypsin activation in isolated pancreatic acini were examined. Caerulein increased markers of zymogen activation in a time- and concentration-dependent manner. Notably, trypsin activity reached a peak value within 30 min, then diminished with time, whereas chymotrypsin activity increased with time. Ethanol (35 mM) sensitized the acinar cells to the effects of caerulein (10(-10) to 10(-7) M) on zymogen activation but had no effect alone. The effects of ethanol were concentration dependent. Alcohols with a chain length of >or=2 also sensitized the acinar cell to caerulein; the most potent was butanol. Branched alcohols (2-propanol and 2-butanol) were less potent than aliphatic alcohols (1-propanol and 1-butanol). The structure of an alcohol is related to its ability to sensitize acinar cells to the effects of caerulein on zymogen activation.

Topics: 2-Propanol; Acute Disease; Alcohols; Animals; Butanols; Ceruletide; Chymotrypsin; Enzyme Activation; Enzyme Precursors; Ethanol; Kinetics; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Trypsin

To compare the effect of different vasoactive mediator antagonists in the same model of severe acute pancreatitis (AP) and to evaluate whether combinations of the agents exhibit synergistic effects.. Prospective experimental study.. Microcirculation and pancreas research laboratory at an university hospital.. Hundred eighty anesthetized male Sprague-Dawley rats.. Six hours after inducing AP by intra-ductal bile salt infusion and i.v. cerulein in 168 rats, these were randomized for therapy with (1) saline, (2) endothelin receptor antagonist (ET-RA), (3) platelet activating factor receptor antagonist (PAF-RA), (4) intercellular adhesion molecule-1 antibody (ICAM-1-AB) or different combinations (5-7). After 24 h the animals underwent a second laparotomy for intra-vital microscopic determination of pancreatic and colonic capillary permeability, blood flow and leukocyte-endothelial interaction.. AP induction decreased capillary blood flow and increased permeability and leukocyte rolling. ET-RA, PAF-RA and ICAM-1-AB decreased capillary permeability, increased blood flow and reduced leukocyte rolling. ET-RA was most effective in decreasing capillary permeability in both organs as well as in increasing pancreatic capillary blood flow. Combining vasoactive mediator blockers did not further improve target parameters.. This study supports previous observations that ET-RA, PAF-RA and ICAM-1-AB improve microcirculation in AP and that ET-RA is more effective than PAF-RA or ICAM-1-AB, especially in counteracting capillary leakage. Although this may suggest that they act through different mechanisms, antagonist combinations failed to improve microcirculation further. We conclude that ET-RA is the most promising candidate for a clinical trial to reduce capillary leakage in patients with AP.

Topics: Acute Disease; Animals; Antibodies; Bile Acids and Salts; Capillary Leak Syndrome; Ceruletide; Endothelin Receptor Antagonists; Intercellular Adhesion Molecule-1; Male; Microcirculation; Pancreatitis; Platelet Membrane Glycoproteins; Prospective Studies; Rats; Rats, Sprague-Dawley; Receptors, Cell Surface; Receptors, G-Protein-Coupled; Vascular Diseases

Lipopolysaccharides (LPS) are responsible for septic shock but low doses of LPS reduce pancreatic damage produced by caerulein-induced pancreatitis (CIP) in rats. Leptin, produced by adipocytes attenuates the severity of CIP. The aim of this study was to evaluate the effect of intracerebroventricular (i.c.v.) administration of LPS on CIP and plasma leptin level and to investigate the involvement of sensory nerves (SN) in the effects of LPS on CIP.. CIP was produced by subcutaneous (s.c.) infusion of caerulein (25 Kg/kg) to conscious rats. SN were deactivated with capsaicin (100 mg/kg s.c.). LPS (0.2, 2, or 20 Kg/rat) were applied to the right cerebral ventricle 30 min prior to CIP.. CIP was manifested by an increase in plasma levels of amylase, lipase, leptin and an anti-inflammatory interleukin 10 (IL-10), (by 400%, 1000%, 700% and 50%, respectively), confirmed by histological examination and accompanied by 40% reduction in pancreatic blood flow. Pretreatment of CIP rats with i.c.v. LPS resulted in significant reduction of CIP accompanied by dose-dependent increase in plasma levels of leptin and IL-10. Deactivation of SN, which by itself failed to affect CIP, completely reversed the beneficial effects of i.c.v. administration of LPS on CIP and reduced plasma leptin and IL-10 concentrations.. Pretreatment with LPS given i.c.v. prevents the development of caerulein-induced pancreatitis through the activation of SN and though the release of leptin.

Topics: Acute Disease; Adipocytes; Afferent Pathways; Amylases; Animals; Capsaicin; Ceruletide; Escherichia coli; Injections, Intraventricular; Interleukin-10; Leptin; Lipase; Lipopolysaccharides; Neurons, Afferent; Pancreatitis; Rats; Rats, Wistar; Sympathectomy, Chemical

We studied the effect of iron chelators, 2,2'-dipyridyl and desferrioxamine, on cerulein-induced pancreatitis in rats. Acute pancreatitis was induced by a single subcutaneous injection of 100 micrograms/kg body weight of cerulein, which caused hyperamylasemia and edematous pancreatitis with neutrophilic infiltration. Blood samples were collected for determination of serum amylase values and the pancreas was removed for the histological examination 6 h after the cerulein injection. Intraperitoneal administration of a ferrous iron chelator, 2,2'-dipyridyl, prior to the cerulein injection resulted in amelioration of hyperamylasemia and histological abnormalities such as edema and inflammation but not of acinar cell vacuolization. In contrast, administration of a ferric iron chelator, desferrioxamine, did not show any beneficial effects. These results indicate that administration of 2,2'-dipyridyl ameliorates the pancreatitis induced by the supramaximal dose of cerulein.

Topics: 2,2'-Dipyridyl; Acute Disease; Animals; Ceruletide; Deferoxamine; Edema; Injections, Intraperitoneal; Iron Chelating Agents; Male; Neutrophil Infiltration; Pancreatitis; Rats; Rats, Wistar

To explore the pathogenesis of acute pancreatitis (AP) and evaluate the effect of a Chinese herb WPY on the course of AP.. Intravital fluorescence microscopy was used. The pancreatic microvascular diameter, RBC velocities and functional capillary density (FCD) were estimated following intracutaneous injection of caerulein alone or with gastropipe WPY.. Caerulein mediated a significant decrease in functional capillary density (FCD), RBC velocities and diameter of interlobular arteriole (P < 0.01). Compared with AP group, WPY was effective in restoring functional capillary density, interlobular arteriole diameter and RBC velocity.. Impairment of pancreatic microcirculation in the early phase of acute pancreatitis may play a key role in the progression of this disease. Possible contributory mechanisms include reduced blood flow and functional capillary density, interlobular arteriole spasm, and leukocyte-endothelial cell interaction. WPY has a beneficial effect on the course of acute pancreatitis. Possible causes include attenuating microcirculatory failure.

Topics: Acute Disease; Animals; Blood Flow Velocity; Ceruletide; Drug Combinations; Drugs, Chinese Herbal; Mice; Microcirculation; Microscopy, Fluorescence; Pancreas; Pancreatitis

We demonstrated that the dynamic aspects of cytokine production in rat acute pancreatitis, which was induced by cerulein and aggravated by subsequent lipopolysaccharide (LPS) injection. A priming effect by induction of mild pancreatitis with cerulein enhanced the subsequent cytokine production by LPS injection. Alternatively, after induction of severe pancreatitis with cerulein and LPS, cytokine production was markedly suppressed for > or = 90 hours. Production of interleukin-2 (IL-2) by splenocytes decreased, and mortality rate after cecal ligation and puncture (CLP) increased significantly after induction of severe acute pancreatitis. These results suggest that the suppression of a cytokine response in severe acute pancreatitis may alter the defense system and, as a result, increase mortality after CLP.

Topics: Acute Disease; Animals; Cells, Cultured; Ceruletide; Cytokines; Endotoxemia; Escherichia coli; Lipopolysaccharides; Male; Pancreatitis; Peritonitis; Rats; Rats, Wistar; Spleen

Reactive oxygen species have been implicated in the pathogenesis of acute pancreatitis. Few studies have focused on the loss of endogenous antioxidants and molecular oxidative damage. Two acute pancreatitis models in rats; taurocholate (3% intraductal infusion) and cerulein (10 microg/kg/h), were used to study markers of oxidative stress: Glutathione, ascorbic acid, and their oxidized forms (glutathione disulfide and dehydroascorbic acid), malondialdehyde, and 4-hydroxynoneal in plasma and pancreas, as well as 7-hydro-8-oxo-2'-deoxyguanosine in pancreas. In both models, pancreatic glutathione depleted by 36-46% and pancreatic ascorbic acid depleted by 36-40% (p <.05). In the taurocholate model, plasma glutathione was depleted by 34% (p <.05), but there were no significant changes in plasma ascorbic acid or in plasma and pancreas dehydroascorbic acid, malondialdehyde, and 4-hydroxynoneal, and no significant changes in the pancreas glutathione disulfide/glutathione ratio. While pancreas glutathione disulfide/glutathione ratio increased in the cerulein model, there were no significant changes in plasma glutathione, plasma, or pancreas ascorbic acid, dehydroascorbic acid, 4-hydroxynoneal, and malondialdehyde, or in pancreas 7-hydro-8-oxo-2'-deoxyguanosine. Reactive oxygen species have a minor role in the intermediate stages of pancreatitis models.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acute Disease; Aldehydes; Animals; Ascorbic Acid; Biomarkers; Ceruletide; Dehydroascorbic Acid; Deoxyguanosine; Glutathione; Glutathione Disulfide; Male; Malondialdehyde; Oxidation-Reduction; Oxidative Stress; Pancreatitis; Rats; Rats, Wistar; Reactive Oxygen Species; Taurocholic Acid

It is well known that endotoxemia, which is caused by a bacterial infection, can exacerbate acute pancreatitis, whereas pancreatitis-associated protein (PAP) has the ability to induce bacterial aggregation. Pancreatitis-associated protein is supposed to protect the tissue from infection during inflammation. In order to clarify the relationship between PAP mRNA expression and endotoxemia during acute pancreatitis, the kinetic patterns of PAP-I mRNA in mouse pancreas treated with either cerulein or lipopolysaccharide (LPS) or both were investigated in this study.. The administration of LPS (5 mg/kg) intraperitoneally resulted in a dramatic upregulation of PAP-I mRNA expression, increasing 18.61-fold to a maximum at 12 h, then decreasing, but still sustaining at a high level and reaching baseline on day five. These changes were accompanied by the upregulation of tumor necrosis factor (TNF)-alpha, interleukin-1beta (IL-1beta), interleukin 6 (IL-6) and interferon gamma (IFNgamma) mRNA expressions in the pancreas, but not by marked alterations of serum amylase, lactic dehydrogenase (LDH) and histology. Cerulein also increased PAP-I mRNA expression. However, the combination of cerulein and LPS was not able to enhance PAP-I mRNA expression further, although more prominent pancreatitis based on significant changes of serum amylase, LDH and histology were observed.. These results suggest that PAP-I mRNA might be modulated by endotoxemia, independent of cerulein-pancreatitis. There were no strong correlations between PAP-I mRNA expression and the severity of pancreatitis.

Topics: Acute Disease; Acute-Phase Proteins; Animals; Antigens, Neoplasm; Biomarkers, Tumor; Blotting, Northern; Ceruletide; Cytokines; Immunohistochemistry; Lectins, C-Type; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Pancreatitis; Pancreatitis-Associated Proteins; Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Up-Regulation

Endothelin-1 has been shown to reduce pancreatic blood flow and cause focal acinar cell necrosis similar to those seen in acute pancreatitis (AP), whereas therapy with endothelin receptor antagonists enhanced pancreatic capillary blood flow (PCBF) and decreased mortality rates. The current study evaluated the role of endothelin in the development of severe AP. Trypsinogen activation peptides, acinar cell necrosis, and PCBF were used as local indicators of disease severity, fluid sequestration, cardiorespiratory and renal parameters, and colonic capillary blood flow as systemic disease indicators. The following groups of animals were examined: 1) rats with mild edematous AP and 2) severe necrotizing AP treated with and without endothelin, 3) transgenic rats overexpressing endothelin with severe AP, and 4) rats with severe AP prophylactically treated with endothelin receptor antagonists. The following observations were made: endothelin superimposed on mild AP caused hemoconcentration, a decrease in PCBF, and necrosis and ascites not seen in this model without endothelin exposure. Endothelin superimposed on severe AP had no significant effects. After induction of severe AP, less PCBF and more acinar cell necrosis were observed in transgenic rats than in their normal littermates. Prophylactic endothelin receptor antagonists improved local (acinar necrosis, PCBF) and systemic parameters (ascites, urine production, colonic capillary blood flow) of disease severity in animals with severe AP. These observations underscore the role of endothelin as a mediator of disease severity in AP and suggest that endothelin receptor blockade may become a promising therapeutic tool in this disease.

Topics: Acute Disease; Animals; Animals, Genetically Modified; Blood Pressure; Capillaries; Ceruletide; Edema; Endothelin-1; Gene Expression; Hematocrit; Male; Pancreas; Pancreatitis; Pancreatitis, Acute Necrotizing; Rats; Rats, Sprague-Dawley

Complement factor C5a acting via C5a receptors (C5aR) is recognized as an anaphylotoxin and chemoattractant that exerts proinflammatory effects in many pathological states. The effects of C5a and C5aR in acute pancreatitis and in pancreatitis-associated lung injury were evaluated using genetically altered mice that either lack C5aR or do not express C5. Pancreatitis was induced by administration of 12 hourly injections of cerulein (50 microg/kg ip). The severity of pancreatitis was determined by measuring serum amylase, neutrophil sequestration in the pancreas, and acinar cell necrosis. The severity of lung injury was evaluated by measuring neutrophil sequestration in the lung and pulmonary microvascular permeability. In both strains of genetically altered mice, the severity of pancreatitis and pancreatitis-associated lung injury was greater than that noted in the comparison wild-type strains of C5aR- and C5-sufficient animals. This exacerbation of injury in the absence of C5a function indicates that, in pancreatitis, C5a exerts an anti-inflammatory effect. Potentially, C5a and its receptor are capable of both promoting and reducing the extent of acute inflammation.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents; Antigens, CD; Capillaries; Ceruletide; Complement C5a; Crosses, Genetic; Inflammation; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreatitis; Peroxidase; Receptor, Anaphylatoxin C5a; Receptors, Complement

Prostaglandins (PG), the products of arachidonate metabolism through cyclooxygenase (COX) pathway, protect the pancreas from the acute damage. The existence of two isoforms of COX was documented including: COX-1, present in normal tissues and COX-2, expressed at the site of inflammation, such as induced by bacterial lipopolysaccharide (LPS). Pretreatment with low dose of LPS and activation of nitric oxide (NO) synthase (NOS) has been shown to prevent the injury caused by caerulein-induced pancreatitis (CIP) in the rat. The aim of this study was to investigate the role of COX-1 and COX-2 in the LPS-induced protection of the pancreas against CIP and the involvement of NOS in the activation of COX-PG system in the rats with CIP. CIP was produced by subcutaneous (s.c.) infusion of caerulein (5 microg/kg-h for 5 h) to the conscious rats. Protective dose of LPS, from Escherichia coli, (1 mg/kg) was given intraperitoneally (i.p.) 15 min prior to the start of CIP. Nonselective inhibitor of COX; indomethacin (5 or 10 mg/kg), selective inhibitor of COX-1: resveratrol, or a highly selective inhibitors of COX-2: rofecoxib or NS-398 (2 or 10 mg/kg) were injected i.p. 15 min prior to the administration of LPS. COX-1 or COX-2 mRNA was determined by reverse transcription-polimerase chain reaction (RT-PCR) in the pancreatic tissue. Pancreatic blood flow (PBF) was measured by a laser Doppler flowmetry. PGE2 content in the pancreas was measured by radioimmunoassay. CIP was manifested by an increase of pancreatic weight and plasma amylase activity (by 500% and 700%, respectively) and it was confirmed by histological examination. CIP slightly increased pancreatic PGE2 generation (by 12%) and diminished PBF (by about 40%). LPS (1 mg/kg i.p.), given prior to the start of CIP, increased PGE2 generation in the pancreas (by 45%), reversed the histological manifestations of pancreatitis, reduced the rise in amylase blood level and improved PBF. Administration of nonselective inhibitor of COX; indomethacin (5 or 10 mg/kg i.p.) prior to the injection of LPS abolished its protective effects on CIP and reduced pancreatic PGE2 generation. Selective inhibitor of COX-1; resveratrol (10 mg/kg i.p.) given prior to the injection of LPS reversed its protective effects against CIP. Pretreatment with a selective inhibitors of COX-2: rofecoxib or NS-398 (10 mg/kg) attenuated LPS-induced pancreatic protection in the CIP rats. COX-1 expression was detected in the intact pancreas and was not signif

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents, Non-Steroidal; Ceruletide; Cyclooxygenase 1; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Cyclooxygenase Inhibitors; Dinoprostone; Enzyme Inhibitors; Indomethacin; Isoenzymes; Lactones; Lipopolysaccharides; Male; Membrane Proteins; Nitric Oxide; Nitric Oxide Donors; Nitroarginine; Nitrobenzenes; Organ Size; Pancreas; Pancreatitis; Penicillamine; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Wistar; Resveratrol; S-Nitroso-N-Acetylpenicillamine; Stilbenes; Sulfonamides; Sulfones

The molecular mechanisms that lead from acute pancreatitis (AP) to multiple organ failure remain to be clarified. We previously reported that ascitic fluids from a rat model of severe acute pancreatitis (pancreatitis-associated ascitic fluids, PAAF) transcriptionally activated endothelial cells and leukocytes in vitro. To clarify the role of ascitic fluids on the development of multiple organ failure in AP, we examined the effects of PAAF on the prognosis and immunohistologic findings in cerulein pancreatitis, an experimental model of mild pancreatitis in vivo. Intraperitoneal injection of PAAF decreased the survival rates in a dose-dependent manner. Histologically, destruction of vessels, alveolar septal thickening, interstitial hypertrophy, and infiltration of inflammatory cells were prominent in the lung of PAAF-injected rats. Transcription factor, nuclear factor KB (NF-kappaB) was activated and the mRNA levels of tumor necrosis factor-alpha and interleukin-1beta were increased in the lung of the PAAF-injected rats. The permeability index assessed by Evans blue assay and the lung myeloperoxidase activity levels were significantly higher in the PAAF-injected rats than in controls. Inhibition of NF-kappaB ameliorated the histologic findings and improved the survival rates. Our results suggest that PAAF play a role in the pathogenesis of lung injury in severe AP, at least in part through the activation of NF-kappaB.

Topics: Acute Disease; Animals; Ascites; Ascitic Fluid; Ceruletide; Disease Models, Animal; Evans Blue; Immunohistochemistry; Interleukin-1; Lung; Lung Diseases; Male; NF-kappa B; Pancreatitis; Peroxidase; Prognosis; Rats; Rats, Wistar; RNA, Messenger; Survival Rate; Tumor Necrosis Factor-alpha

Although the pancreatic heat shock response has already been reported to confer protective effects during experimental pancreatitis, the mechanism of action remains unknown. We investigated the effects of hyperthermia in cerulein-induced pancreatitis. Heat shock protein 70 (HSP70) expression in rats was induced by a 20-min period of water immersion (42 degrees C). The severity of pancreatitis as well as the pancreatic expression of cytokines, nuclear factor-kappaB (NF-kappaB), and inhibitory factor kappaB-alpha (IkappaB-alpha) were evaluated in the presence and absence of hyperthermia. We found that hyperthermia resulted in time-dependent expression of HSP70 within the pancreas associated with a reduction in the severity of acute pancreatitis. Tumor necrosis factor-alpha and intercellular adhesion molecule-1 expression was significantly reduced in the presence of hyperthermia. Moreover, NF-kappaB activity was delayed in the presence of hyperthermia whereas IkappaB-alpha was stabilized in the cytoplasm. These results suggest that hyperthermia decreases the severity of cerulein-induced pancreatitis by decreasing cytokine expression in the pancreas through the modulation of NF-kappaB activity.

Topics: Acute Disease; Animals; Ceruletide; Fever; HSP70 Heat-Shock Proteins; Intercellular Adhesion Molecule-1; Male; NF-kappa B; Osmolar Concentration; Pancreatitis; Rats; Rats, Wistar; Reference Values; Tumor Necrosis Factor-alpha

Taurine, or 2-aminoethane sulfonic acid, is an intracellular amino acid and has been suggested to have a function in protecting biological systems from oxidative tissue damage. The aim of this study was to determine the effect of taurine against cerulein-induced acute pancreatitis in rats. Acute pancreatitis was induced by administering three subcutaneous injections of cerulein (40 microg/kg body weight) at 1-hour intervals, while taurine was administered intravenously at graded doses (30, 100, or 300 mg/kg, respectively) following the first cerulein injection. The severities of pancreatitis and lung injury were determined by measuring biochemical parameters, tissue myeloperoxidase (MPO), and histological changes. To clarify the mechanism of taurine, serum IL-1beta and TNF-alpha levels and tissue concentrations of malondialdehyde (MDA) were evaluated. In cerulein-induced acute edematous pancreatitis, treatment with taurine significantly decreased hyperamylasemia, tissue MPO, pancreatic edema, and the extent of pancreatic and pulmonary injury. Taurine decreased MDA concentration in the pancreas and lung, but not the serum cytokine concentration. We would conclude that taurine has beneficial effects in cerulein-induced acute pancreatitis and lung injuries by preventing the production of oxygen free radicals.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Interleukin-1; Lung Diseases; Male; Malondialdehyde; Organ Size; Oxidative Stress; Pancreas; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley; Taurine; Tumor Necrosis Factor-alpha

To investigate the effects of somatostatin analogs (SSa) on apoptosis of pancreatic acinar cells and apoptosis-regulated gene bax, and p53 in treating acute pancreatitis in mice.. In cerulein-induced pancreatitis, with or without treatment of somatostatin, analogs (Octreotide) in CD-1 (BALB/c x DBetaAlpha/1) mice, apoptosis of pancreatic acinar cells was detected by using the TdT-mediated dUTP nick-end labeling (TUNEL) method, and the expression of apoptosis-regulated gene bax and p53 was determined by using the streptavidin-peroxidase immunohistochemical technique and the RT-PCR method, respectively.. On HE staining, acinar cells in the pancreas showed pyknotic nuclei and the formation of apoptotic bodies, which are the typical morphological features of apoptosis. Regarding TUNEL use, the apoptotic index of pancreatic acinar cells in the non-treated group at 5 and 14 h after induction of acute pancreatitis was significantly lower than those of the SSa-treated group, respectively (P < 0.01). On immunohistochemistry and RT-PCR, there was an expression of neither bax nor p53 in normal pancreatic tissues. The expression of bax in the SSa-treated group at 5 and 14 h after treatment of SSa was markedly higher than those of the non-treated group, respectively (P < 0.01), but there was no significant difference in the expression of p53 between the SSa-treated group and the non-treated group.. The induction of apoptosis in pancreatic acinar cells injury to reduce inflammatory reaction might be one of the mechanisms of SSa in treating acute pancreatitis in mice, and the mechanisms of apoptosis probably correlated with the expression of apoptosis-regulated gene bax, but have no relationship with the expression of p53.

Topics: Acute Disease; Animals; Apoptosis; bcl-2-Associated X Protein; Ceruletide; Female; Genes, p53; Hormones; Immunohistochemistry; In Situ Nick-End Labeling; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Octreotide; Pancreatitis; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Reverse Transcriptase Polymerase Chain Reaction; Somatostatin

The interactions between inflammatory cells and their mediators play important roles in many inflammatory processes, but their importance during acute experimental pancreatitis and pancreatitis-associated lung injury is unclear. To address the role of the interaction between CD40 and its ligand CD40L, molecules that mediate major immunoregulatory functions, pancreatitis was induced by administering supramaximal doses of cerulein in mice that do not express CD40L.. The severity of pancreatitis was measured by serum amylase activity, pancreatic edema, acinar cell necrosis, and pancreas myeloperoxidase activity (an indicator of neutrophil infiltration). Lung injury was quantitated by evaluating lung microvascular permeability and lung myeloperoxidase activity.. In pancreatic tissue from control mice and cerulein-treated mice, the expression of both CD40 and CD40L was detected. Immunohistochemical analysis performed in isolated acini from wild-type pancreata showed that both CD40 and CD40L were expressed on the acinar cell surface. Interestingly, pancreatitis and pancreatitis-associated lung injury were markedly decreased in mice deficient in CD40L compared with wild-types.. These observations indicate that CD40L plays an important proinflammatory role in pancreatitis and pancreatitis-associated lung injury.

Topics: Acute Disease; Amylases; Animals; CD40 Ligand; Ceruletide; Lung Diseases; Male; Mice; Pancreatitis; Tumor Necrosis Factor-alpha

In experimental models of acute pancreatitis (AP), acinar cell death occurs by both necrosis and programmed cell death or apoptosis. Apoptosis is an active form of cell death associated with a tightly regulated expression of gene products that are either pro- or antiapoptotic. The aim of this study was to characterize pancreatic mRNA levels by Northern blotting analysis of apoptosis-associated genes used during the course of cerulein-induced AP in mice. Histone H3 mRNA levels were also examined as an indicator of cell proliferation. Acinar cell apoptosis was confirmed histologically. The findings show that AP modifies pancreatic mRNA levels of both pro- and antiapoptotic genes simultaneously. Pancreatic bclXL, bax, and p53 mRNA levels increased significantly in a temporal fashion during induction of AP. Pancreatic bcl-2 mRNA levels were unchanged during AP. Pancreatic mRNA levels of insulin-like growth factor-1 (IGF-1), a mitogen and cell survival factor, and its receptor (IGF-1R) also increased in a temporal fashion during induction of AP. In summary, this study indicates that acinar cell death during cerulein-induced AP in mice can occur by the apoptotic pathway. Since factors promoting and antagonistic for cell survival are activated simultaneously, regulation of acinar cell survival appears complex and dynamic during AP.

Topics: Acute Disease; Amylases; Animals; Apoptosis; bcl-2-Associated X Protein; Blotting, Northern; Blotting, Western; Cell Survival; Ceruletide; Female; Genes, p53; Histones; Insulin-Like Growth Factor I; Mice; Pancreatitis; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; RNA, Messenger

Acute severe (necrotizing) pancreatitis is often associated with pancreatic or peripancreatic infection. Decreased bacterial clearance due to impaired immune defense may cause local infection. We investigated expressions of surface opsonin receptors (CD11b, complement receptor 3; CD32/CD16, immunoglobulin G Fc receptor) on local and circulatory neutrophils, in murine acute pancreatitis. The mild and severe forms of acute pancreatitis were induced by seven and 13 subcutaneous injections of caerulein, respectively. Peritoneal exudative and circulatory neutrophils were counted and assayed for receptor expressions by flow cytometry, serially at 1-72 hours after pancreatitis induction. Histologically, mild and severe forms showed edematous and necrotizing pancreatitis, respectively. The peritoneal exudative neutrophil count was greater in mild than in severe pancreatitis. Expressions of CD11b and CD32/CD16 on local neutrophils were upregulated early in mild pancreatitis. This upregulation was attenuated in severe pancreatitis. The circulatory neutrophil count was elevated in severe pancreatitis but was unchanged in mild pancreatitis. Opsonin receptor expression on circulatory neutrophils showed a transient, modest upregulation in the early phase of mild pancreatitis. Receptor-positive circulatory neutrophils showed a marked elevation that persisted throughout the course of severe pancreatitis. In conclusion, severe (necrotizing) pancreatitis is associated with reduced opsonin receptor expression on local neutrophils and enhanced expression on circulatory neutrophils, as compared with mild (edematous) pancreatitis. These changes may contribute to local infectious complications and multiple organ failure, in severe pancreatitis.

Topics: Acute Disease; Animals; Ascitic Fluid; Ceruletide; Complement Activation; Disease Progression; Drug Administration Schedule; Edema; Female; Leukocyte Count; Macrophage-1 Antigen; Mice; Mice, Inbred BALB C; Neutrophils; Pancreatitis; Pancreatitis, Acute Necrotizing; Phagocytosis; Receptors, Fc; Receptors, IgG; Receptors, Immunologic

Production of nitric oxide (NO) by inducible nitric oxide synthase (iNOS) has been proposed as a pathogenic factor in acute pancreatitis, but its role has still not been fully examined. The present study explored the role of iNOS in cerulein-induced acute pancreatitis using iNOS-deficient mice. Twelve- to 14-week-old male mice (C57B1/6 and iNOS-deficient) were administered cerulein by intraperitoneal (i.p.) injection at hourly intervals for 7 hours and killed 24 hours later after the first dose. Pancreatic wet weight, pancreatic myeloperoxidase (MPO) activity, and levels of plasma nitrite and serum amylase were measured. In another experiment isosorbide dinitrate (an NO donor) was given by oral gavage every 6 hours for 24 hours beginning simultaneously with cerulein injections in iNOS-deficient mice. Cerulein administration dose-dependently increased pancreatic wet weight, myeloperoxidase activity, and levels of nitrite and amylase in C57B1/6 mice. These parameters (except nitrite levels) were significantly intensified in iNOS-deficient mice. At the dose employed, cerulein failed to increase nitrite levels in iNOS-deficient mice. The susceptibility to cerulein toxicity in iNOS-deficient mice was abolished by NO donor treatment. NO release from an iNOS source appears to play a protective role in cerulein-induced pancreatitis. At least in part, NO may prevent neutrophil accumulation after cerulein administration.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Genetic Predisposition to Disease; Injections, Intraperitoneal; Isosorbide Dinitrate; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Nitric Oxide Donors; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Pancreatitis; Peroxidase

An in vivo microscopic technique was used to clarify the increase in microvascular permeability and enhanced leukocyte-endothelium interaction of pancreatic microcirculation in experimental pancreatitis of differing severity. Using bovine albumin fluorescein isothiocyanate (FITC) and carboxyfluorescein diacetate succinimidyl ester (CFDASE) as tracers, the change in permeability and the behavior of leukocytes in the acinar microcirculation were quantified during the initial 1, 2, 6, and 12h after the induction of caerulein pancreatitis in mice. Cold stress was added to produce the severe model. It was revealed that the early microcirculatory changes in the pancreas of caerulein pancreatitis included the increased permeability of endothelial lining and an accumulation of extravasated fluid in the perilobular space, which were more severe if cold stress was added. A decrease in flow velocity was also noted 2h after the onset of severe pancreatitis. Leukocyte adherence to the endothelial cells was not observed during the first 12h in either model of severity. In contrast, observation of the hepatic microcirculation revealed a significant number of adherent leukocytes 2h after the induction of severe pancreatitis. These results suggest that during the early course of acute pancreatitis, leukocyte adherence in the pancreatic microcirculation is a secondary event following the increase in pancreatic vascular permeability.

Topics: Acute Disease; Amylases; Animals; Aspartate Aminotransferases; Blood Flow Velocity; Capillary Permeability; Cell Adhesion; Ceruletide; Disease Models, Animal; Endothelium, Vascular; Hemorheology; Leukocytes; Lipase; Male; Mice; Mice, Inbred ICR; Microcirculation; Microscopy, Fluorescence; Pancreas; Pancreatitis; Signal Processing, Computer-Assisted; Time Factors

Intra-acinar cell activation of digestive enzyme zymogens including trypsinogen is generally believed to be an early and critical event in acute pancreatitis. We have found that the phosphatidylinositol 3-kinase inhibitor wortmannin can reduce the intrapancreatic activation of trypsinogen that occurs during two dissimilar experimental models of rodent acute pancreatitis, secretagogue- and duct injection-induced pancreatitis. The severity of both models was also reduced by wortmannin administration. In contrast, the NF-kappa B activation that occurs during the early stages of secretagogue-induced pancreatitis is not altered by administration of wortmannin. Ex vivo, caerulein-induced trypsinogen activation is inhibited by wortmannin and LY294002. However, the cytoskeletal changes induced by caerulein were not affected by wortmannin. Concentrations of caerulein that induced ex vivo trypsinogen activation do not significantly increase phosphatidylinositol-3,4-bisphosphate or phosphatidylinositol 3,4,5-trisphosphate levels or induce phosphorylation of Akt/PKB, suggesting that class I phosphatidylinositol 3-kinases are not involved. The concentration of wortmannin that inhibits trypsinogen activation causes a 75% decrease in phosphatidylinositol 3-phosphate, which is implicated in vesicle trafficking and fusion. We conclude that a wortmannin-inhibitable phosphatidylinositol 3-kinase is necessary for intrapancreatic activation of trypsinogen and regulating the severity of acute pancreatitis. Our observations suggest that phosphatidylinositol 3-kinase inhibition might be of benefit in preventing acute pancreatitis.

Topics: Acute Disease; Androstadienes; Animals; Cells, Cultured; Ceruletide; Chromones; Cytoskeleton; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Inhibitors; Lysosomes; Male; Mice; Morpholines; Necrosis; NF-kappa B; Pancreatitis; Phosphatidylinositol 3-Kinases; Phosphatidylinositol Phosphates; Phosphorylation; Rats; Time Factors; Trypsinogen; Wortmannin

External perfusions of the pancreatic glands of Wistar-rats were done, using a modified Krebs-Ringer-Buffer (KRB). We looked for an elevation of amylase, lipase and lactate-dehydrogenase in the effusion fluid (portal outflow fluid). We investigated a normal perfusion (KRB, 60 minutes), a long term perfusion (KRB, 240 minutes) and a perfusion (60 minutes) including an additive of the detergents triton x-100 or the cholecystokinin analogue ceruletid (10(-8) M).. An isolated external perfusion of a rat pancreas is possible without inducing any increase of parameters of damage such as amylase, lipase or lactate-dehydrogenase in the outflow medium. The perfusion time should be limited to 80 minutes including a 20 minutes equilibration period. A damage of pancreatic parenchyma is indicated by increased levels of pancreatic enzymes in the perfusion medium. Such damage can be induced by various noxious substances like detergents or cerulein, which has a significance in the pathophysiology of experimental acute pancreatitis. A significant increase (p < 0.01) of lactate-dehydrogenase, lipase and amylase was found 10, 20 and 30 minutes after an application of triton x-100. During a perfusion with the cholecystokinin analogue ceruletid (10(-8) M) we found an increase of lipase (p < 0.05) after 30 minutes and an increase of amylase (p < 0.05) after 50 minutes perfusion.. The isolated perfused rat pancreas is a valuable experimental model to investigate the early phase of pathophysiology in acute pancreatitis.

Topics: Acute Disease; Amylases; Analysis of Variance; Animals; Ceruletide; Detergents; Gastrointestinal Agents; In Vitro Techniques; L-Lactate Dehydrogenase; Lipase; Male; Models, Animal; Pancreas; Pancreatitis; Perfusion; Polyethylene Glycols; Rats; Rats, Wistar; Time Factors

The pancreas of 24 male Wistar rats was perfused extracoporally by modified Krebs-Ringer-buffer for 80 minutes (including a 20 minutes equilibration period). To verify any organ damage we measured the activity of pancreatic enzymes like amylase, lipase and lactatdehydrogenase in the portal effluent. Furthermore histological changes were analysed after perfusion. Organ damage was induced by adding cerulein in a physiological dose (10(-10) M, n = 6) and in a supramaximal dose (10(-8) M, n = 6) and by intraductal injection of taurocholate (3.5 %, n = 6).. Already 10 minutes after stimulation with cerulein (10(-8) M) and after intraductal injection of taurocholate increased activities (p < 0.01) of amylase and lipase were measured in the portal effluent compared to the group without any treatment. Lactatdehydrogenase levels did not changed. Apart from marked oedema in both groups considerable zones of necrosis could be noticed especially in the taurocholate group.. Our data suggest that the isolated perfused rat pancreas (IPRP) is a valuable experimental tool to verify pathophysiological changes in the early phase of acute pancreatitis (AP). Various established models of AP such as by cerulein hyperstimulation or intraductal injection of taurocholate, could be applied to the IPRP. We conclude that this method enlarges the spectrum of established experimental models of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cholagogues and Choleretics; Detergents; Disease Models, Animal; Gastrointestinal Agents; In Vitro Techniques; L-Lactate Dehydrogenase; Lipase; Male; Necrosis; Pancreas; Pancreatitis; Perfusion; Rats; Rats, Wistar; Taurocholic Acid; Time Factors

The function of primary sensory neurons is to receive and transmit information from external environment and these neurons are able to release neuromediators from the activated peripheral endings. The aim of this study was to determine the influence of sensory nerves and administration of their mediator--calcitonin gene related peptide (CGRP) on the course of acute pancreatitis (AP). Ablation of sensory nerves was performed by neurotoxic dose of capsaicin (100 mg/kg). Single or repeated episodes of AP were induced by caerulein infusion (10 microg/kg/h for 5 h). Five repeated AP were performed once a week. Capsaicin at the dose which stimulates sensory nerves (0.5 mg/kg/dose) or CGRP (10 microg/kg/dose) was administrated before and during or after single induction of AP, as well as, after each induction of repeated AP. Rats were killed at the time 0, 3 or 9 h after single induction of AP or two weeks after last induction of repeated AP. Ablation of sensory nerves aggravated pancreatic damage in caerulein-induced AP. Treatment with stimulatory doses of capsaicin or CGRP before and during single induction of AP attenuated the pancreatic damage in morphological examination. This effect was also manifested by partial reversion of AP evoked drop in DNA synthesis and pancreatic blood flow (PBF). Administration of CGRP after single AP induction aggravated histologically manifested pancreatic damage. The further decrease in PBF and DNA synthesis was also observed. Animals with five episodes of AP showed almost full pancreatic recovery two weeks after last induction of AP concerning all parameters tested. In stimulatory doses of capsaicin treated rats, we observed the decrease in pancreatic amylase and fecal chymotrypsin activity, as well as, the drop in DNA synthesis. Similar but less pronounced effects were observed after treatment with CGRP. We conclude that effect of sensory nerves and CGRP on AP is two-phase and time dependent. Stimulation of sensory nerves or the administration of CGRP during development of AP exhibits protective effects against pancreatic damage induced by caerulein overstimulation. After induction of AP, persistent activity of sensory nerves and presence of CGRP aggravate pancreatic damage and lead to functional insufficiency typical for chronic pancreatitis.

Topics: Acute Disease; Amylases; Animals; Calcitonin Gene-Related Peptide; Capsaicin; Ceruletide; DNA; Interleukin-1; Male; Neurons, Afferent; Nitric Oxide; Pancreas; Pancreatitis; Rats; Rats, Wistar

To examine the role of p38 during acute experimental cerulein pancreatitis.. Rats were treated with cerulein with or without a specific JNK inhibitor (CEP1347) and/or a specific p38 inhibitor (SB203580) and pancreatic stress kinase activity was determined. Parameters to assess pancreatitis included trypsin, amylase, lipase, pancreatic weight and histology.. JNK inhibition with CEP1347 ameliorated pancreatitis, reducing pancreatic edema. In contrast, p38 inhibition with SB203580 aggravated pancreatitis with higher trypsin levels and, with induction of acinar necrosis not normally found after cerulein hyperstimulation. Simultaneous treatment with both CEP1347 and SB203580 mutually abolished the effects of either compound on cerulein pancreatitis.. Stress kinases modulate pancreatitis differentially. JNK seems to promote pancreatitis development, possibly by supporting inflammatory reactions such as edema formation while its inhibition ameliorates pancreatitis. In contrast, p38 may help reduce organ destruction while inhibition of p38 during induction of cerulein pancreatitis leads to the occurrence of acinar necrosis.

Topics: Acute Disease; Animals; Carbazoles; Ceruletide; Enzyme Inhibitors; Imidazoles; Indoles; Mitogen-Activated Protein Kinases; Models, Animal; Necrosis; p38 Mitogen-Activated Protein Kinases; Pancreatitis; Pyridines; Rats; Trypsin

In this study we aimed to clarify the role of mast cells in the development and progression of inflammation in cerulein-induced acute pancreatitis (AP) in rats. We have also examined the effects of ketotifen; a mast-cell stabilizing agent in the treatment of acute pancreatitis and its relation with nitric oxide (NO) synthesis.. In the first part of the study we planned to examine the effects mast cell stabilization in acute pancreatitis, while the second part was focused on examining the relation between NO synthesis and the potential effects of ketotifen in AP. Wistar albino rats were randomly divided into 6 groups (n: 10). In the first part of the study, AP was induced by four subcutaneous (sc) injections of 20 microg/kg body weight of cerulein at hourly intervals in Groups A and B while Group C was treated with saline as the control group. Group B was pretreated with ketotifen 1 mg/kg (ip). In the second part, the study design was similar except for the inhibition of nitric oxide synthesis by N-nitro L-arginine methyl ester (L-NAME) 30 mg/kg (ip) in Groups D, E and F. Group D was treated with L-NAME and cerulein and Group E was treated with ketotifen, L-NAME and cerulein. Group F was treated with L-NAME and saline as the control group. Serum amylase activity and pancreatic myeloperoxidase activity (MPO) were measured. Pancreatic histology and mast-cell count in pancreatic tissue were evaluated.. Mast cell count was found to be increased in the pancreatic tissue in cerulein-induced AP. (2.93+/-0.26 vs 1.98+/-0.26; p<0.001). Ketotifen treatment significantly reduced cerulein induced edema (1.30+/-0.21 vs 0.70+/-0.15; p<0.001), neutrophil infiltration (1.50+/-0.16 vs 0.60+/-0.16; p<0.001) and attenuated the increase in amylase (4394.0+/-149.5 U/L vs 3350.5+/-216.9 U/L; p<0.05) and MPO activity (1.14+/-0.13 U/gr tissue vs 0.54+/-0.08 U/gr tissue; p<0.001). Mast-cell count in pancreatic tissue was also decreased significantly with ketotifen pretreatment (2.93+/-0.26 vs 1.70+/-0.21; p<0.05). Inhibition of NO synthesis with L-NAME treatment decreased the beneficial effects of ketotifen.. It seems likely that mast cell activity may play an important role in the initiation and progression of acute pancreatitis. Ketotifen treatment may reduce the severity of AP in rats. The protective action of ketotifen in cerulein-induced acute pancreatitis is most probably owing to mast cell stabilization and stimulation of NO synthesis.

Topics: Acute Disease; Animals; Anti-Allergic Agents; Ceruletide; Drug Therapy, Combination; Enzyme Inhibitors; Ketotifen; Male; Mast Cells; NG-Nitroarginine Methyl Ester; Pancreatitis; Rats; Rats, Wistar

We previously reported that serum hepatocyte growth factor (HGF) levels are elevated in patients with acute pancreatitis and that pancreatitis-associated ascitic fluid (PAAF) contains cytotoxic factor(s) inducing apoptosis on Madin-Darby canine kidney (MDCK) cells. In this study, plasma HGF levels and HGF tissue distribution were investigated in rats with experimental acute pancreatitis, and the effects of HGF on the cytotoxic activity and apoptosis-inducing activity of PAAF also were examined. Plasma HGF levels were elevated in rats with two experimental pancreatitis models of different grades of severity. The degree of its elevation was correlated with the severity and the organ dysfunctions. In rats with severe pancreatitis, HGF protein and messenger RNA (mRNA) levels significantly increased in liver, kidney, and lung, which were injured organs. When anti-HGF neutralizing antibody was administered in severe pancreatitis, liver dysfunction worsened, and apoptotic cells increased in kidney. Recombinant HGF inhibited the cytocidal activity of PAAF on MDCK cells in a dose-dependent manner. Moreover, recombinant HGF prevented the apoptotic cell death (DNA fragmentation, nuclear fragmentation, and caspase-3 activation) induced by PAAF. These results suggest that HGF is produced in injured organs and may function as an organotrophic and antiapoptotic factor against the organ injuries in acute pancreatitis.

Topics: Acute Disease; Animals; Apoptosis; Caspase 3; Caspases; Cell Line; Ceruletide; Deoxycholic Acid; DNA Fragmentation; Dogs; Edema; Hepatocyte Growth Factor; Kidney; Liver; Lung; Male; Multiple Organ Failure; Pancreatitis; Pancreatitis, Acute Necrotizing; Rats; Rats, Wistar; Recombinant Fusion Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spleen

Activation of the receptor c-met stimulates motility, mitosis, morphogenesis, processes involved in organ regeneration, or progression of malignancies. In the present study we investigated the expression of c-met protein in the regenerating pancreas and characterized the influence of cytokines on c-met expression.. Acute pancreatitis was induced in rats by cerulein injection. Rat acini and rat and human pancreatic cancer cells were stimulated with interleukin-1alpha (IL-1alpha), IL-6, tumor necrosis factor-alpha (TNF-alpha) or transforming growth factor-beta1 (TGF-beta1). C-met expression was analyzed by means of Western blotting and localization in pancreatic tissue by immunohistochemistry.. C-met protein expression was significantly upregulated in the regenerating pancreas and localized in areas of regenerating tissue. Stimulation with cytokines resulted in a two- to threefold increase of c-met expression in vitro.. Enhanced c-met expression after acute pancreatitis suggests that HGF/met has an important role in pancreatic regeneration, which is probably mediated by cytokines. This regulatory mechanism is also of importance in pancreatic cancer.

Topics: Acute Disease; Animals; Blotting, Western; Cells, Cultured; Ceruletide; Cytokines; Hepatocyte Growth Factor; Humans; Immunohistochemistry; Interleukin-1; Interleukin-6; Male; Pancreas; Pancreatic Neoplasms; Pancreatitis; Proto-Oncogene Proteins c-met; Rats; Rats, Wistar; Regeneration; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

Pancreatic stellate cells may be a major source of extracellular matrix deposition during injury. This study was undertaken to establish whether pancreatic stellate cells are a source of Type I collagen in vivo and whether they continue to be a source of matrix production in the post-injury fibrotic pancreas. To induce pancreatic fibrogenesis, acute pancreatic injury was induced in mice three times weekly with supraphysiologic doses of cerulein. Animals were treated for 6 weeks and allowed to recover for an additional 6 weeks. Stellate cell activation and pancreatic collagen expression were measured by immunohistochemistry, whole tissue RNA analysis, and in situ hybridization. Histology and digital image analysis demonstrated the development of substantial pancreatic fibrosis after 6 weeks of treatment. During recovery, incomplete resolution of the fibrosis was found. Procollagen alpha1(I) mRNA increased more than 15-fold during treatment and continued to be 5-fold elevated during the post-injury phase. In situ hybridization studies demonstrated that collagen gene expression was colocalized to activated pancreatic stellate cells. Collagen expression and fibrosis persisted in focal areas during recovery. These findings show that pancreatic stellate cells are the major source of collagen during repetitive injury in vivo. Additionally, focal areas of sustained pancreatic fibrogenesis persist after cessation of cerulein treatment, and these areas may contribute to sustained total organ collagen expression in the absence of ongoing injury.

Topics: Acute Disease; Animals; Ceruletide; Female; Immunohistochemistry; In Situ Hybridization; Mice; Pancreas; Pancreatitis; Procollagen; RNA, Messenger

Few data are available on the potential role of T lymphocytes in experimental acute pancreatitis. The aim of this study was to characterize their role in the inflammatory cascade of acute pancreatitis.. To type this issue, acute pancreatitis was induced by repeated injections of cerulein in nude mice and in vivo CD4(+) or CD8(+) T cell-depleted mice. The role of T lymphocyte-costimulatory pathways was evaluated using anti-CD40 ligand or anti-B7-1 and -B7-2 monoclonal blocking antibodies. The role of Fas-Fas ligand was explored using Fas ligand-targeted mutant (generalized lymphoproliferative disease) mice. Severity of acute pancreatitis was assessed by serum hydrolase levels and histology. Intrapancreatic interleukin 12, interferon gamma, Fas ligand, and CD40 ligand messenger RNA were detected by reverse-transcription polymerase chain reaction. Intrapancreatic T lymphocytes were identified by immunohistochemistry.. In control mice, T cells, most of them CD4(+) T cells, are present in the pancreas and are recruited during acute pancreatitis. In nude mice, histological lesions and serum hydrolase levels are significantly decreased. T-lymphocyte transfer into nude mice partially restores the severity of acute pancreatitis and intrapancreatic interferon gamma, interleukin 12, and Fas ligand gene transcription. The severity of pancreatitis is also reduced by in vivo CD4(+) (but not CD8(+)) T-cell depletion and in Fas ligand-targeted mutant mice. Blocking CD40-CD40 ligand or B7-CD28 costimulatory pathways has no effect on the severity of pancreatitis.. T lymphocytes, particularly CD4(+) T cells, play a pivotal role in the development of tissue injury during acute experimental pancreatitis in mice.

Topics: Acute Disease; Animals; B7-1 Antigen; CD28 Antigens; CD4-Positive T-Lymphocytes; CD40 Antigens; Ceruletide; Fas Ligand Protein; Female; Ligands; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Pancreas; Pancreatitis; Reference Values

The role of repetitive acute injury in the pathogenesis of chronic pancreatitis remains unknown. To determine if repetitive injury induced by pancreatic hyperstimulation would reproduce the characteristic features of human chronic pancreatitis, acute reversible pancreatic injury was induced in mice by twice weekly cerulein treatment, 50 microg/kg/hr x 6 hr, for 10 weeks. Procollagen alpha1(I) mRNA was markedly increased by week 2. Sirius red staining of interstitial collagen demonstrated progressive accumulation of extracellular matrix surrounding acinar units and in interlobular spaces. Atrophy, transdifferentiation of acinar units to ductlike tubular complexes, and dilatation of intraacinar lumina also developed. Electron microscopy demonstrated the presence of stromal cells in areas of fibrosis with morphologic characteristics of pancreatic stellate cells. These findings demonstrate that, in a murine model, repetitive acute injury to the pancreas by hyperstimulation can reproduce the major morphological characteristics of human chronic pancreatitis.

Topics: Acute Disease; Animals; Atrophy; Ceruletide; Chronic Disease; Extracellular Matrix; Female; Fibrosis; Gene Expression Regulation; Humans; Mice; Microscopy, Electron; Pancreatitis; Procollagen

We investigated whether the timing of administration of contrast medium after onset of acute pancreatitis is critical in determining the magnitude of microcirculatory derangement.. An acute pancreatitis model in male Sprague-Dawley rats (225-275 g) was established by continuous infusion of cerulein (15 mg/kg per hour). The mean arterial pressure was monitored continuously by means of a femoral artery catheter. Diatrizoate (Hypaque-76), a water-soluble contrast medium, was delivered through a femoral vein catheter at doses corresponding to those given to humans, either 1, 2, or 3 hours after pancreatitis induction. In vivo microscopy and laser-Doppler flowmetry were used to investigate microcirculatory derangement. The water contents of the pancreas and lung, the malondialdehyde levels of the pancreas, and the trypsinogen activation peptide levels in the serum were measured at the end of the experiment (8 hours after infusion of cerulein).. Early administration of contrast medium (1 hour after pancreatitis induction) resulted in significantly greater changes in microcirculation and mean arterial pressure than did late administration (2 or 3 hours after pancreatitis induction). Rats given contrast medium 1 hour after induction also had highest pancreas and lung water contents, the highest pancreas malondialdehyde levels, and the highest serum trypsinogen activation peptide levels.. These results show that a water soluble contrast medium that is often used for computed tomographic imaging of the pancreas can adversely affect the pancreatic microcirculatory parameters, such as tissue perfusion and leukocyte sticking, and hemodynamics in a cerulein-induced model of acute pancreatitis. Early administration seems to cause more severe derangement of the pancreatic microcirculation.

Topics: Acute Disease; Animals; Blood Pressure; Ceruletide; Contrast Media; Diatrizoate; Laser-Doppler Flowmetry; Male; Malondialdehyde; Microcirculation; Microscopy; Oligopeptides; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Trypsinogen

Pancreatic oedema occurs early in the development of acute pancreatitis, and the overall extent of fluid loss correlates with disease severity. The tachykinin substance P (SP) is released from sensory nerves, binds to the neurokinin-1 receptor (NK1-R) on endothelial cells and induces plasma extravasation, oedema, and neutrophil infiltration, a process termed neurogenic inflammation. We sought to determine the importance of neurogenic mechanisms in acute pancreatitis. Pancreatic plasma extravasation was measured using the intravascular tracers Evans blue and Monastral blue after administration of specific NK1-R agonists/antagonists in rats and NK1-R(+/+)/(-/-) mice. The effects of NK1-R genetic deletion/antagonism on pancreatic plasma extravasation, amylase, myeloperoxidase (MPO), and histology in cerulein-induced pancreatitis were characterized. In rats, both SP and the NK1-R selective agonist [Sar(9) Met(O(2))(11)]SP stimulated pancreatic plasma extravasation, and this response was blocked by the NK1-R antagonist CP 96,345. Selective agonists of the NK-2 or NK-3 receptors had no effect. In rats, cerulein stimulated pancreatic plasma extravasation and serum amylase. These responses were blocked by the NK1-R antagonist CP 96,345. In wildtype mice, SP induced plasma extravasation while SP had no effect in NK1-R knockout mice. In NK1-R knockout mice, the effects of cerulein on pancreatic plasma extravasation and hyperamylasemia were reduced by 60%, and pancreatic MPO by 75%, as compared to wildtype animals. Neurogenic mechanisms of inflammation are important in the development of inflammatory oedema in acute interstitial pancreatitis.

Topics: Acute Disease; Amylases; Animals; Blood Pressure; Ceruletide; Edema; Gastrointestinal Agents; Inflammation; Male; Mice; Neurokinin-1 Receptor Antagonists; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley; Receptors, Neurokinin-1; Substance P

The connective tissue component hyaluronan is accumulated locally in the damaged tissue during various inflammatory conditions. Owing to the strong water-binding capacity of this glycosaminoglycan, increased tissue content of hyaluronan is paralleled by the development of interstitial edema. The aim with the current experiment was to investigate whether hyaluronan is accumulated in acute pancreatitis and if increased levels of hyaluronan can be correlated to the inflammation of the pancreatic tissue.. Acute pancreatitis was induced in Sprague-Dawley rats by the administration of supramaximal doses of the cholecystokinin analogue caerulein. The animals were followed for 5 hours (n = 4), 24 hours (n = 6), or 48 hours (n = 5), and the pancreata were then investigated for hyaluronan and water content, hyaluronan distribution, general morphology and the presence of CD44-positive cells, macrophages, and T lymphocytes.. Hyaluronan accumulated in the edematous interstitium during acute pancreatitis. Twenty-four hours after the induction of pancreatitis, the hyaluronan content of the pancreata had increased by more than 100%. Simultaneously, CD44-positive cells infiltrated the tissue. However, no correlation between hyaluronan and water was seen at any time point.. This study shows that acute pancreatitis is associated with a strong but transient increase in interstitial hyaluronan and an infiltration of CD44-positive cells located mainly in the same region as the accumulated hyaluronan.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Edema; Hyaluronic Acid; Hyaluronoglucosaminidase; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Water

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Edema; Humans; Hyaluronic Acid; Pancreatitis; Rats

Whether acute pancreatitis induced by cerulein was aggravated in human interleukin 6 (IL-6) transgenic mice and whether a specific anti-IL-6 receptor antibody improved pancreatitis were investigated. To induce acute pancreatitis, cerulein (50 microg/kg, seven injections) with or without 1 mg/kg lipopolysaccharides (LPS) was injected intraperitoneally every hour. In some mice, a monoclonal anti-IL-6 receptor antibody was administered before the first cerulein injection. The animals were killed 1 hour after the last injection. The pancreatic wet weight induced by cerulein alone was significantly higher in IL-6 transgenic mice compared with wild-type mice, but pretreatment with a specific anti-IL-6 receptor antibody did not reduce interstitial edema. When cerulein was administered with LPS, the pancreatic wet weight increased much more than when pancreatitis was induced by cerulein alone in both genotypes, and pretreatment with the anti-IL-6 receptor antibody decreased the pancreatic edema only in human-IL-6 transgenic mice. These results suggest that anticytokine antibodies may be effective in improving acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Antibodies, Monoclonal; Ceruletide; Edema; Humans; Interleukin-6; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Organ Size; Pancreas; Pancreatitis; Receptors, Interleukin-6

Adenosine has been shown to modulate various pathophysiologic conditions through receptor-mediated mechanisms. However, the role of adenosine in the pathogenesis of acute pancreatitis has not been described. We examined the effect of adenosine-receptor stimulation or inhibition on the pathologic changes of the pancreas.. Rats received intraperitoneal injections of selective agonists of A1, A2a, and A3 adenosine receptors: 2-chloro-N(6)-cyclopentyladenosine (CCPA), CGS-21680 (CGS), or 1-deoxy-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-be ta-D-ribofuranuronamide (IB-MECA), respectively. Serum amylase activity and pathologic changes of the pancreas were evaluated. The effects of a specific A1-receptor antagonist (FK-838) on the pathologic findings of cerulein- and taurocholate-induced pancreatitis were also examined.. Administration of a selective A1 agonist induced hyperamylasemia and morphologic changes in the pancreas characterized by interstitial edema and leukocyte infiltration; neither A2a nor A3 agonist produced such changes. Treatment with an A1-receptor antagonist significantly attenuated the outcome induced by A1 agonist stimulation. In addition, the A1-receptor antagonist significantly ameliorated pancreatic edema in both pancreatitis models, although it did not improve the acinar cell damage of the pancreas or the increase of serum amylase.. Activation of the adenosine A1-receptor pathway may have an important role in the pathogenesis of acute pancreatitis.

Topics: Acute Disease; Adenosine; Amylases; Animals; Ceruletide; Edema; Leukocytes; Male; Pancreas; Pancreatic Diseases; Pancreatitis; Phenethylamines; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Pyrazoles; Pyridines; Rats; Rats, Wistar; Receptor, Adenosine A3; Receptors, Purinergic P1; Taurocholic Acid

Autodigestion of the pancreas by its own prematurely activated digestive proteases is thought to be an important event in the onset of acute pancreatitis. The mechanism responsible for the intrapancreatic activation of digestive zymogens is unknown, but a recent hypothesis predicts that a redistribution of lysosomal cathepsin B (CTSB) into a zymogen-containing subcellular compartment triggers this event. To test this hypothesis, we used CTSB-deficient mice in which the ctsb gene had been deleted by targeted disruption. After induction of experimental secretagogue-induced pancreatitis, the trypsin activity in the pancreas of ctsb(-/-) animals was more than 80% lower than in ctsb(+/+) animals. Pancreatic damage as indicated by serum activities of amylase and lipase, or by the extent of acinar tissue necrosis, was 50% lower in ctsb(-/-) animals. These experiments provide the first conclusive evidence to our knowledge that cathepsin B plays a role in intrapancreatic trypsinogen activation and the onset of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Cathepsin B; Ceruletide; Disease Models, Animal; Edema; Enzyme Activation; Gene Deletion; Gene Targeting; Humans; Lipase; Mice; Mice, Knockout; Necrosis; Pancreas; Pancreatitis; Phenotype; Trypsinogen

Stimulation of capsaicin sensitive nerves or administration of calcitonin gene-related peptide (CGRP) before induction of acute pancreatitis (AP) attenuates pancreatic damage, whereas CGRP administration after development of AP aggravates lesion of pancreatic tissue. The aim of this study was to determine the effect of prolonged activity of sensory nerves or CGRP administration on the pancreatic repair after repeated episodes of AP. Five episodes of acute caerulein-induced pancreatitis (10 microg/kg/h for 5 h s.c.) were performed at weekly intervals in rats receiving either vehicle or capsaicin at the sensory nerve stimulatory dose (0.5 mg/kg, 3 times daily), or CGRP (10 microg/kg, 3 times daily). Two weeks after the last induction of AP morphological signs of pancreatic damage, pancreatic blood flow (PBF), serum and pancreatic amylase activity, fecal chymotrypsin activity, pancreatic weight, pancreatic RNA and DNA content, as well as, serum interleukin-1beta (Il-1beta ) were assessed. Pancreata of animals receiving vehicle alone showed almost full recovery within two weeks after last episode of pancreatitis induction. In capsaicin-treated group of rats, we observed the increase in PBF by 44% and in serum Il-1beta concentration by 91%. The pancreatic amylase activity, fecal activity of chymotrypsin, pancreatic nucleic acids content and DNA synthesis were decreased. In rats treated with CGRP the alterations in PBF, serum Il-1beta concentration, as well as, in pancreatic and fecal activity of enzymes were similar to capsaicin treated group but less pronounced. We conclude that prolonged activity of capsaicin-sensitive sensory nerves and the presence of their main mediator-CGRP during pancreatic regeneration after AP leads to pancreatic functional insufficiency typical for chronic pancreatitis.

Topics: Acute Disease; Amylases; Animals; Calcitonin Gene-Related Peptide; Capsaicin; Ceruletide; Chymotrypsin; DNA; Interleukin-1; Male; Neurons, Afferent; Pancreas; Pancreatitis; Rats; Rats, Wistar; RNA, Messenger

Mitogen-activated protein kinase (MAPK) family members such as c-jun N-terminal kinase (JNK) may act as signal transducers early during pancreatitis development and evidence indicates that MAPK phosphatases (MKP) downregulate MAPK. We therefore investigated expression and regulation of pancreatic MKP in vivo. Pancreatic MKP mRNA levels were near or below the detection threshold in unstimulated animals. Cerulein hyperstimulation strongly induced MKP-1, MKP-3, and MKP-5 expression, peaking 30 to 60 min after treatment. Thus, MKP's clearly are early responsive genes during pancreatitis induction. Interestingly, inhibition of MKP-1 expression by Ro-31-8220 maximally induced activation of JNK but not of p38 and ERK in acutely isolated acini. These effects indicate that JNK may indeed be a preferred MKP-1 substrate in vivo.

Topics: Acute Disease; Animals; Cell Cycle Proteins; Ceruletide; Dual Specificity Phosphatase 1; Enzyme Inhibitors; Female; Gene Expression Regulation, Enzymologic; Immediate-Early Proteins; Indoles; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 4; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Pancreatitis; Phosphoprotein Phosphatases; Protein Phosphatase 1; Protein Tyrosine Phosphatases; Rats; Rats, Wistar; RNA, Messenger; Signal Transduction

To find out if two immunomodulatory drugs used in organ transplantation (FK506 (tacrolimus) and OKT3 (Orthoclone) would reduce early inflammatory complications in experimental acute pancreatitis.. Laboratory study.. University hospital, Germany.. 36 Balb/c mice.. Pancreatitis induced by 7 intraperitoneal injections of cerulein 50 microg/kg at hourly intervals followed by FK506 0.32 mg/kg, OKT3 0.6 mg/kg, or 0.9% sodium chloride (controls) (n = 12 in each group). 12 hours after induction of pancreatitis the animals were killed.. Serum amylase activity and interleukin-6 (IL-6) concentrations; histological damage to pancreas and lungs, apoptotic cells in pancreas; and myeloperoxidase activity in lungs.. No animal died during the experiment. At 12h serum amylase activity and IL-6 concentrations were increased in all 3 groups, but highest in the OKT3 group. The pancreatic histological score, apoptosis, and inflammatory infiltration were lower in the two experimental groups than controls, but the degree of vacuolisation of acinar cells was similar. Packed cell volume was higher in the control than the experimental groups, and pulmonary damage and myeloperoxidase activity were less in the experimental groups than the controls.. Single therapeutic doses of FK506 and OKT3 reduced the early severity of pancreatitis, pulmonary damage, and haemoconcentration in mice. Single doses of FK506 or OKT3 may therefore be effective in preventing the early complications of pancreatitis.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Ceruletide; Female; Immunosuppressive Agents; Interleukin-6; Lung; Mice; Mice, Inbred BALB C; Muromonab-CD3; Pancreas; Pancreatitis; Peroxidase; Tacrolimus

Heat shock proteins (HSPs) 70 and 27 are stress-responsive proteins that are important for cell survival after injury; the expression of these HSPs is regulated primarily by the transcription factor heat shock factor-1 (HSF-1). The purpose of this study was to determine the effect of acute pancreatitis on pancreatic HSPs (70, 27, 60, and 90) expression and to assess potential mechanisms for HSP induction using a murine model of cerulein-induced pancreatitis. We found an increase of both HSP70 and HSP27 levels with expression noted throughout the pancreas after induction of pancreatitis. HSP60 and HSP90 levels were constitutively expressed in the pancreas and did not significantly change with acute pancreatitis. HSF-1 DNA binding activity increased in accordance with increased HSP expression. We conclude that acute pancreatitis results in a marked increase in the expression of HSP70 and HSP27. Furthermore, the induction of HSP70 and HSP27 expression was associated with a concomitant increase in HSF-1 binding activity. The increased expression of both HSP70 and HSP27 noted with pancreatic inflammation may confer a protective effect for the remaining acini after acute pancreatitis.

Topics: Acute Disease; Animals; Blotting, Western; Ceruletide; DNA; DNA-Binding Proteins; Electrophoresis; Female; Heat Shock Transcription Factors; Heat-Shock Proteins; HSP70 Heat-Shock Proteins; Mice; Molecular Chaperones; Neoplasm Proteins; Pancreatitis; Transcription Factors

alpha 1-Acid glycoprotein (AAG), a highly negatively charged glycoprotein, well known for its capillary stabilizing effect, was tested in rat models of acute edematous pancreatitis, acute hemorrhagic-necrotizing pancreatitis, and acute respiratory distress syndrome (ARDS). In cerulein-elicited edematous pancreatitis AAG improved histological alterations at 200 mg/kg i.v. and plasma amylase activity at 1800 or 4200 mg/kg i.v. All other parameters (edema, plasma lipase) were not affected in a biologically relevant manner. In glycodeoxycholic acid-induced hemorrhagic-necrotizing pancreatitis AAG was without effect on parameters measured (plasma amylase, plasma lipase activity, histological scores) at 1800 or 4200 mg/kg i.v. At the extremely high dose of 1500 mg/kg i.v. plasma amylase and lipase levels were decreased. In lipopolysaccharide-mediated ARDS, AAG was tested at 50, 200 or 600 mg/kg i.v. AAG, but also the placebo formulation decreased the myeloperoxidase content in the bronchoalveolar lavage fluid. Histological alterations were improved by AAG, however, not by the placebo formulation. Lung water content was not significantly influenced by AAG, whereas Evans blue extravasation was significantly diminished by all three doses of AAG. It is concluded that the edematous pancreatitis is the first in vivo condition with increased extravascular fluid accumulation, in which AAG is not effective. Based on data presented here and literature data, there is evidence for a beneficial effect of AAG in acute lung injury.

Topics: Acute Disease; Animals; Bronchoalveolar Lavage Fluid; Ceruletide; Edema; Glycodeoxycholic Acid; Hemorrhage; Lipopolysaccharides; Lung Diseases; Male; Orosomucoid; Pancreatitis; Rats; Rats, Sprague-Dawley; Respiratory Tract Diseases

The regenerative process after acute inflammation of the pancreas is characterized by cell proliferation as well as synthesis and transient deposition of extracellular matrix. Although the regulation of these processes is still unknown, there is growing evidence that the coordinated activity of various growth factors plays an important role in regeneration. Cerulein-induced pancreatitis in the rat was used to analyze whether growth factors and their receptor concentrations are changed in the acute pancreatitis. Messenger RNA hybridization revealed an individual expression pattern for each analyzed growth factor. The mRNA levels of platelet-derived growth factor B (PDGF B), epidermal growth factor (EGF), and insulin-like growth factor 2 (IGF-2) were not altered, whereas fibroblast growth factor-1 (FGF-1) and 2, IGF-1, transforming growth factor-alpha (TGFalpha), and hepatocyte growth factor (HGF) showed markedly increased concentrations with different expression maxima and duration compared with mRNA levels in healthy pancreata. The FGF-2 and IGF-1 expressions were increased between 1 and 3 days after induction of pancreatitis with maxima at day 2. HGF and FGF-1 mRNAs were upregulated between days 3 and 5. In contrast, TGFalpha exhibited the most prolonged overexpression. In the corresponding receptors, only c-met, the HGF-binding protein, showed higher mRNA and protein levels, whereas the expression of the other receptors did not change. Furthermore, in cultured pancreatic epithelial cells, HGF stimulated the expression of its own receptor in an autocrine manner. These results point out that the highly coordinated process of regeneration after pancreatitis may be influenced by a sequential induction and expression of peptide growth factors and their receptors.

Topics: Acute Disease; Animals; Ceruletide; Fibroblast Growth Factor 1; Fibroblast Growth Factor 2; Gene Expression Regulation; Hepatocyte Growth Factor; Insulin-Like Growth Factor I; Male; Pancreatitis; Rats; Rats, Wistar; Receptors, Growth Factor; RNA, Messenger; Time Factors; Transcription, Genetic; Transforming Growth Factor alpha

This presentation reviews the role of cholecystokinin (CCK) as a contributory factor for the development and progression of acute pancreatitis (AP) and the perspective of CCK receptor antagonists for treatment of AP. High, supraphysiological concentrations of CCK induce AP in various species including man. There is also evidence that physiological increases in plasma CCK deteriorates AP in several animal models. The latter findings support the hypothesis that CCK plays a contributory or permissive role for the development of AP. The majorities of experimental studies show that the prophylactic and therapeutic use of CCK antagonists ameliorates AP. The latter effects were clearly shown for models of biliary AP in which plasma CCK is increased due to a feedback mechanism. However, CCK antagonists also had beneficial effects in models in which plasma CCK is not increased. In animal strains which do not have a CCK-A-receptor due to a genetic abnormality AP induced by a certain noxious factor does not develop to the same severity when compared to animals with a normal CCK-A-receptor. Thus, CCK acts as a permissive or contributory factor for the development and progression of AP. There is also evidence that CCK antagonists have a potential therapeutic benefit. Clinical studies will evaluate their therapeutic potential for patients with AP.

Topics: Acute Disease; Animals; Ceruletide; Cholecystokinin; Devazepide; Disease Models, Animal; Disease Progression; Humans; Indoles; Pancreatitis; Rats; Receptors, Cholecystokinin

Intercellular adhesion molecule 1 (ICAM-1) and neutrophils play important roles in many inflammatory processes, but their importance in both acute pancreatitis and pancreatitis-associated lung injury has not been defined.. To address this issue, mice that do not express ICAM-1 were used and depleted of neutrophils by administration of antineutrophil serum. Pancreatitis was induced by administering either supramaximal doses of the secretagogue cerulein or feeding a choline-deficient, ethionine-supplemented diet. The severity of pancreatitis was evaluated by quantitating serum amylase, pancreatic edema, acinar cell necrosis, and pancreas myeloperoxidase activity (i.e., neutrophil content). Lung injury was evaluated by quantitating lung myeloperoxidase activity and pulmonary microvascular permeability. ICAM-1 was quantitated by enzyme-linked immunosorbent assay and was localized by light-microscopic immunohistochemistry.. It was found that serum, pancreas, and lung ICAM-1 levels increase during pancreatitis. Both pancreatitis and the associated lung injury are blunted, but not completely prevented, in mice deficient in ICAM-1. Neutrophil depletion also reduces the severity of both pancreatitis and lung injury. However, the combination of neutrophil depletion with ICAM-1 deficiency does not reduce the severity of pancreatitis or lung injury to a greater extent than either neutrophil depletion or ICAM-1 deficiency alone. Neither pancreatitis nor pancreatitis-associated lung injury are completely prevented by ICAM-1 deficiency, neutrophil depletion, or combined ICAM-1 deficiency plus neutrophil depletion.. The observations indicate that ICAM-1 plays an important, neutrophil-mediated, proinflammatory role in pancreatitis and pancreatitis-associated lung injury. The studies also indicate that ICAM-1 and neutrophil-independent events also contribute to the evolution of pancreatitis and lung injury in these models.

Topics: Acute Disease; Animals; Capillary Permeability; Ceruletide; Choline Deficiency; Death; Ethionine; Female; Intercellular Adhesion Molecule-1; Lung; Male; Mice; Mice, Knockout; Microcirculation; Neutrophils; Pancreatitis; Peroxidase; Pulmonary Circulation

The therapeutic effects of an intravenously injected carboxamide derivative (IS-741) on lung injury were studied in rats with cerulein-induced pancreatitis complicated by endotoxemia. Pancreatitis was induced by four intramuscular injections of cerulein (50 microg/kg at 1-hr intervals). Pancreatitis rats were injected intraperitoneally with 10 mg/kg of lipopolysaccharide (LPS) 6 hr following the first cerulein injection as a challenge of endotoxemia. Rats were divided into four groups: group I, pancreatitis with LPS; group II, pancreatitis with LPS treated with a continuous intravenous injection of IS-741 at 0.03 mg/kg/hr); group III, pancreatitis with LPS treated with a continuous intravenous injection of IS-741 at 0.3 mg/kg/hr); and group IV, pancreatitis with LPS treated with a continuous intravenous injection of IS-741 at 3 mg/kg/hr). IS-741 was administered 30 min before the endotoxemia challenge. Intense mononuclear cell infiltration and lung hemorrhage occurred in untreated pancreatitis rats with LPS (group I), but hemorrhage was not seen in group IV rats receiving a continuous injection of IS-741 shortly before the induction of endotoxemia. The IS-741-treated rats (groups II, III, and IV) had lower serum concentrations of cytokine-induced neutrophil chemoattractant (CINC), as well as fewer pulmonary infiltrates immunoreactive for CINC or Mac-1 (CD11b/CD18). The number of neutrophils infiltrating the lung in groups II, III, and IV was significantly lower than that of group I. Conversely, CINC production by bronchoalveolar macrophages in vitro were stimulated by LPS but were reduced by the presence of IS-741. The carboxamide derivative IS-741 effectively prevented pancreatitis-associated lung injury following the challenge of endotoxemia.

Topics: Acute Disease; Animals; Bronchoalveolar Lavage Fluid; Cells, Cultured; Ceruletide; Endotoxemia; Enzyme Inhibitors; Hemorrhage; Infusions, Intravenous; Interleukin-16; Lung; Lung Diseases; Macrophages; Male; Neutrophils; Pancreatitis; Phospholipases A; Pyridines; Rats; Rats, Sprague-Dawley

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Enzyme Activation; Oligopeptides; Pancreatitis; Rats; Trypsinogen

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Dizocilpine Maleate; Gastrointestinal Agents; Liver; Male; Microscopy, Electron; Mitochondria; Mitochondrial Swelling; Neuroprotective Agents; Pancreatitis; Rats; Rats, Wistar

The amount of enzymes stored in individual zymogen granules and the glycosylation of their membrane have been analysed in rats with acute pancreatitis induced by caerulein after hydrocortisone treatment. The consequences of prolonging hydrocortisone administration after pancreatitis and the use of the cholecystokinin (CCK) receptor antagonist, L-364,718, have also been evaluated.. Analysis was performed using flow cytometry.. Caerulein-induced pancreatitis in rats previously treated for 7 days with hydrocortisone (10 mg kg-1 per day) revealed alterations in enzyme storage in the pancreas. Significant increases in amylase and trypsinogen contents in zymogen granules were observed, an effect associated with a reduction in L-fucose glycoconjugates. Pancreatitis persists 7 days later if hydrocortisone treatment is prolonged. At this stage, a reduced granule fucosylation was still observed, and a significant decrease in the amount of trypsinogen stored in the granules was found. However, hydrocortisone administration led to an increase in intragranular amylase quantities up to normal values, even when L-364,718 was simultaneously administered, but it reverted to plasma as a consequence of pancreatitis. The amount of N-acetyl D-glucosamine in the zymogen granule membrane was not altered by caerulein acute pancreatitis induced under continuous hydrocortisone treatment, but it was decreased by the administration of L-364,718 over 7 days after pancreatitis induction.. The administration of hydrocortisone after the development of pancreatitis prevented recurrence of the disease. L-364,718 proved to be detrimental, not only failing to reduce the symptoms of pancreatitis but also altering the glycoproteins of zymogen granule membrane.

Topics: Acetylglucosamine; Acute Disease; Amylases; Animals; Ceruletide; Cytoplasmic Granules; Devazepide; Enzyme Precursors; Hematocrit; Hydrocortisone; Intracellular Membranes; Male; Pancreatitis; Rats; Rats, Wistar; Receptors, Cholecystokinin; Trypsinogen

Full recovery is always achieved after caerulein induced pancreatitis. Cyclosporin stimulates transforming growth factor beta (TGF-beta) and may interfere with pancreatic regeneration.. To investigate the effects of cyclosporin after caerulein induced pancreatitis or after caerulein injury.. Protocol A: rats received cyclosporin daily (20 mg/kg) and caerulein pancreatitis was induced on days 2 and 8. Protocol B: six courses of caerulein pancreatitis were induced at weekly intervals. Cyclosporin was administered on induction and the day before. Rats recovered for two weeks before being killed. Control groups received saline, cyclosporin, or caerulein alone.. Protocol A: plasma TGF-beta1 and tissue collagenase rose after pancreatitis but decreased towards baseline values on day 15, matching a low collagen content. Morphology disclosed minimal inflammatory infiltration and some interstitial cells immunoreactive for smooth muscle alpha-actin (SMA). TGF-beta1 increased, and remained high in cyclosporin treated groups (cyclosporin alone and cyclosporin plus caerulein). Rats treated with cyclosporin and caerulein showed severe pancreatic weight reduction, abundant inflammatory infiltrates, increased SMA immunoreactive interstitial cells, high collagen content, and delayed collagenase response. No SMA immunoreactive cells were detected in normal rats. Cyclosporin alone also increased SMA immunoreactive cells, despite the absence of inflammatory infiltration and fairly conserved pancreatic structure. Protocol B: the combined pulse treatment induced appreciable collagen deposition and resulted in a smaller pancreas than controls. Morphological examination showed atrophy, fibrosis, fibroblast proliferation, and mononuclear infiltrates.. Cyclosporin greatly distorts pancreatic repair, transforming caerulein induced pancreatitis into a fibrotic chronic-like disease. The mechanism involves TGF-beta, myofibroblasts, and defective collagenase activation.

Topics: Actins; Acute Disease; Animals; Cell Division; Ceruletide; Collagenases; Cyclosporine; Fibrosis; Gastrointestinal Agents; Hydroxyproline; Immunosuppressive Agents; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Regeneration; Transforming Growth Factor beta

Proteases and protease inhibitors are important in acute pancreatitis (AP), although little is known about the time course in cerulein-induced AP in the rat.. AP was induced by supramaximal stimulation of cerulein, 10 microgram/kg/h, and during 72 h we measured lipase, amylase, albumin, prekallikrein, factor X, alpha(1)-protease inhibitor, alpha(1)-macroglobulin, alpha(2)-antiplasmin, antithrombin III (all in plasma) and macroscopic and histologic variables.. Within 12 h an edematous pancreatitis was evident with peak values of peritoneal exudate, pancreatic wet weight ratio, and plasma amylase and lipase activities. Histologically, edema and vacuolization were prominent already after 3 and 6 h, respectively, while inflammation, necrosis, and total histological score gradually increase to reach peak levels at 48 h. Proenzymes and most plasma protease inhibitors decreased to low levels after 6-12 h followed by a gradual increase. The sequential changes over time indicate that kallikrein - kinin activation, and plasminogen activation are probably early events in cerulein-induced AP in rats. alpha(1)-Macroglobulin and alpha(1)-protease inhibitor gradually decreased during the whole study period, probably being "second line" defense inhibitors. Levels above normal were seen for alpha(2)-antiplasmin and factor X at 48 h, normalizing at 72 h.. These results suggest that protease activation and protease inhibitor consumption occur in cerulein-induced AP in the rat.

Topics: Acute Disease; alpha-2-Antiplasmin; alpha-Macroglobulins; Amylases; Animals; Antithrombin III; Ceruletide; Complement C1 Inactivator Proteins; Endopeptidases; Factor X; Infusion Pumps, Implantable; Lipase; Male; Pancreatitis; Prekallikrein; Protease Inhibitors; Rats; Rats, Wistar; Serum Albumin; Time Factors

This study was done to evaluate the possible preventive effects of reactive oxygen species (ROS) scavenger agent desferrioxamine (DFX) and platelet-activating factor (PAF) antagonist agent ginkgo biloba (GB) in an experimental acute pancreatitis model. Seventy-eight CD-1 mice were divided into six groups consisting of 10-13 mice. Induction of pancreatitis was achieved by cerulein injection in groups 2-5. The first group was control, whereas DFX and GB were used alone or in combinations as preventive agents in groups 3-5. DFX or GB were injected to the mice in groups 6 and 7 to evaluate any toxic effect. The assessment of the pancreatic edema and inflammation, the measurement of the amylase and the pancreatic weight and the measurement of the pancreatic tissue oxidative capacity by chemiluminescence method were the parameters to evaluate pancreatitis. Although the results indicate DFX and GB alone or in combinations have significant preventive roles, this was not a complete prevention.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Deferoxamine; Disease Models, Animal; Edema; Free Radical Scavengers; Ginkgo biloba; Inflammation; Leukocytes; Luminescent Measurements; Male; Mice; Pancreas; Pancreatitis; Phytotherapy; Plants, Medicinal; Platelet Activating Factor

The preventive effect of a novel synthetic serine protease inhibitor, sepimostat mesilate (sepimostat), on acute alcohol pancreatic injury, induced by exocrine hyperstimulation and ethanol administration, was assessed and compared with that of a similar protease inhibitor, camostat mesilate (camostat). Conscious rats were infused with 1 microg mL(-1) h(-1) caerulein intravenously for 6 h and with 0.1 g mL(-1) h(-1) ethanol for 9 h, with the latter infusion beginning 3 h after the start of the caerulein infusion. Sepimostat or camostat was administered orally 1 h before the caerulein infusion. Rats infused with caerulein plus ethanol showed increased plasma amylase and lipase activities, and aggravated pancreatic interstitial oedema when compared with rats given caerulein alone. Sepimostat at 10 and 30 mg kg(-1) prevented the increase in plasma amylase and lipase activities caused by caerulein plus ethanol infusion. Sepimostat at 30 mg kg(-1) suppressed the histological change. Camostat did not show any preventive effects at the equivalent dose. When conscious rats were infused with 1 microg mL(-1) h(-1) caerulein alone intravenously for 6 h, plasma amylase and lipase activities were increased compared with rats given saline. Neither drug prevented the increase in these activities at 30mg kg(-1). Our results suggest that sepimostat has superior preventive effects on alcohol-induced acute pancreatic injury compared with camostat. Sepimostat may thus be a useful drug in the therapy of alcohol-induced pancreatitis.

Topics: Acute Disease; Administration, Oral; Amylases; Animals; Ceruletide; Esters; Ethanol; Gabexate; Guanidines; Imidazoles; Lipase; Male; Pancreas; Pancreatitis; Protease Inhibitors; Rats; Rats, Wistar

The aim of the study was to investigate the impact of L-arginine (nitric oxide donor), L-NNA (NO synthase inhibitor), heparin and procaine on the pancreas' microcirculation, serum interleukin 6 (IL-6) level, and microscopic alterations of the pancreatic gland in acute pancreatitis (AP) in rats. AP was induced by 4 i.p. injections of cerulein (15 micrograms/kg/h). Microcirculatory values of the pancreas were measured by means of laser Doppler flowmetry 5 h after the first cerulein injection. Remarkable morphologic changes in the pancreas, including parenchymal necrosis, an elevation of serum IL-6 activity, and significant drop of pancreatic capillary perfusion was observed in rats with NO synthase inhibition. L-arginine improved the pancreatic microcirculation but worsened the microscopic alterations within the pancreas. Heparin had a beneficial effect on the microcirculatory values, serum IL-6 activity, and morphologic changes. Procaine had no effect on the course of AP. Authors conclude that heparin, improving the pancreatic capillary blood perfusion, may be considered as a promising therapeutic agent in acute pancreatitis.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Heparin; Interleukin-6; Male; Microcirculation; Nitric Oxide; Nitroarginine; Pancreas; Pancreatitis; Procaine; Rats; Rats, Wistar

Nitric oxide (NO) is involved in the control of pancreatic blood flow and secretion, and its role in the maintenance of pancreatic tissue has been suggested. The aim of our study was to evaluate the influence of NO on pancreatic trophic parameters during acute pancreatitis induction and the pancreas recovery process.. Acute pancreatitis was induced in rats by a supramaximal dose of caerulein. During acute pancreatitis induction, rats were treated with L-arginine (a substrate for NO synthesis), glyceryl trinitrate (NTG, NO donor), glycine, N(G)-nitro-L-arginine (L-NNA, NO synthase inhibitor), L-arginine + L-NNA, or saline. Pancreatic weight, total contents of RNA, DNA, protein, amylase and chymotrypsin as well as pancreas histology and the number of proliferating acinar cells were evaluated after pancreatitis induction in all groups and additionally after 7 and 14 days of recovery in untreated acute pancreatitis rats or those injected with L-NNA and/or L-arginine.. Pancreas destruction after acute pancreatitis was evidenced by similar decreases of all parameters in untreated acute pancreatitis rats or those treated with L-NNA or glycine when compared to control healthy animals. The recovery process after acute pancreatitis was not affected by L-NNA injections; however, the increased cell proliferation occurred later than in untreated rats. NTG and especially L-arginine treatment resulted in significant attenuation of pancreas damage (partially prevented by L-NNA treatment) as evidenced by biochemical data with a slight improvement in morphology. Treatment with L-arginine alone or in combination with L-NNA resulted in augmented cell proliferation after acute pancreatitis induction followed by earlier recovery in comparison to untreated acute pancreatitis rats or the L-NNA-injected group.. The present study suggests the involvement of NO in mild acute pancreatitis and in the recovery process after inflammatory damage.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Male; Nitric Oxide; Nitric Oxide Synthase; Nitroglycerin; Pancreas; Pancreatitis; Rats; Rats, Wistar

Pancreatic proteases are secreted in acute pancreatitis, but their contribution to associated lung injury is unclear. Applying models of mild edematous (intravenous caerulein) and severe necrotizing (intraductal glycodeoxycholic acid) pancreatitis in rats, we showed that both trypsinogen and trypsin concentrations in peripheral blood, as well as lung injury, correlate with the severity of the disease. To isolate the potential contribution of proteases to lung injury, trypsin or trypsinogen was injected into healthy rats or trypsinogen secreted in caerulein pancreatitis was activated by intravenous enterokinase. Pulmonary injury induced by protease infusions was dose dependent and was ameliorated by neutrophil depletion. Trypsinogen activation worsened lung injury in mild pancreatitis. In vitro incubation of leukocytes with trypsinogen showed that stimulated leukocytes can convert trypsinogen to trypsin. In conclusion, this study demonstrates that the occurrence and severity of pancreatitis-associated lung injury (PALI) corresponds to the levels of circulating trypsinogen and its activation to trypsin. Neutrophils are involved in both protease activation and development of pulmonary injury.

Topics: Acute Disease; Animals; Carcinogens; Ceruletide; Detergents; Endopeptidases; Enteropeptidase; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Glycodeoxycholic Acid; Leukocytes; Lung; Lung Diseases; Male; Oligopeptides; Pancreas; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley; Tetradecanoylphorbol Acetate; Trypsin; Trypsinogen

Little is known about the changes in pancreatic enzyme storage in acute pancreatitis. We have performed flow cytometric studies of zymogen granules from rats with acute pancreatitis induced by hyperstimulation with caerulein. A comparison was made with rats treated with hydrocortisone (10 mg/kg/day) over 7 days before inducing pancreatitis in order to find out whether the amount of enzymes stored in the pancreas plays a key role in the development of pancreatitis. The potentially therapeutic effect of L-364,718 (0.1 mg/kg/day, for 7 days), a CCK receptor antagonist, was assayed in the rats with caerulein-induced pancreatitis which had previously received the hydrocortisone treatment. A significant increase in the intragranular enzyme content was observed 5 h after hyperstimulation with caerulein. The highest values were reached in the rats previously treated with hydrocortisone. The greatest pancreatic enzyme load was parallel to the highest values in plasma amylase, edema and haematocrit observed. Acute pancreatitis was reversed seven days later. At this stage smaller granules appeared in the pancreas whose enzyme content was similar to that of controls when no treatment was applied after pancreatitis. In contrast, L-364,718 administration prevented the favourable evolution of pancreatitis since the antagonism exerted on CCK receptors induced a blockade of secretion of the large amounts of enzymes stored in the pancreas. Moreover, the enzyme content in zymogen granules was below normal values since the stimulatory CCK action on enzyme synthesis can be inhibited by L-364,718. Our results suggest that the efficiency of CCK antagonists, as potential therapy, would also depend on the load of enzymes in the pancreas when acute pancreatitis is produced.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cytoplasmic Granules; Devazepide; Enzyme Precursors; Hormone Antagonists; Hydrocortisone; Male; Pancreatitis; Rats; Rats, Wistar; Receptor, Cholecystokinin A; Receptors, Cholecystokinin; Trypsinogen

Since oxygen free radicals and lipid peroxidation have been implicated in the pathogenesis of an early stage of acute pancreatitis, we examined whether melatonin, a recently discovered free-radical scavenger, could attenuate pancreatic injury in Sprague-Dawley rats with cerulein-induced pancreatitis. Acute pancreatitis was induced by four intraperitoneal injections of cerulein (50 microg/kg body wt) given at 1-hr intervals. Thirty minutes after the last cerulein injection, the rats were killed and the degree of pancreatic edema, the level of lipid peroxidation in the pancreas, and serum amylase activity were increased significantly. Pretreatment with melatonin (10 or 50 mg/kg body wt) 30 min before each cerulein injection resulted in a significant reduction in pancreatic edema and the levels of lipid peroxidation. Serum amylase activity, however, was not significantly influenced by either dose of melatonin. Moreover, we found that cerulein administration was associated with stomach edema as well as high levels of lipid peroxidation in the stomach and small intestine, which were also reduced by melatonin. Melatonin's protective effects in cerulein-treated rats presumably relate to its radical scavenging ability and to other antioxidative processes induced by melatonin.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Edema; Free Radical Scavengers; Lipid Peroxidation; Male; Malondialdehyde; Melatonin; Pancreatitis; Rats; Rats, Sprague-Dawley

Analogues of the previously reported potent and highly selective CCK(1) receptor antagonist (4aS, 5R)-2-benzyl-5-(N-Boc-tryptophyl)amino-1,3-dioxoperhydropyrido-[1, 2-c]pyrimidine (2a) were prepared to explore the structural requirements at the Boc-tryptophan domain for CCK(1) receptor affinity. Structural modifications of 2a involved the Trp side chain, its conformational freedom, the Boc group, and the carboxamide bond. Results of the CCK binding and in vitro functional activity evaluation showed three highly strict structural requirements: the type and orientation of the Trp side chain, the H-bonding acceptor carbonyl group of the carboxamide bond, and the presence of the Trp amino protection Boc. Replacement of this acid-labile group with 3, 3-dimethylbutyryl or tert-butylaminocarbonyl conferred acid stability to analogues 14a and 15a, which retained a high potency and selectivity in binding to CCK(1) receptors, as well as an in vivo antagonist activity against the acute pancreatitis induced by caerulein in rats. Oral administration of compounds 14a and 15a also produced a lasting antagonism to the hypomotility induced by CCK-8 in mice, suggesting a good bioavailability and metabolic stability.

Topics: Acute Disease; Administration, Oral; Animals; Cerebral Cortex; Ceruletide; Hyperkinesis; In Vitro Techniques; Injections, Intraperitoneal; Male; Mice; Pancreas; Pancreatitis; Pyridines; Pyrimidinones; Rats; Rats, Wistar; Receptors, Cholecystokinin; Sincalide; Structure-Activity Relationship; Tryptophan

The interrelationship between acinar cell apoptosis and tubular complex formation was examined in caerulein-induced pancreatitis using histology, immunohistochemistry, electron microscopy and DNA gel electrophoresis. Rats were given 8 hourly subcutaneous injections of caerulein, 24 micrograms/kg, for up to 2 days. Morphologically and biochemically typical apoptosis affected 4.6 and 8.9% of acinar cells at 1 and 2 days, respectively, resulting in removal of most acinar cells by 2 days. Consequently, pancreatic ducts, the lining cells expressing bcl-2 and therefore resistant to apoptosis, became much more closely approximated to form the basis of tubular complexes; small numbers of immunohistochemically discrete acinar cells in their lining were either pre-apoptotic resistant to it or newly formed. Proliferation of duct-like lining cells was associated with apoptosis, an increase in islet cells and acinar cell regeneration. There was evidence of duct to acinar cell differentiation but the main increase in acinar cell numbers appeared to derive from proliferation of newly formed acinar cells.

Topics: Acute Disease; Animals; Apoptosis; Cell Division; Ceruletide; Electrophoresis, Agar Gel; Immunoenzyme Techniques; Male; Microscopy, Electron; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

We investigated how heat shock protein 27 (HSP27) and its phosphorylation are involved in the action of cholecystokinin (CCK) on the actin cytoskeleton by genetic manipulation of Chinese hamster ovary (CHO) cells stably transfected with the CCK-A receptor. In these cells, as in rat acini, CCK activated p38 mitogen-activated protein (MAP) kinase and increased the phosphorylation of HSP27. This effect could be blocked with the p38 MAP kinase inhibitor SB-203580. Examination by confocal microscopy of cells stained with rhodamine phalloidin showed that CCK dose-dependently induced changes of the actin cytoskeleton, including cell shape changes, which were coincident with actin cytoskeleton fragmentation and formation of actin filament patches in the cells. To further evaluate the role of HSP27, CHO-CCK-A cells were transfected with expression vectors for either wild-type (wt) or mutant (3A, 3G, and 3D) human HSP27. Overexpression of wt-HSP27 and 3D-HSP27 inhibited the effects on the actin cytoskeleton seen after high-dose CCK stimulation. In contrast, overexpression of nonphosphorylatable mutants, 3A- and 3G-HSP27, or inhibition of phosphorylation of HSP27 by preincubation of wt-HSP27 transfected cells with SB-203580 did not protect the actin cytoskeleton. These results suggest that phosphorylation of HSP27 is required to stabilize the actin cytoskeleton and to protect the cells from the effects of high concentrations of CCK.

Topics: Actin Cytoskeleton; Actins; Acute Disease; Animals; Cell Fractionation; Ceruletide; CHO Cells; Cricetinae; Culture Media, Serum-Free; Detergents; Enzyme Activation; Enzyme Inhibitors; Fluorescent Antibody Technique; Gene Expression; Heat-Shock Proteins; Humans; Imidazoles; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; Mutagenesis; Octoxynol; p38 Mitogen-Activated Protein Kinases; Pancreatitis; Phosphorylation; Pyridines; Rats; Receptor, Cholecystokinin A; Receptors, Cholecystokinin; Sincalide; Transfection

Nitric oxide (NO) as a unique biological mediator that has been implicated in many physiological and pathophysiological processes may have a significant influence on the course of acute pancreatitis and the recovery process. The aim of the study was to evaluate the effect of a NO synthase inhibitor or a substrate for NO endogenous production on the ultrastructural features of the acinar cells in the course of caerulein-induced acute pancreatitis. Acute pancreatitis was induced in the rats by a supramaximal dose of caerulein. During acute pancreatitis induction, the rats were treated with L-arginine (the substrate for NO synthesis), NG-nitro-L-arginine (L-NNA, NO synthase inhibitor), L-arginine + L-NNA or saline. Light and electron microscopy examinations were performed in all groups after pancreatitis induction and additionally after 7 and 14 days of recovery. The study demonstrated that the NO synthase inhibitor given during pancreatitis induction in rats enhances the damage to the acinar cells, detected ultrastructurally, and increases the cellular inflammatory infiltration. In the later period, the considerable damage to the mitochondria and the changes in secretory compartment were observed, including dilated cisternae of Golgi apparatus, focal degranulation of rough endoplasmic reticulum, and reduced number of zymogen granules and condensing vacuoles. L-arginine reversed to some extent the deleterious effect of L-NNA, although when administered alone it had no apparent effect on the ultrastructure of pancreatic acinar cells compared with untreated animals. The obtained results indicate that the NO synthase inhibitor enhances the ultrastructural degenerative alterations in the pancreatic acinar cells in the course of caerulein-induced acute pancreatitis and confirm the protective role of endogenous nitric oxide in this disease.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Enzyme Inhibitors; Male; Microscopy, Electron; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pancreas; Pancreatitis; Rats; Rats, Wistar

This study was designed to evaluate the protective effect of thromboxane A2 (TXA2) receptor antagonist, seratrodast, against pancreatic injuries during acute pancreatitis. Acute pancreatitis was induced in rats by intravenous infusion of a supramaximal dose of caerulein (5 microg/kg x h for 4 h). In this model, marked hyperamylasemia, a significant increase in pancreatic water content, and a significant increase in pancreatic micro-vascular leakage of Evans blue dye were observed. Pancreatic subcellular redistribution of a lysosomal enzyme, cathepsin B from the lysosomal fraction to the zymogen fraction as well as a significant increase in pancreatic trypsin content were also observed. Pretreatment with seratrodast at a dose of 2 mg/kg (twice, 8 and 4 h before caerulein infusion) significantly inhibited these pancreatic injuries including hyperamylasemia, increased pancreatic microvascular leakage, redistribution of cathepsin B and increased pancreatic trypsin content. These results suggest that TXA2 may be involved in the pathogenesis of acute pancreatitis in the early stage of the disease and that TXA2 receptor antagonist might be of therapeutic value for treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Body Water; Capillary Permeability; Cathepsin B; Ceruletide; Male; Microcirculation; Pancreas; Pancreatitis; Rats; Rats, Wistar; Receptors, Thromboxane; Subcellular Fractions; Tissue Distribution; Trypsin

We have reported previously that cerulein-induced edematous pancreatitis would transform into hemorrhagic pancreatitis by administration of endothelin-1 in rats. In the present study, we tried to protect rat model from developing into hemorrhagic pancreatitis with BQ123 (an ETA receptor antagonist).. The rat model was made by 5-hour restraint water-immersion stress and two intraperitoneal injections of cerulein (40 micrograms/kg) at hourly interval. BQ123 (3 or 6 mg/kg) was administered intravenously 30 minutes before and 2 hours after the first cerulein injection.. Acute hemorrhagic pancreatitis was induced in all rats treated with cerulin + stress. The score for pancreatic hemorrhage was 2.4 +/- 0.2 in this group. In the rats pretreated with BQ123, the score was reduced to 1.0 +/- 0.0, pancreas wet weight and serum amylase activity were significantly reduced, and histologic alterations in the pancreas lightened, also the local pancreatic blood flow improved without affecting the systemic blood pressure.. These results suggest that endothelin-1 should play a role in aggravating the development of acute hemorrhagic pancreatitis, through its action on the pancreatic microcirculation.

Topics: Acute Disease; Animals; Ceruletide; Endothelin Receptor Antagonists; Endothelin-1; Male; Pancreatitis; Peptides, Cyclic; Rats; Rats, Sprague-Dawley

To study the feature of pancreatic microcirculatory impairment in caerulein-induced acute pancreatitis.. The AP model of subcutaneous injection of caerulein was studied by intravital fluorescence microscopy of erythrocytes labeled by FITC (FITC-RBC) and by scanning electron microscopy of vascular corrosion casts and light microscopy of Chinese ink-injected/cleared tissues.. Animals treated with caerulein showed hyper-amylases. Contractions of intralobular arteriolar sphincter, presence of vacuoles in all the layers of sphincter, gross irregularity in capillary network of acini were observed in caerulein-induced groups. The decrease of pancreatic capillary blood flow (P < 0.01), reduction of functional capillary density, and irregular intermittent perfusion (P < 0.05) were found.. Impairment and contraction of pancreatic intralobular arteriolar sphincter are the initial microcirculatory lesions in the early phase of acute pancreatitis induced by Caerulein, and play a key role in the pancreatic ischemia and pancreatic microvascular failure in acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Injections, Subcutaneous; Male; Microcirculation; Pancreas; Pancreatitis; Rats; Rats, Wistar

We tested the hypothesis that membrane wounding of acinar cells is one of the earliest changes during the induction of acute pancreatitis. Wounding of cell membranes was detected by the penetration of the animals own albumin into cells. The pancreatitis was induced by the intraperitoneal injection of supramaximal doses of caerulein. The controls received saline. Fifteen to 180 min. after the injection the animals were perfused with buffer followed by fixative. Frozen sections of pancreas were processed identically for immunocytological localization of albumin. The intensity of staining was quantified by image analysis. Animals receiving caerulein consistently display significantly greater (p < 0.001) anti-albumin immunostaining in the cytoplasm of acinar cells than controls. The penetration of albumin into acinar cells indicates that wounding of their plasma membrane occurs during the onset of acute pancreatitis. Wounding of membranes may allow the exit of molecules such as enzymes from the acinar cells during this period.

Topics: Acute Disease; Animals; Cell Membrane; Cell Membrane Permeability; Ceruletide; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Serum Albumin

We developed a new method to quantitate leukocyte accumulation in tissues and used it to examine the time course and severity of acute experimental pancreatitis.. Leukocyte activation and infiltration are believed to be critical steps in the progression from mild to severe pancreatitis and responsible for many of its systemic complications.. Pancreatitis of graded severity was induced in Sprague-Dawley rats with a combination of caerulein and controlled intraductal infusion. Technetium-99m (99mTc)-labeled leukocytes were quantified in pancreas, lung, liver, spleen, and kidney and compared with myeloperoxidase activity. The severity of pancreatitis was ascertained by wet/dry weight ratio, plasma amylase, and trypsinogen activation peptide in the pancreas. The time course of leukocyte accumulation was determined over 24 hours.. Pancreatic leukocyte infiltration correlated well with tissue myeloperoxidase concentrations. In mild pancreatitis, leukocytes accumulated only in the pancreas. Moderate and severe pancreatitis were characterized by much greater leukocyte infiltration in the pancreas than in mild disease (p < 0.01), and increased 99mTc radioactivity was detectable in the lung as early as 3 hours. 99mTc radioactivity correlated directly with the three levels of pancreatitis.. Mild pancreatitis is characterized by low-level leukocyte activation and accumulation in the pancreas without recruitment of other organs; marked leukocyte accumulation was found in the pancreas and in the lung in more severe grades of pancreatitis. These findings provide a basis for the pathophysiologic production of cytokines and oxygen free radicals, which potentiate organ injury in severe pancreatitis. This study validates a new tool to study local and systemic effects of leukocytes in pancreatitis as well as new therapeutic hypotheses.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Disease Progression; Gastrointestinal Agents; Leukocytes; Male; Pancreatitis; Radiopharmaceuticals; Random Allocation; Rats; Rats, Sprague-Dawley; Reproducibility of Results; Severity of Illness Index; Technetium Tc 99m Exametazime

We recently showed that activation of the hypothalamus-pituitary-adrenal axis may mitigate the progress of acute pancreatitis. To clarify the mechanism, the role of endogenous glucocorticoids in pancreatic acinar cell death was examined.. The occurrence of apoptosis was studied in adrenalectomized or sham-operated rats with or without cerulein-induced pancreatitis. The effects of RU38486, a glucocorticoid-receptor antagonist, on the survival of cultured acinar cells (AR42J) were also examined.. Adrenalectomy caused increases in terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end labeling (TUNEL) of acinar nuclei depending on the time after adrenalectomy but not of other cell types in the pancreas and in other digestive organs. Electron microscopy showed the characteristic features of apoptosis in the TUNEL-labeled acinar cells. In cerulein pancreatitis of adrenalectomized rats, the TUNEL-labeled acinar nuclei increased remarkably depending on the time after cerulein infusion. Replacement of glucocorticoids blocked the occurrence of apoptosis in these experiments. RU38486 induced dose dependently the apoptosis of AR42J cells.. These results provide evidence that endogenous glucocorticoids are an important factor for acinar cell survival. Endogenous glucocorticoids may protect acinar cells by decreasing their sensitivity to the induction of cell death during acute pancreatitis.

Topics: Acute Disease; Adrenalectomy; Animals; Apoptosis; Cells, Cultured; Ceruletide; Dexamethasone; DNA Fragmentation; Gastrointestinal Agents; Glucocorticoids; Hormone Antagonists; Male; Microscopy, Electron; Mifepristone; Pancreatitis; Rats; Rats, Wistar; Receptors, Glucocorticoid

The effect of B2 receptor bradykinin antagonist icatibant on postcapillary leukostasis, microcirculatory stasis, and tissue necrosis was studied in acute pancreatitis. In rats, pancreatitis was induced by intraductal injection of sodium taurocholate (ST), intravenous caerulein and intraductal infusion of glucodeoxycholic acid (GDOC), or intravenous caerulein infusion alone. Intravital pancreatic microcirculation was observed. Icatibant or vehicle was given 30 min before induction of pancreatitis. In ST pancreatitis, the number of perfused capillaries increased in icatibant-pretreated rats (77% vs. 0% for controls, P < 0.001). Capillary flow was preserved in icatibant-treated rats; total stasis was observed in controls. Mean venular leukocyte adherence decreased in icatibant-treated rats (26% vs. 74% for controls, P < 0.001), and median histopathologic score was reduced (icatibant vs. controls, 5.0 vs. 12 points, respectively; P < 0.01). Kinase II inhibitor captopril or exogenous bradykinin in addition to an otherwise effective dosage of icatibant resulted in microcirculatory stasis, extensive venular leukocyte adherence, and severe histological damage. With a 100 times greater icatibant dosage, this adverse effect was compensated. The beneficial effects of icatibant were also observed in intermediate pancreatitis (caerulein + GDOC). In ST and intermediate pancreatitis, icatibant preserved microcirculation, reduced venular leukocyte adherence, and prevented pancreatic tissue damage. B2 receptor bradykinin-mediated postcapillary leukostasis plays an important role in the pathogenesis of severe forms of acute pancreatitis.

Topics: Acute Disease; Animals; Arterioles; Bradykinin; Bradykinin Receptor Antagonists; Capillaries; Cell Adhesion; Ceruletide; Edema; Female; Hemorrhage; Leukocytes; Microcirculation; Pancreas; Pancreatitis; Rats; Rats, Inbred Lew; Receptor, Bradykinin B2; Regional Blood Flow; Taurocholic Acid; Time Factors; Vasoconstriction; Venules

To investigate the debated role of intracellular trypsinogen activation and its relation to lysosomal enzyme redistribution in the pathogenesis of acute pancreatitis, rats were infused with the cholecystokinin analog caerulein at 5 micrograms.kg-1.h-1 for intervals up to 3 h, and the changes were contrasted with those in animals receiving saline or 0.25 microgram.kg-1.h-1 caerulein. Saline or 0.25 microgram.kg-1.h-1 caerulein did not induce significant changes. In contrast, 5 micrograms.kg-1.h-1 caerulein caused significant hyperamylasemia and pancreatic edema within 30 min. Pancreatic content of trypsinogen activation peptide (TAP) increased continuously (significant within 15 min). TAP generation was predominantly located in the zymogen fraction during the first hour but expanded to other intracellular compartments thereafter. Cathepsin B activity in the zymogen compartment increased continuously throughout the experiments and correlated significantly with TAP generation in the same compartment. Total trypsinogen content increased to 143% with marked interstitial trypsinogen accumulation after 3 h. Supramaximal caerulein stimulation causes trypsinogen activation by 15 min that originates in the zymogen compartment and is associated with increasing cathepsin B activity in this subcellular compartment. However, a much larger pool of trypsinogen survives and accumulates in the extracellular space and may become critical in the evolution of necrotizing pancreatitis.

Topics: Acute Disease; Amylases; Animals; Cathepsin B; Cell Fractionation; Ceruletide; Edema; Enzyme Activation; Kinetics; Male; Oligopeptides; Pancreatitis; Rats; Rats, Sprague-Dawley; Subcellular Fractions; Trypsinogen

We examined the effects of treatment with cholecystokinin (CCK) octapeptide (CCK-8) and the CCK receptor antagonist loxiglumide on the recovery of exocrine pancreas in post-acute pancreatitic rats. Acute pancreatitis was induced in rats by intravenous infusion of 20 microg/kg/h cerulein for 4 h. At 24 h after the start of cerulein infusion, rats were divided into nine treatment groups: oral administration of saline (control), or oral administration of 10 or 50 mg/kg body weight loxiglumide twice daily for the first 3 days, followed by saline administration (Loxi-1 and Loxi-2), 10 or 50 mg/kg body weight loxiglumide twice daily for 6 days (Loxi-3 and Loxi-4), oral administration of saline or 10 or 50 mg/kg body weight loxiglumide twice daily for the first 3 days, followed by subcutaneous injection of 2.5 microg/kg body weight CCK-8 twice daily for the next 3 days (CCK-1, CCK-2, and CCK-3), and subcutaneous injection of 2.5 microg/kg body weight CCK-8 twice daily for 6 days (CCK-4). Pancreatic wet weight and biochemical changes were evaluated on day 8 at 12 h after the last treatment. Treatment with loxiglumide (Loxi-3 and Loxi-4) or CCK-8 for 6 days (CCK-4) or with a high dose of loxiglumide for the first 3 days (Loxi-2) significantly suppressed the recovery of pancreatic weight and DNA content compared to saline treatment or to the untreated normal control rats. However, when loxiglumide treatment was followed by 3 days of CCK-8 injections (CCK-2 and CCK-3), pancreatic protein and DNA content recovered to levels comparable to or above the control levels. The most remarkable increase in enzyme content was obtained in postpancreatitic rats treated with high-dose loxiglumide for the first 3 days, followed by CCK-8 injection (CCK-3). On the other hand, 6 days of CCK-8 treatment (CCK-4) had no significant influences on pancreatic enzyme contents. These results suggest that the most favorable strategy for the treatment of acute pancreatitis is to give high-dose loxiglumide during the early stage for only a short period, followed by CCK-8 administration.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cholecystokinin; DNA; Hormone Antagonists; Lipase; Male; Organ Size; Pancreas; Pancreatitis; Proglumide; Rats; Rats, Wistar; Receptors, Cholecystokinin; Regeneration; Sincalide; Trypsin

Subcellular organelles and membranes were the structures most protected by allopurinol, indirectly demonstrating their role of main and early target of oxygen-derived free radicals in the pathogenesis of acute pancreatitis.. The present work evaluates the ultrastructural changes during cerulein-induced acute pancreatitis in rat, with and without treatment with allopurinol.. Supramaximal doses of cerulein, injected intraperitoneally (50 microg/kg) twice, at 1-h interval, caused severe subcellular alterations, including zymogen distribution, pathological vacuoles, and damage to organelles and membranes. Cotreatment (40 mg/kg ip twice with 1-h interval; n = 10 rats) and, most of all, pretreatment (40 mg/kg ip allopurinol, 1 h; 20 mg/kg ip allopurinol + 50 microg/kg ip cerulein, 30 min; 40 mng/kg ip allopurinol, 30 min; 50 microg/kg ip cerulein; n = 10 rats) with allopurinol showed significant morphological improvement.

Topics: Acute Disease; Allopurinol; Animals; Ceruletide; Free Radicals; Male; Microscopy, Electron; Pancreas; Pancreatitis; Rats; Rats, Wistar

Loxiglumide ((+/-)-4-(3,4-dichlorobenzamido)-N-(3-methoxypropyl)-N- pentylglutaramic acid, CAS 107097-80-3, CR 1505) is a cholecystokinin-A (CCK-A) receptor antagonist. In this report, the effects of loxiglumide and gabexate mesilate were studied on three experimental acute pancreatitis models induced by caerulein, sodium taurocholate + caerulein and closed duodenal loop. The intravenous injection of loxiglumide at 3 and 10 mg/kg (6 times at hourly intervals) significantly inhibited an increase in serum amylase activity induced by the intraperitoneal injection of caerulein (50 micrograms/kg i.p., 6 times at hourly intervals) in mice. But gabexate mesilate at 10, 30 and 60 mg/kg did not. The intravenous infusion of loxiglumide at 18 and 60 mg/kg/h showed a life prolonging effect in the lethal necrotizing pancreatitis, induced by the subcutaneous injection of caerulein (50 micrograms/kg s.c., 4 times at 2 h intervals) after the injection of sodium taurocholate (10%, 0.1 ml/body) into the common bile duct, cumulative survival rates being 86 and 90%, respectively. Gabexate mesilate at 180 mg/kg/h showed the prolonging effect (cumulative survival rates 75%). The intravenous injection of loxiglumide at 6, 18 and 60 mg/kg/h significantly inhibited an increase in total ascitic lipase activity, and plasma amylase and lipase activity of rats with closed duodenal loop. These results suggest that CCK plays an important role in the progression of acute pancreatitis, and that loxiglumide may have a therapeutic potential for pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Duodenum; Female; Gabexate; Hormone Antagonists; Injections, Intravenous; Male; Mice; Mice, Inbred ICR; Organ Size; Pancreas; Pancreatitis; Proglumide; Rats; Rats, Sprague-Dawley; Receptors, Cholecystokinin; Serine Proteinase Inhibitors

Substantial quantities of interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) are produced within the pancreatic parenchyma during acute pancreatitis. Recent evidence suggests that IL-1 beta and TNF-alpha propagate acute pancreatitis and intensify the resulting pancreatic acinar cell death. This study examines the direct effect of IL-1 beta and TNF-alpha on pancreatic acinar cells. Human pancreata (n = 6), harvested during organ procurement, were perfused ex vivo through the splenic artery using a sterile, oxygenated colloid solution. Each pancreas was perfused with either recombinant human IL-1 beta or TNF-alpha for 2 h and subsequently with the cholecystokinin analogue caerulein (positive control). Venous effluent was collected continuously and amylase and lipase were determined at 15-min intervals. Pancreatic histology was graded at baseline and following cytokine and caerulein perfusion. To examine the long-term effects of these cytokines on acinar cell viability, additional in vitro studies utilized the AR42J acinar cell line which was exposed to either IL-1 beta or TNF-alpha with survival determined daily by MTT assay. Perfusion of the human pancreas with either IL-1 beta or TNF-alpha did not alter amylase, lipase, or histology. Caerulein did induce pancreatitis as measured by increased amylase, lipase, and pancreatic histology. Survival of pancreatic acinar cells decreased when they were incubated with TNF-alpha but not IL-1 beta. Although present in large amounts within the pancreas during acute pancreatitis, IL-1 beta and TNF-alpha have no direct effect on acinar cell viability or exocrine function acutely nor do they induce pancreatitis. When present for more than 24 h, however, TNF-alpha but not IL-1 beta has a dramatic effect on acinar cell survival.

Topics: Acute Disease; Amylases; Cell Survival; Ceruletide; Humans; In Vitro Techniques; Interleukin-1; Lipase; Pancreas; Pancreatitis; Perfusion; Tumor Necrosis Factor-alpha

Substance P, acting via the neurokinin 1 receptor (NK1R), plays an important role in mediating a variety of inflammatory processes. However, its role in acute pancreatitis has not been previously described. We have found that, in normal mice, substance P levels in the pancreas and pancreatic acinar cell expression of NK1R are both increased during secretagogue-induced experimental pancreatitis. To evaluate the role of substance P, pancreatitis was induced in mice that genetically lack NK1R by administration of 12 hourly injections of a supramaximally stimulating dose of the secretagogue caerulein. During pancreatitis, the magnitude of hyperamylasemia, hyperlipasemia, neutrophil sequestration in the pancreas, and pancreatic acinar cell necrosis were significantly reduced in NK1R-/- mice when compared with wild-type NK1R+/+ animals. Similarly, pancreatitis-associated lung injury, as characterized by intrapulmonary sequestration of neutrophils and increased pulmonary microvascular permeability, was reduced in NK1R-/- animals. These effects of NK1R deletion indicate that substance P, acting via NK1R, plays an important proinflammatory role in regulating the severity of acute pancreatitis and pancreatitis-associated lung injury.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Gene Deletion; Lung; Mice; Mice, Knockout; Pancreas; Pancreatitis; Receptors, Neurokinin-1; Substance P

Pancreatitis complicated with infection often results in the development of multiple organ failure. We investigated the role of altered intracellular calcium as a priming signal for cytokine-induced neutrophil chemoattractant expression in this process. Agents modulating cytosolic Ca2+ were utilized to study the in vivo and in vitro cytokine-induced neutrophil chemoattractant expression for macrophages in rats with cerulein-induced pancreatitis after intraperitoneal administration of lipopolysaccharide as a septic challenge. Pretreatment with the calcium channel blocker verapamil significantly reduced serum cytokine-induced neutrophil chemoattractant concentrations in rats with cerulein-induced pancreatitis after septic challenge. Lipopolysaccharide-stimulated in vitro cytokine-induced neutrophil chemoattractant (CINC) production by peritoneal macrophages was significantly enhanced by pretreatment with thapsigargin (an inhibitor of the endoplasmic reticulum-resident Ca2+-ATPase), but not by A23187 (a calcium-specific ionophore, extracellular Ca2+ influx). Pretreatment with U73122 (a phospholipase C inhibitor) inhibited lipopolysaccharide-stimulated but not basal cytokine-induced neutrophil chemoattractant production, while verapamil (a calcium channel blocker), TMB-8 (an inhibitor of calcium release from endoplasmic reticulum), and W7 (calmodulin antagonist) completely abrogated the chemoattractant production. Altered intracellular calcium, due to Ca2+ efflux from intracellular stores, may be involved in the "priming" of macrophages to release cytokine-induced neutrophil chemoattractant following triggering with lipopolysaccharide during acute cerulein pancreatitis.

Topics: Acute Disease; Animals; Calcium; Calcium Channel Blockers; Ceruletide; Chemokines, CXC; Chemotactic Factors; Growth Substances; Intercellular Signaling Peptides and Proteins; Lipopolysaccharides; Macrophages, Peritoneal; Male; Pancreatitis; Rats; Rats, Wistar; RNA, Messenger

Overexpression of transforming growth factors (TGF) in acute pancreatitis (AP) suggested that these substances play an important role in pancreatic repair and remodeling but the contribution of epidermal growth factor (EGF), that is well known to promote cell growth and regeneration, has not been investigated. The aim of this study was to compare the gene and immunohistochemical expression of EGF and TGF-beta1, cell proliferation, and biochemical parameters in AP induced by infusion of a supramaximal dose of caerulein in rats. The rats were sacrificed at 0, 12, 24, 48, 72 h, 5 and 10 days after the termination of caerulein infusion. Pancreatic tissue DNA synthesis, cell proliferation, histological and immunohistochemical assessments and plasma amylase were estimated following induction of AP. The mRNA expression for EGF and TGF-beta1 was evaluated by reverse transcription-polymerase chain reaction. During 10 days of the study after induction of AP a gradual normalization of biochemical and histological parameters was observed. DNA synthesis and cell proliferation which were significantly decreased at 0 and 24 h, increased significantly at 48 and 72 h, and then gradually decreased reaching at day 10 the values similar to those of vehicle-treated control rats. In these control rats the EGF mRNA or immunohistochemical expression was not detected, while the TGF-beta1 expression was weak. After induction of AP, the mRNA and immunohistochemical expression of EGF showed an increase during the initial 5 days, while those of TGF-beta1 showed a marked increase between 0 and 48 h and then again at day 10. We confirm that: (1) the expression of TGF-beta1 during AP is biphasic with an initial increase probably related to pancreatic damage and inhibition of cell proliferation and with the later phase of increase accompanied by the stimulation of the synthesis of extracellular matrix components and (2) AP is accompanied by an induction of synthesis of EGF that occurs in the initial phase of AP, probably limiting the extent of AP, and enhancing the stimulation of the pancreatic repair and regeneration.

Topics: Acute Disease; Animals; Biomarkers; Cell Division; Ceruletide; DNA; Epidermal Growth Factor; Gastrointestinal Agents; Gene Expression; Immunohistochemistry; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; RNA, Messenger; Transforming Growth Factor beta

To test the hypothesis that the specific acute-phase response program of the pancreas is a powerful emergency defense mechanism that is beneficial during acute pancreatitis.. Prospective, randomized, controlled animal study.. Research laboratory in a university medical school.. Female Wistar rats, weighing 250 to 300 g.. An acute-phase response was induced in rats subjected to hyperstimulation with cerulein. The development of the acute-phase reaction was monitored by the expression of the pancreatitis-associated protein I. In control animals, no acute-phase response was induced. After the first experimental procedure at periods of 2, 48, or 168 hrs, the pancreas was challenged by inducing severe necrotizing pancreatitis with retrograde infusion of taurocholate into the pancreatic duct. The course of necrohemorrhagic pancreatitis and survival of the rats to the challenge was monitored with time.. Forty-eight hours after the onset of edematous pancreatitis, the acute-phase response was strong, as judged by the overexpression of mRNA, which encoded the pancreatitis-associated protein I, and the resulting increase in concentrations of this protein in the pancreas. When necrotizing pancreatitis was induced, the survival rate was significantly higher than in the corresponding control group. In contrast, expression of the pancreatitis-associated protein I was not detectable after 2 hrs, indicating that the acute phase had not fully developed, nor after 168 hrs when the acute phase had ended. In both cases, challenge by necrotizing pancreatitis led to similar survival rates in cerulein-treated and control rats.. The acute-phase response of the pancreas seems to be a powerful emergency defense mechanism against further pancreatic aggression, as shown by the improved survival of the animals. The factors mediating this protection are unknown. Due to the strong overexpression of the pancreatitis-associated protein I during the climax of the acute phase, this protein might be involved in the defense mechanism.

Topics: Acute Disease; Acute-Phase Proteins; Acute-Phase Reaction; Animals; Antigens, Neoplasm; Biomarkers, Tumor; Ceruletide; Disease Models, Animal; Female; Gastrointestinal Agents; Lectins, C-Type; Pancreas; Pancreatitis; Pancreatitis-Associated Proteins; Pancreatitis, Acute Necrotizing; Rats; Rats, Wistar

1-Cyano-2-hydroxy-3-butene (CHB) has been reported to cause cell death in rat pancreatic acini. In this report, we describe the time-dependent effects of CHB on mouse acinar cell apoptosis and the effects of CHB-induced acinar cell apoptosis on the severity of secretagogue-induced acute pancreatitis in mice. CHB administration to mice resulted in a time-dependent increase in pancreatic apoptosis, which was maximal 12 hours after CHB administration. The severity of pancreatitis was significantly reduced by prior CHB administration and maximal protection was observed when the caerulein injections were started 12 hours after CHB administration. These observations indicate that induction of apoptosis can reduce the severity of pancreatitis and they suggest that induction of pancreatic acinar cell apoptosis may be beneficial in the clinical management of acute pancreatitis.

Topics: Acute Disease; Alkenes; Amylases; Animals; Apoptosis; Ceruletide; Disease Models, Animal; Female; Mice; Nitriles; Pancreas; Pancreatitis; Rats; Time Factors

In acute pancreatitis, two different types of secretory phospholipase A2 (PLA2) have been found: pancreatic type I PLA2 and non-pancreatic type II PLA2. In this study a potent new PLA2 inhibitor effective against type II PLA2 was used in an experimental model of acute pancreatitis.. In 70 rats the efficacy of the compound was analysed in two experimental models of acute pancreatitis: cerulein- and taurocholate-induced acute pancreatitis, imitating mild and severe disease respectively. Serum rat type I PLA2 protein concentration and type I and type II PLA2 catalytic activities were measured while giving the inhibitor therapeutically. In a prophylactic protocol the effect on histology was analysed.. In the taurocholate model, type II PLA2 activity was found to be nine-fold higher than in the cerulein model (P < 0.002), whereas the activity of type I PLA2 was not increased. The inhibitor significantly decreased serum type II PLA2 activity in the taurocholate model of acute pancreatitis (P < 0.05) but type I PLA2 protein concentration and type I PLA2 activity were not affected. The inhibitor also reduced histological tissue damage, with significant differences at 3 and 12 h (P < 0.01).. The PLA2 inhibitor significantly reduced type II PLA2 activity and was able to protect the pancreas against tissue damage. PLA2 inhibition offers the possibility of a treatment for acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Edema; Enzyme Inhibitors; Female; Necrosis; Pancreatitis; Phospholipases A; Phospholipases A2; Rats; Rats, Wistar; Taurocholic Acid

Pancreatitis-associated protein I (PAP I) is a pancreatic secretory protein strongly expressed during acute pancreatitis in the rat and human. We hypothesized that its expression was part of a general and coordinated response of the organ against aggression. An opposite pattern of PAP I mRNA expression has recently been described in the mouse. The murine PAP I mRNA was described to be highly expressed in normal pancreas and down-regulated during pancreatitis. The important implications of these unexpected findings led us to investigate the expression of murine PAP I in cerulein-induced pancreatitis. Northern blot analysis demonstrated a very low level of PAP I mRNA in the healthy mouse pancreas and strong overexpression during acute pancreatitis. Western blot analysis confirmed that changes in pancreatic PAP I levels were parallel to those of the mRNA and the protein was localized by immunohistochemistry to the acinar cells. It was concluded that, during the course of acute pancreatitis, the pattern of PAP I expression in the mouse pancreas was comparable to that already observed in the rat and human. Although we have no explanation for the discrepancy between our results and those recently reported, the expression pattern of PAP I in the mouse exocrine pancreas described in the present study suggests that the pancreatic response to aggression might be conserved in mammals.

Topics: Acute Disease; Acute-Phase Proteins; Animals; Antigens, Neoplasm; Biomarkers, Tumor; Ceruletide; Gastrointestinal Agents; Immunohistochemistry; Lectins; Lectins, C-Type; Mice; Mice, Inbred BALB C; Pancreas; Pancreatitis; Pancreatitis-Associated Proteins; Proteins; RNA, Messenger; Up-Regulation

This study was designed to investigate hyperbaric oxygen (HBO) therapy as a treatment for managing animals with induced acute pancreatitis. Forty-five anesthetized male Sprague-Dawley rats were studied. A severe acute pancreatitis model was established by combining an intravenous infusion of cerulein (15 microg/kg/h) and an intraductal injection of 0.1 ml of glycodeoxycholic acid (5 mM). Pathology, serum amylase level, pancreatic malondiadehyde levels and water content of the lungs and the pancreas were used to evaluate the severity of disease. Moreover, an in vivo microscopic technique was used to investigate microcirculatory derangement in the pancreas, i.e., flow velocity and leukocytes sticking in postcapillary venules. HBO was delivered in three regimens, i.e., 100% oxygen at 2.5 absolute atmospheric pressure (AAP), 40% oxygen at 2.5 AAP, and 100% oxygen at 1 AAP, 6 h after the initiation of induction of acute pancreatitis. All animals survived until the end of the experiments. HBO significantly improved the pathologic conditions and pancreatic malondiadehyde levels. Concomitantly, it also significantly lessened the severity of lung edema and improved the microcirculatory environment in the pancreas. Our results support the findings that HBO therapy has a beneficial effect on pancreatic microcirculation and lung edema during acute pancreatitis in rats.

Topics: 3,4-Methylenedioxyamphetamine; Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Extravascular Lung Water; Glycodeoxycholic Acid; Hyperbaric Oxygenation; Lung; Male; Microcirculation; Pancreas; Pancreatitis; Pulmonary Edema; Rats; Rats, Sprague-Dawley; Water

Supramaximal stimulation of the pancreas with the CCK analog caerulein causes acute edematous pancreatitis. In this model, active trypsin can be detected in the pancreas shortly after the start of supramaximal stimulation. Incubation of pancreatic acini in vitro with a supramaximally stimulating caerulein concentration also results in rapid activation of trypsinogen. In the current study, we have used the techniques of subcellular fractionation and both light and electron microscopy immunolocalization to identify the site of trypsinogen activation and the subsequent fate of trypsin during caerulein-induced pancreatitis. We report that trypsin activity and trypsinogen-activation peptide (TAP), which is released on activation of trypsinogen, are first detectable in a heavy subcellular fraction. This fraction is enriched in digestive enzyme zymogens and lysosomal hydrolases. Subsequent to trypsinogen activation, both trypsin activity and TAP move to a soluble compartment. Immunolocalization studies indicate that trypsinogen activation occurs in cytoplasmic vacuoles that contain the lysosomal hydrolase cathepsin B. These observations suggest that, during the early stages of pancreatitis, trypsinogen is activated in subcellular organelles containing colocalized digestive enzyme zymogens and lysosomal hydrolases and that, subsequent to its activation, trypsin is released into the cytosol.

Topics: Acute Disease; Animals; Cathepsin B; Cell Fractionation; Ceruletide; Edema; Enzyme Activation; Immunohistochemistry; Kinetics; Lysosomes; Male; Oligopeptides; Pancreas; Pancreatitis; Rats; Rats, Wistar; Subcellular Fractions; Time Factors; Trypsin; Trypsinogen; Vacuoles

The aim of this study was to test the hypothesis that apoptosis can protect against experimental pancreatitis and induction of apoptosis by an extract of Artemisia asiatica (DA-9601) is beneficial in cerulein-induced pancreatis in rats. Pancreatitis was induced in 6-week-old male SPF Sprague-Dawley rats by two intravenous (i.v.) administrations of 40 microg/kg cerulein. To investigate the effects of DA-9601 on the severity of pancreatitis and extent of apoptosis, rats were treated with intragastric DA-9601, 30 mg/kg (D30), 100 mg/kg (D100), or 300 mg/kg (D300), intraperitoneal superoxide dismutase, 10,000 U/kg (SOD), and i.v. gabexate mesilate, 40 mg/kg (Foy), three times (30 min before cerulein injection, 30 and 90 min after cerulein injection). The control group was administered vehicle alone. Ten rats were included in each treatment group and control group. Rats were sacrificed 5 h after cerulein treatment. Serum amylase, histological activity index (HAI), pancreatic lipid peroxide levels, and apoptotic index [in situ hybridization by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL)] were determined. Gel electrophoresis was performed for the presence of DNA fragmentations. The results were as follows. Serum amylase was significantly increased in all cerulein-treated groups compared to normal controls (p < 0.001). The HAI was significantly decreased in only the D300 group compared to the controls (p < 0.05). The apoptotic index of the cerulein-alone group was 3.8 +/- 2.7, but the mean apoptotic indexes of the SOD and Foy groups were 16.4 +/- 4.6 and 13.3 +/- 1.8, respectively, a significant increase (p < 0.01). The apoptotic index was more significantly increased in the DA-9601-treated groups, dose dependently (8.4 +/- 3.4 in D30, 14.8 +/- 4.3 in D100, 24.2 +/- 4.7 in D300). A smearing pattern of DNA electrophoresis was noted in the DA-9601-treated groups. In conclusion, DA-9601, an extract of Artemisia, induced apoptosis of pancreatic acinar cells dose dependently and concomitantly attenuated the severity of pancreatitis.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Artemisia; Ceruletide; DNA Fragmentation; Gabexate; Lipid Peroxidation; Male; Organ Size; Pancreas; Pancreatitis; Plant Extracts; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Specific Pathogen-Free Organisms; Superoxide Dismutase

To assess diaphragmatic function in vitro during experimental cerulein-induced acute pancreatitis.. Prospective, randomized, controlled animal trial.. Research laboratory at a large university medical center.. Twenty male Sprague-Dawley rats, weighing 180 to 200 g.. Sodium chloride 0.9% or cerulein (5 microg/kg/hr) was infused for 6 hrs.. Isometric force generated during electrical stimulation of costal diaphragmatic strips was measured 6 hrs after the end of infusion. Diaphragmatic strength was assessed at different frequencies (10, 20, 30, 50, and 100 Hz). Endurance index was the time until the force generated during the 30 Hz repetitive stimulation decreased to 50% of the initial value (T50%). Histologic examination of the diaphragm was performed. A decrease averaging 40% in diaphragmatic strength generation was observed for all frequencies of stimulation in the pancreatitis group. Compared with the control group, this decrease was associated with a reduction in T50% (30.9 +/- 12.5 [SD] and 46.4 +/- 10.8 secs in pancreatitis and control, respectively; p< .05). No histologic alteration of the diaphragm was observed.. Acute pancreatitis induced marked diaphragmatic dysfunction. Although the precise mechanisms responsible for this alteration are not precisely determined, diaphragmatic dysfunction may play a role in pancreatitis-associated respiratory failure.

Topics: Acute Disease; Animals; Ceruletide; Diaphragm; Disease Models, Animal; Electric Stimulation; Gastrointestinal Agents; Male; Muscle Contraction; Pancreatitis; Physical Exertion; Prospective Studies; Random Allocation; Rats; Rats, Sprague-Dawley

Protective effect of trifluoroacetyl-L-lysyl-L-alaninanilide hydrochloride (compound 1), a pancreatic elastase inhibitor, on three types of acute pancreatitis models was examined in rats. Mild, moderate and severe acute pancreatitis were induced by cerulein, the closed duodenal loop method and retrograde injection of a taurocholate plus trypsin solution into the pancreatic duct, respectively. Intravenous infusion of compound 1 at a dose of 30 mg/kg/hr resulted in lower increases in serum amylase, lipase, blood urea nitrogen (BUN) and creatinine levels in rats with mild cerulein-induced edematous pancreatitis. Compound 1 had no beneficial effect on pancreatitis in rats with moderate pancreatitis. In rats with severe pancreatitis, prophylactic treatment of compound 1 (30 mg/kg/hr) reduced both elevated serum BUN level and ascitic volume, and it histologically inhibited the extent of pancreatic edema and hemorrhage. These results suggest that pancreatic elastase is partially responsible for pancreatic edema and hemorrhage exhibited by rats with severe acute pancreatitis.

Topics: Acute Disease; Anilides; Animals; Ceruletide; Dipeptides; Disease Models, Animal; Kidney; Liver; Lung; Male; Pancreas; Pancreatic Elastase; Pancreatitis; Rats; Rats, Wistar; Serine Proteinase Inhibitors

Oxidative stress has been proposed to play a role in the early events of acute pancreatitis, and metallothionein (MT) can provide protection against oxidative stress. Using transgenic mice, we characterized the effects of depletion of MT-I and -II, or overexpression of MT-I, on pancreatic responses during cerulein-induced acute pancreatitis. In MT-I/-II knockout mice, repeated injections of cerulein caused (a) higher serum amylase levels at 3 and 7 h after the initiation of acute pancreatitis; (b) earlier and stronger upregulation of oxidative stress-responsive genes, including heme oxygenase (HO)-1 and c-fos; and (c) exacerbated tissue damage (edema and polymorphonuclear neutrophil infiltration) compared with nontransgenic 129/SvCPJ mice. Total pancreatic glutathione (GSH + GSSG) content was similar between the knockout and nontransgenic 129/SvCPJ mice. Interestingly, during acute pancreatitis, CD-1 mice pretreated with L-buthionine-[S,R]-sulfoximine (BSO), which dramatically depleted pancreatic GSH, also had more severe pancreatitis, based on the same three criteria listed above, relative to untreated controls. No effects were observed with BSO treatment alone. Finally, during cerulein-induced acute pancreatitis, MT-I overexpressing transgenic mice (>20-fold increase in pancreatic MT-I content) had lower serum alpha-amylase levels between 7 and 24 h and delayed upregulation of HO-1 mRNA levels, but no difference in c-fos mRNA induction relative to the appropriate strain of nontransgenic mice. Diminished tissue damage (particularly cellular necrosis) was noted in these MT-I overexpressing transgenic mice. Total pancreatic GSH content was similar in these transgenic and nontransgenic mice during cerulein-induced acute pancreatitis. These studies suggest that pancreatic MT can function as an intracellular antioxidant as does GSH and that these intracellular antioxidants play a protective role during cerulein-induced acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Buthionine Sulfoximine; Ceruletide; Glutathione; Heme Oxygenase (Decyclizing); Male; Metallothionein; Mice; Mice, Knockout; Mice, Transgenic; Pancreas; Pancreatitis; Proto-Oncogene Proteins c-fos; RNA, Messenger

The effects of a new benzodiazepine-derivative, cholecystokinin receptor antagonist, TS-941, on experimental acute pancreatitis were studied in rats. Hemorrhagic pancreatitis was induced by an infusion of a mixture of trypsin and taurocholate into the pancreatic duct. Edematous pancreatitis was induced by intraperitoneal injection of 40 microg/kg body weight of cerulein at 0 and 1 h after the start of the experiment. TS-941 (3 mg/kg) was injected subcutaneously immediately and 3 h after the induction of pancreatitis. In trypsin-taurocholate-induced pancreatitis, TS-941, with or without the synthetic trypsin inhibitor ONO-3403, had no beneficial effects on the survival rate, pancreatic wet weight, and serum pancreatic enzymes. In cerulein-induced pancreatitis, the treatment with TS-941 significantly reduced the increases of pancreatic wet weight and serum amylase and lipase. Plasma trypsinogen activation peptide (TAP) significantly rose 1 h after the first injection of cerulein. TS-941 inhibited the liberation of TAP in cerulein-induced pancreatitis. These results show that TS-941 is effective for prevention of cerulein-induced edematous pancreatitis. ONO-3403 has beneficial effects on trypsin-taurocholate-induced hemorrhagic pancreatitis, but the combination of TS-941 and ONO-3403 has no additive effect.

Topics: Acute Disease; Allylglycine; alpha-Macroglobulins; Amylases; Animals; Benzamidines; Benzodiazepines; Ceruletide; Drug Therapy, Combination; Lipase; Male; Oligopeptides; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Receptors, Cholecystokinin; Taurocholic Acid; Trypsin

Recent reports suggest that platelet-activating factor (PAF) plays a role in pancreatitis and pancreatitis-associated lung injury. In this study, the effects on these processes of termination of PAF action by recombinant PAF-acetylhydrolase (rPAF-AH) were investigated.. Rats were given rPAF-AH and then infused with a supramaximally stimulating dose of cerulein to induce mild pancreatitis. Opossums underwent biliopancreatic duct ligation to induce severe pancreatitis, and rPAF-AH administration was begun 2 days later.. In mild, secretagogue-induced pancreatitis, rPAF-AH given before the cerulein reduced hyperamylasemia, acinar cell vacuolization, and pancreatic inflammation but did not alter pancreatic edema or pulmonary microvascular permeability. In severe, biliopancreatic duct ligation-induced pancreatitis, rPAF-AH delayed and reduced the extent of inflammation and acinar cell injury/necrosis and completely prevented lung injury even though the rPAF-AH administration was begun after the onset of pancreatitis.. PAF plays an important role in the regulation of pancreatic injury but not pancreatic edema or increased pulmonary microvascular permeability in mild, secretagogue-induced pancreatitis. PAF plays a critical role in the regulation of progression of pancreatic injury and mediation of pancreatitis-associated lung injury in severe biliary pancreatitis. Amelioration of pancreatitis and prevention of pancreatitis-associated lung injury can be achieved with rPAF-AH even if treatment is begun after pancreatitis is established.

Topics: 1-Alkyl-2-acetylglycerophosphocholine Esterase; Acute Disease; Animals; Bile Ducts; Ceruletide; Disease Models, Animal; Gastrointestinal Agents; Ligation; Lung; Lung Diseases; Male; Opossums; Pancreas; Pancreatic Ducts; Pancreatitis; Phospholipases A; Platelet Activating Factor; Rats; Rats, Wistar; Recombinant Proteins

The pathological activation of zymogens within the pancreatic acinar cell plays a role in acute pancreatitis. To identify the processing site where activation occurs, antibodies to the trypsinogen activation peptide (TAP) were used in immunofluorescence studies using frozen sections from rat pancreas. Saline controls or animals receiving caerulein in amounts producing physiological levels of pancreatic stimulation demonstrated little or no TAP immunoreactivity. However, after caerulein hyperstimulation (5 micrograms. kg-1. h-1) for 30 min and the induction of pancreatitis, TAP immunoreactivity appeared in a vesicular, supranuclear compartment that demonstrated no overlap with zymogen granules. The number of vesicles and their size increased with time. After 60 min of hyperstimulation with caerulein, most of the TAP reactivity was localized within vacuoles >/=1 micrometer that demonstrated immunoreactivity for the granule membrane protein GRAMP-92, a marker for lysosomes and recycling endosomes. Pretreatment with the protease inhibitor FUT-175 blocked the appearance of TAP after hyperstimulation. These studies provide evidence that caerulein hyperstimulation stimulates trypsinogen processing to trypsin in distinct acinar cell compartments in a time-dependent manner.

Topics: Acute Disease; Animals; Biomarkers; Ceruletide; Endosomes; Lysosomes; Male; Membrane Proteins; Oligopeptides; Organelles; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Time Factors; Trypsinogen; Vacuoles

To investigate the pathobiology of severe acute pancreatitis, we studied the expression of inducible nitric oxide synthase (iNOS) in peritoneal macrophages of experimental pancreatitis. Taurocholate (TCA) pancreatitis and cerulein (CE) pancreatitis were used as models of lethal and self-limited pancreatitis, respectively, and the mechanism of iNOS expression in peritoneal macrophages was studied. Serum nitrate and nitrite (NOx) concentrations increased during the course of TCA pancreatitis, and iNOS-immunoreactivity was detected in the peritoneal macrophages 12 h after the induction of TCA pancreatitis, but these phenomena were not observed in CE pancreatitis. Despite the difference in the iNOS expression, the iNOS messenger RNA (mRNA) and the activation of nuclear factor-kappa B (NF-kappa B) were detected in the peritoneal macrophages of both pancreatitis models. The supernatant of TCA pancreatitis ascites could induce iNOS in the peritoneal macrophages of normal rats in vitro, but the peritoneal lavage fluid of CE pancreatitis rats could not. The results indicated that there may be qualitative or quantitative differences in the macrophage activation between the two types of experimental pancreatitis and suggested that the ascites of rats with lethal acute pancreatitis contains some soluble factors that activate the macrophage/monocyte system and cause an overproduction of NO by the iNOS expression.

Topics: Acute Disease; Animals; Ascitic Fluid; Ceruletide; Enzyme Inhibitors; Gene Expression; Lipopolysaccharides; Macrophages, Peritoneal; Male; NF-kappa B; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Pancreatitis; Rats; Rats, Wistar; RNA, Messenger; Taurocholic Acid

Nitric oxide (NO) blockade by L-nitroarginine methyl ester (L-NAME) inhibits pancreatic secretion in vivo and aggravates caerulein induced pancreatitis. Nitric oxide synthase (NOS) is present in pancreatic islets, endothelium, and nerve fibres. L-NAME blocks all known NOS isoforms.. To investigate the source of NO blocked by L-NAME that inhibits amylase secretion.. Amylase output was measured in rats in response to caerulein (0.1-50 microg/kg) alone or with indazole. Baseline secretion and the response to supramaximal caerulein were also examined after administration of indazole, L-NAME, haemoglobin, or aminoguanidine under continuous blood pressure measurement. In separate experiments, pancreatic secretion was measured after blockade of afferent nerve fibres by either systemic or local capsaicin. The effect of neural NOS inhibition on caerulein induced pancreatitis was also investigated.. L-NAME, haemoglobin, and supramaximal caerulein (10 microg/kg) increased blood pressure, whereas indazole and suboptimal caerulein (0.1 microg/kg) did not. Indazole and capsaicin decreased basal amylase output. L-NAME and haemoglobin reduced basal amylase output to a lesser extent and potentiated the inhibitory response to supramaximal caerulein. In contrast, full neural NOS inhibition by L-NAME partially reversed the expected caerulein induced suppression of amylase output. This effect was reproduced by indazole and capsaicin. Indazole did not alter responses to either optimal (0.25 microg/kg) or suboptimal (0.1 microg/kg) caerulein, nor, in contrast with L-NAME, aggravate the outcome of caerulein induced pancreatitis.. Reduction of circulating NO availability, probably of endothelial origin, is responsible for the decrease in amylase secretion observed in the early response to L-NAME. Nitrergic neurotransmission plays an important role in the control of pancreatic secretion and may induce opposite effects to endothelial NOS activity.

Topics: Acute Disease; Amylases; Animals; Capsaicin; Ceruletide; Enzyme Inhibitors; Gastrointestinal Agents; Hemoglobins; Male; Nerve Fibers; Neurons, Afferent; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

Overproduction of nitric oxide (NO) via induction of the inducible NO synthase (iNOS) is an important factor in the haemodynamic disturbances of several inflammatory states.. To identify the role of NO in a caerulein induced model of acute pancreatitis in the rat.. Arterial blood pressure and plasma NO metabolites were measured at zero and seven hours in adult male Wistar rats administered caerulein (n=10) or saline (n=10). Pancreatic activity of NOS (inducible and constitutive) was assayed biochemically. The pancreatic expression and cellular localisation of NOS and nitrotyrosine (a marker of peroxynitrite induced oxidative tissue damage) were characterised immunohistochemically.. Compared with controls at seven hours, the pancreatitis group displayed raised plasma NO metabolites (mean (SEM) 70.2 (5.9) versus 22.7 (2.2) micromol/l, p<0.0001) and reduced mean arterial blood pressure (88.7 (4.6) versus 112.8 (4.1) mm Hg, p=0.008). There was notable iNOS activity in the pancreatitis group (3.1 (0.34) versus 0.1 (0.01) pmol/mg protein/min, p<0.0001) with reduced constitutive NOS activity (0.62 (0.12) versus 0.96 (0.08) pmol/mg protein/min, p=0.031). The increased expression of iNOS was mainly localised within vascular smooth muscle cells (p=0.003 versus controls), with positive perivascular staining for nitrotyrosine (p=0.0012 versus controls).. In this experimental model of acute pancreatitis, iNOS induction and oxidative tissue damage in the pancreas is associated with raised systemic NO and arterial hypotension. Excess production of NO arising from the inducible NO synthase may be an important factor in the systemic and local haemodynamic disturbances associated with acute pancreatitis.

Topics: Acute Disease; Animals; Blood Pressure; Ceruletide; Endotoxins; Immunohistochemistry; Male; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Pancreatitis; Rats; Rats, Wistar

Alcohol consumption causes acute alcohol pancreatitis and worsens the prognosis; however, there is no useful model for elucidation of the mechanism underlying this worsening. The aim of our study was to establish a new prognostic model of acute alcohol pancreatitis in rats. To ascertain the effect of continuous infusion of ethanol on each phase, i.e., progression and recovery, in caerulein-induced pancreatic injury in rats, we infused a physiological or supramaximal dose of caerulein intravenously to conscious Wistar rats for up to 6 h (time: 0-6 h) with or without ethanol infusion for 9 h (time: 3-12 h). Ethanol did not induce the pancreatic injury alone or when combined with a physiological dose of caerulein. In the progression phase, ethanol infusion for 3 h (time: 6 h) did not aggravate the pancreatic injury induced by a supramaximal dose of caerulein in terms of plasma amylase and lipase activities but did increase the pancreatic calcium level. In the recovery phase, however, ethanol infusion for 9 h (time: 12 h) significantly restrained the recovery from pancreatic injury as monitored in terms of these activities. Further, ethanol infusion for 9 h significantly increased the cumulative urinary excretion of amylase from 12 to 27 h but did not do the same from 0 to 12 h. In the histological evaluation at 27 h, the induction of acinar cell vacuolization and dilation of the glandular lumina and ducts were significant in the caerulein plus ethanol-treated group. Our findings suggest that ethanol administration delays the recovery rather than worsens the progression in acute pancreatic injury induced by exocrine hyperstimulation, and we consider our experimental model to be a useful tool for studying the pathogenesis of worsening prognosis in acute alcohol pancreatitis.

Topics: Acute Disease; Amylases; Animals; Calcium; Central Nervous System Depressants; Ceruletide; Disease Progression; Ethanol; Lipase; Male; Pancreas; Pancreatitis, Alcoholic; Prognosis; Rats; Rats, Wistar; Stimulation, Chemical

Ectopic protease activation, microcirculatory changes, and leucocyte activation are the main events in the pathogenesis of acute pancreatitis. Nitric oxide (NO) is known to be a key mediator in the normal and inflamed pancreas.. To investigate the targets on which NO exerts its effect in caerulein induced pancreatitis.. Acute pancreatitis was induced in rats which additionally received either the NO synthase substrate, L-arginine; the NO donor, sodium nitroprusside; or the NO synthase inhibitor, N-nitro-L-arginine methyl ester (L-NAME). At six hours, pancreatic injury (oedema, leucocyte content, ectopic trypsinogen activation) was analysed and pancreatic oxygenation and perfusion were determined. A direct influence of NO on amylase secretion and trypsinogen activation was evaluated separately in vitro.. Both NO donors reduced the grade of inflammation. L-NAME increased the severity of inflammation, while decreasing pancreatic tissue oxygenation. Although neither amylase secretion nor intracellular trypsinogen activation in caerulein stimulated pancreatic acini was influenced by either NO donors or inhibitors, both NO donors decreased intrapancreatic trypsinogen activation peptide (TAP) and pancreatic oedema in vivo, and L-NAME increased TAP.. NO protects against injury caused by pancreatitis in the intact animal but has no discernible effect on isolated acini. It is likely that in pancreatitis NO acts indirectly via microcirculatory changes, including inhibition of leucocyte activation and preservation of capillary perfusion.

Topics: Acute Disease; Animals; Arginine; Cells, Cultured; Ceruletide; Enzyme Inhibitors; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroprusside; Oligopeptides; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Rats, Wistar; Vasodilator Agents

The purpose of the study was to compare the morphological alterations of the liver in two models of acute pancreatitis: caerulein-induced (edematous) and taurocholate-induced (necro haemorrhagic one). The experiments were performed on 24 male, Wistar rats, weighing 240-260 g. In group I (n = 8) the supramaximal stimulation with i.v. caerulein (5 micrograms/kg/h) during 12 h was applied (C-AP). Control animals (group II, n = 4) received i.v. saline (C-C). In group III (n = 8) 5% sodium taurocholate (0.2 ml/min) was injected into the bile-pancreatic duct during sterile laparotomy (T-AP). In group IV (n = 4) animals were sham operated (T-C). The specimens of the liver were excised after decapitation of rats at 12 h after beginning of caerulein infusion or intraductal injection of sodium taurocholate. The light and electron microscopy was performed. The marked hepatic lesion were found in both variants of experimental pancreatitis, however they were far more advanced in taurocholate pancreatitis. In light microscopy the dispersed foci of colliquative necrosis, degeneration of hepatocytes, swelling of Kupffer cells predominated in taurocholate pancreatitis. The glycogen deposits were depleted but lipid droplets were increased in size and number. The swelling of mitochondria, degeneration of their matrix and cristae, increase of autophagocytosis and numerous lysosomes, the lesions of sinusoids with increased activity of phagocytic cells were more evident in taurocholate pancreatitis--(more severe model of the disease). These findings document severe injury to the liver in acute pancreatitis depending on the severity of inflammatory process in pancreas. They also suggest that the liver could be not only passive target of pancreatogenic noxa in acute pancreatitis, but it could be also a defensive barrier against spreading of injuring agents on other system. This role seems to be especially evident in more severe form--taurocholate induced pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Culture Techniques; Disease Models, Animal; Gastrointestinal Agents; Liver; Male; Microscopy, Electron; Pancreatitis; Rats; Rats, Wistar; Reference Values; Taurocholic Acid

The role of bronchoalveolar macrophages (BAMs) in the aggravation of cerulein-induced pancreatitis was studied by measuring expression of cytokine-induced neutrophil chemoattractant (CINC) in vitro. Pancreatitis was induced by four intramuscular injections of cerulein (50 microg/kg at 1-hr intervals). Pancreatitis rats were injected intraperitoneally with 30 mg/kg lipopolysaccharide (LPS) 6 hr following the first cerulein injection as a septic challenge. Rats were divided into four groups: group I, nonpancreatitis without LPS; group II, pancreatitis without LPS; group III, nonpancreatitis with LPS; and group IV, pancreatitis with LPS. Hyperactivity of BAMs in response to LPS was assessed as a function of in vitro CINC production. CINC concentrations of the serum and bronchoalveolar lavage fluid in group IV were significantly higher than those in groups I, II, and III. BAMs in group II harvested 6 hr following the first cerulein injection had significantly greater CINC production than those in group I. Northern blot analysis revealed abundant CINC mRNA transcripts in BAMs from groups III and IV. Additionally, myeloperoxidase activity in the lung of group IV rats 8 and 12 hr following the first cerulein injection was significantly higher than that in group I, II, and III rats. Significant differences in static lung compliance in group IV were found compared with groups I, II, and III. These results indicate that BAMs from rats with cerulein-induced pancreatitis were primed and had enhanced release of CINC following LPS exposure. Enhanced expression of CINC may modulate the pathogenesis of pancreatitis-associated lung injury complicated with sepsis.

Topics: Acute Disease; Animals; Blotting, Northern; Bronchoalveolar Lavage Fluid; Ceruletide; Chemokines, CXC; Chemotactic Factors; Growth Substances; In Vitro Techniques; Intercellular Signaling Peptides and Proteins; Lipopolysaccharides; Lung; Lung Compliance; Macrophages, Alveolar; Male; Pancreatitis; Peroxidase; Rats; Rats, Wistar

In chronic pancreatitis the exocrine pancreatic tissue is replaced by extracellular matrix deposits from fibroblasts. We have stimulated fibrogenesis in the pancreas by inducing a single episode of cerulein pancreatitis (10 microg/kg/h for 12 h). We have used a spectrophotometrical assay to measure the tissue hydroxyproline content and immunohistochemistry to study the transient deposition of extracellular matrix in the pancreas. We have investigated whether a potent prolyl 4-hydroxylase inhibitor (HOE-077) can influence the deposition of extracellular matrix in the pancreas. Three days after the induction of the pancreatitis we found the maximum increase in pancreatic hydroxyproline content (by 63%), the maximum decrease in total protein content and amylase activity (by -39 and -86%, respectively), as well as a significant increase in DNA content and the deposition of interstitial collagen fibers on electron microscopy. By immunohistochemistry the largest expansion of extracellular matrix components was found for fibronectin. HOE 077, regardless of the concentration administered, failed to affect any of these parameters. We conclude that the induction of a single episode of cerulein pancreatitis and the serial determination of pancreatic hydroxyproline content represents a simple method to induce and monitor experimental fibrogenesis in the pancreas. Prolyl 4-hydroxylase inhibition did not affect the course of extracellular matrix deposition in the pancreas.

Topics: Acute Disease; Animals; Ceruletide; DNA; Dose-Response Relationship, Drug; Enzyme Inhibitors; Extracellular Matrix; Extracellular Matrix Proteins; Gastrointestinal Agents; Hydroxyproline; Immunohistochemistry; Male; Microscopy, Electron; Pancreatitis; Procollagen-Proline Dioxygenase; Pyridines; Rats; Rats, Wistar; Spectrophotometry

This study was designed to evaluate the protective effect of a peptide leukotriene receptor antagonist, pranlukast hydrate, against pancreatic injuries during acute pancreatitis.. Acute pancreatitis was induced in rats by intravenous infusion of a supramaximal dose of cerulein (5 micrograms/kg h for 4 h). In this model marked hyperamylasemia, a significant increase in pancreatic water content, and a significant increase in pancreatic microvascular leakage of Evans blue dye were observed. Pancreatic subcellular redistribution of the lysosomal enzyme cathepsin B from the lysosomal fraction to the zymogen fraction was also observed.. Pretreatment with pranlukast hydrate at a dose of 10 micrograms/kg (twice, 8 and 4 h before cerulein infusion) significantly inhibited these pancreatic injuries, including hyperamylasemia, increased pancreatic microvascular permeability, and redistribution of cathepsin B in pancreatic acinar cells.. These results suggest that peptide leukotrienes may be involved in the pathogenesis of acute pancreatitis in the early stage of the disease and that peptide leukotriene receptor antagonist might be of therapeutic value for treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Body Water; Cathepsin B; Ceruletide; Chromones; Edema; Leukotriene Antagonists; Male; Pancreas; Pancreatic Diseases; Pancreatitis; Rats; Rats, Wistar

Infusion of a supramaximally stimulating dose of the pancreatic secretagogue caerulein (10 micrograms.kg-1.h-1) for 4 h induces interstitial edematous acute pancreatitis in rats. This model of acute pancreatitis is associated with evidence of acute lung injury, including sequestered neutrophils within the pulmonary microvasculature, increased microvascular permeability, and interstitial pulmonary edema. Infusion of prostaglandin E1 (PGE1; 50 ng.kg-1.min-1) along with caerulein does not alter the severity of secretagogue-induced pancreatitis, but it does reduce the severity of pancreatitis-associated acute lung injury. The rise in lung weight, lung water content, and pulmonary microvascular permeability and the sequestration of neutrophils within the pulmonary microvasculature that accompany secretagogue-induced pancreatitis are all reduced by infusion of PGE1. Infusion of PGE1 does not interfere with polymorphonuclear neutrophil sequestration in the pancreas or reduce the enhanced expression of CD11b/c receptors on circulating neutrophils. Our observations indicate that PGE1 reduces the severity of pancreatitis-associated acute lung injury by preventing neutrophil sequestration within the lung. We speculate that PGE1 interferes with neutrophil sequestration by dilating pulmonary vasculature, increasing pulmonary flow rate, and reducing neutrophil-endothelial cell interaction and attachment.

Topics: Acute Disease; Alprostadil; Animals; Ceruletide; Integrin alphaXbeta2; Lipopolysaccharides; Lung; Lung Diseases; Macrophage-1 Antigen; Male; Neutrophils; Pancreas; Pancreatitis; Rats; Rats, Wistar

Pancreatic exocrine function was examined in rats during the early stage of acute pancreatitis induced by four subcutaneous injections of 20 micrograms/kg body weight of cerulein at hourly intervals. Basal pancreatic fluid secretion at 6 hr after the first of four cerulein injections was significantly elevated (27.6 +/- 3.7 vs 17.4 +/- 2.1 microliters/30 min in control, P < 0.01) and further increased with time, reaching the peak level at 24 hr (105.1 +/- 4.6 microliters/30 min). Intravenous infusion of loxiglumide (50 mg/kg body wt/hr), atropine (100 micrograms/kg body wt/hr), or anti-secretin serum did not modify the fluid hypersecretion observed at 24 hr after induction of acute pancreatitis. Loxiglumide, when given 30 min before the first cerulein injection, markedly reduced fluid secretion, but could not inhibit the fluid hypersecretion when applied after the last cerulein injection. Leakage of Evans blue dye into pancreatic juice was slightly but significantly increased in postpancreatitic rats compared with that in the control rats (1.30 +/- 0.17 vs 0.75 +/- 0.08 micrograms/ml, P < 0.01), whereas that in the pancreas was not different from the control rats. In vivo labeling with 5-bromo-2'-deoxyuridine showed active proliferation of acinar and ductular cells at 6 hr. In addition, the fluid was rich in chloride (137.1 +/- 2.5 at 24 hr vs 92.4 +/- 3.3 meq/liter in control, P < 0.01) but poor in bicarbonate concentration (39.0 +/- 2.0 at 24 hr vs 46.5 +/- 1.9 mmol/liter in control, P < 0.01), indicating acinar cell secretion. These results indicate that pancreatic fluid secretion during the early stage of acute pancreatitis induced by supramaximal doses of cerulein was markedly increased not by CCK-, secretin-, or cholinergic-dependent mechanisms but probably by acinar cell proliferation.

Topics: Acute Disease; Animals; Antibodies; Atropine; Bicarbonates; Cell Division; Ceruletide; Chlorides; Cholinergic Antagonists; Hormone Antagonists; Male; Pancreas; Pancreatic Juice; Pancreatitis; Proglumide; Proteins; Rats; Rats, Wistar; Receptors, Cholecystokinin; Secretin

Interleukin-1 beta (IL-1) is a proinflammatory cytokine which is produced within the pancreas during acute pancreatitis reaching levels which are toxic to many cell types. Since antagonism of this cytokine provides dramatic survival benefits during lethal pancreatitis, we hypothesized that IL-1 had direct secretagogue and cytolytic effects within the pancreas. The effect of IL-1 on pancreatic exocrine function and tissue viability was assessed in vivo by blockade of IL-1 with varying doses of IL-1 receptor antagonist (IL-1ra) prior to the induction of either moderate (caerulein-induced) or severe (choline deficient diet-induced) necrotizing pancreatitis. Subsequent in vitro studies were conducted to determine the direct effect of IL-1 on dispersed rat acini prepared through collagenase digestion. Amylase release was measured after a 30-min incubation with varying doses of recombinant IL-1 beta. Viability was determined in the presence of IL-1 via trypan blue exclusion at multiple time points. Blockade of the IL-1 receptor decreased pancreatic amylase release and tissue necrosis in both models of pancreatitis in a dose-dependent fashion (1.0 mg/kg, P = NS; 10 mg/kg, P < 0.05; 100 mg/kg, P < 0.05). Despite these in vivo findings, the addition of IL-1 to acini in vitro had no effect on exocrine function and failed to decrease acini viability (both, P = NS). Pancreatic amylase release and tissue necrosis are significantly attenuated during experimental pancreatitis by IL-1 antagonism. These changes do not appear to be due to the direct action of IL-1 on pancreatic acini and are likely due to more complex interactions between acini and cytokine-producing leukocytes.

Topics: Acute Disease; Amylases; Animals; Cell Survival; Ceruletide; Dose-Response Relationship, Drug; Exocrine Glands; Interleukin-1; Male; Mice; Necrosis; Pancreatitis; Receptors, Interleukin-1; Recombinant Proteins

Effects of a new cholecystokinin (CCK)A-receptor antagonist, T-0632 [sodium (S)-1-(2-fluorophenyl)-2, 3-dihydro-3-[(3-isoquinolinylcarbonyl) amino]-6-methoxy-2-oxo-1H-indole-3-propanoate], on caerulein-induced and pancreatic duct ligation-induced pancreatitis models were studied and compared with the CCKA-receptor antagonist loxiglumide and the orally active protease inhibitor camostate, respectively. In rats, orally administered T-0632 potently prevented the caerulein-induced increases in pancreatic digestive enzymes in plasma and suppressed the histological changes in the pancreas. The estimated ED50 values of T-0632 and loxiglumide were 0.0092 and 8.9 mg/kg, respectively. In dogs, T-0632 (0.1, 1 mg/kg, i.d.) prevented the caerulein-induced increase in plasma amylase activity in a dose-dependent manner. Loxiglumide (100 mg/kg, i.d.) did not show any preventive effects. In pancreatic duct ligation (6 hr)-induced pancreatitis of the rat, T-0632 (0.001-0.1 mg/kg, p.o.) partially prevented both the increase in plasma amylase activity and the histological changes in the pancreas, whereas camostate (10, 100 mg/kg, p.o.) did not show any preventive effects. In pancreatic duct ligation (3 hr)-induced pancreatitis, caerulein injection (1 microgram/kg, s.c.) caused a further increase in plasma amylase activity, and T-0632 (0.01, 0.1 mg/kg, p.o.) dose-dependently decreased the aggravation by caerulein. We conclude that T-0632 showed preventive effects on all of these pancreatitis models by oral or intraduodenal administration. These results suggest that CCK plays an important role in progression and aggravation of acute pancreatitis, and T-0632 may have a therapeutic value in these disease states.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Dogs; Esters; Female; Gabexate; Guanidines; Hormone Antagonists; Indoles; Ligation; Male; Pancreatic Ducts; Pancreatitis; Proglumide; Protease Inhibitors; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptor, Cholecystokinin A; Receptors, Cholecystokinin

Effects of green tea catechins (GTC) on cerulein-induced acute pancreatitis in rats were examined. The acute pancreatitis induced by cerulein (cerulein pancreatitis) was characterized by interstitial edema and vacuolation. When cerulein pancreatitis was induced, prior administration of 0.1% GTC in drinking water for 1 week before the induction significantly decreased the wet weight of the pancreas, the serum level of amylase, and the tissue concentration of lipid peroxides in the pancreas compared with those in nonmedicated rats supplied with plain tap water only. Furthermore, the pancreatic tissue alterations of the medicated rats were milder than those of the nonmedicated rats. These data suggest that GTC have a protective effect on the pathogenesis of cerulein pancreatitis.

Topics: Acute Disease; Amylases; Animals; Catechin; Ceruletide; Lipid Peroxides; Male; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Tea

Growth factors such as TGF-beta 1 and EGF are known to modulate the deposition of extracellular matrix components and tissue repair and to affect the cellular growth but their expression in the course of pancreatitis has not been studied. In this study we investigated the gene expression of TGF-beta 1 mRNA and EGF mRNA and other parameters of the pancreas including DNA synthesis, blood flow (PBF), tissue protein content and plasma amylase during the induction of acute pancreatitis. Supramaximal dose of cearulein (10 mg/kg/h s.c.) was infused for 5 h to induce pancreatitis. Animals were killed after 1, 2, 3, 4 and 5 h of infusion. The PBF was measured, blood samples were withdrawn to determine serum amylase concentration, biopsy samples were taken to measure the protein content and DNA synthesis. Expression of TGF-beta 1 and EGF mRNA was studied by reverse-transcription of polymerase chain reaction (RT-PCR). Caerulein infused caused a time-dependent decrease in DNA synthesis accompanied by gradual decrease of PBF and significant increase in pancreatic weight. The pancreatic protein content and plasma amylase showed progressive rise during 5 h of cearulein infusion. Histology revealed tissue edema, cellular vacuolization and prominent leukocyte infiltration after 3 h of cearulein infusion. TGF-beta 1 mRNA was strongly expressed at each time interval beginning from the 1 h after the start of cearulein infusion. In contrast, EGF mRNA was detected only at 5 h after induction of pancreatitis. We conclude that 1) the development of caerulein-induced pancreatitis results in the inhibition of pancreatic growth and the reduction in PBF accompanied by enhanced expression of TGF-beta 1; 2) The expression of EGF that was observed at the end of the induction of pancreatitis may indicate the initiation of pancreatic repair; 3) TGF-beta 1 seems to lead to subsequent induction of EGF that may stimulate the regeneration of injured pancreas.

Topics: Acute Disease; Amylases; Animals; Ceruletide; DNA; DNA Primers; Epidermal Growth Factor; Gene Expression Regulation; Male; Organ Size; Pancreas; Pancreatitis; Polymerase Chain Reaction; Protein Biosynthesis; Rats; Rats, Wistar; Regional Blood Flow; RNA; Transforming Growth Factor beta

TdT-mediated dUTP-biotin nick end labelling (TUNEL) has been widely used for detecting cells with DNA fragmentation or apoptotic cells. However, since the concept of apoptosis is based on cellular ultrastructure, it is important to identify the morphological features of TUNEL-positive cells. In this study, we performed TUNEL and electron microscopic observation on serial semithin and ultrathin sections of pancreas from bilaterally adrenalectomized rats with caerulein-induced pancreatitis. TUNEL-positive cells were identified with two different ultrastructural patterns. One was characteristic of apoptosis, with condensed nuclei, intact mitochondria, and zymogen granules. The other pattern was one of marked cellular degeneration, possibly representing the end stage of cell death. Cells which did not demonstrate these ultrastructural patterns were not labelled by the TUNEL method. The three-dimensional structure of TUNEL-positive cells was also investigated by confocal laser scanning microscopy (CLSM), which showed the apoptotic nuclei exhibited various three-dimensional structures. These results confirm the utility of the TUNEL method in detecting apoptosis; application of the technique reported in this study will contribute to the further characterization of individual TUNEL-positive cells.

Topics: Acute Disease; Animals; Apoptosis; Ceruletide; Deoxyuracil Nucleotides; Male; Microscopy, Confocal; Microscopy, Electron; Pancreas; Pancreatitis; Rats; Rats, Wistar; Tissue Embedding; Tissue Fixation

In human acute pancreatitis two different types of secretory phospholipase A2 (PLA2) have been found.. To analyse the specific pattern of distribution of these PLA2 activities and their pathophysiological role in experimental acute pancreatitis.. Catalytic activities of secretory type I (pancreatic) and type II (non-pancreatic) PLA2 and the protein concentration of immunoreactive pancreatic PLA2 (IR-PLA2) in serum and pancreatic tissue of rats with cerulein (mild form) and sodium taurocholate (severe form) induced acute pancreatitis were determined.. Cerulein infusion caused a significant increase in type I PLA2 activity (p < 0.01) and IR-PLA2 protein concentration (p < 0.01) in serum and pancreas, whereas type II PLA2 activity remained unchanged during the 12 hour observation period. Histology showed no significant tissue destruction. In sodium taurocholate induced acute pancreatitis type II PLA2 activity significantly increased, reaching values over 10-fold higher than controls (p < 0.01), whereas IR-PLA2 protein concentration and type I PLA2 activity were only marginally increased. In this severe model of acute pancreatitis significantly lower values were detected than in the control pancreas (p < 0.002) for PLA2 activity and IR-PLA2 protein concentration. Histology showed parenchymal and fat necroses with haemorrhage, oedema, and inflammatory cell infiltration.. Type I PLA2 activity is dependent on the IR-PLA2 protein concentration in serum and pancreatic tissue. The type II PLA2 activity is not stimulated by cerulein, which indicates an extra-acinar origin of this enzyme. Type II PLA2 activity is significantly increased in sodium taurocholate induced acute pancreatitis indicating its role in the local necrotising process and involvement in the systemic effects in severe acute pancreatitis.

Topics: Acute Disease; Animals; Catalysis; Ceruletide; Female; Pancreas; Pancreatitis; Phospholipases A; Phospholipases A2; Rats; Rats, Wistar; Taurocholic Acid

Transforming growth factor beta isoforms (TGF beta s) belong to a family of multifunctional regulators of cellular growth and differentiation. They are mitogenic and chemotactic for fibroblasts and are potent stimulators of extracellular matrix production (collagen) and deposition. Upregulation of TGF beta transcription has been reported for several in vivo systems during repair after injury.. To study the expression of the three mammalian isoforms of TGF beta (TGF beta 1-3) and their relation to collagen expression as a marker for fibroblast response in acute oedematous pancreatitis in rats.. Using northern blot analysis and immunohistochemistry, the expression and localisation of TGF beta isoforms, collagen, and amylase were analysed during the course of acute oedematous pancreatitis in rats, experimentally induced by intravenous caerulein infusion.. Induction of acute pancreatitis resulted in a biphasic peak pattern of expression of TGF beta 1, beta 2, and beta 3 mRNA, with a pronounced increase from day 1 to day 3 (sixfold, 2.5-fold, fivefold, respectively) and again from day 5 to day 7 (three-fold, 2.3-fold, 3.5-fold, respectively). The temporal changes in TGF beta mRNA identically paralleled the expression in collagen mRNA. In contrast, amylase mRNA expression, used as a general indicator of acinar cell integrity, was slightly decreased after induction of acute pancreatitis. Immunohistochemical analysis of pancreatitis tissue showed that increased expression of TGF beta s was mainly present in the pancreatic acinar and ductal cells; this was evident within one day after pancreatitis induction.. Overexpression of TGF beta s after induction of acute pancreatitis suggests a role for these proteins in pancreatic repair and remodelling. The increased levels of TGF beta s may help suppress immune activation, and may contribute to the increase in the extracellular matrix including collagen and to the repair of the pancreatic parenchyma.

Topics: Acute Disease; Amylases; Animals; Blotting, Northern; Ceruletide; Collagen; Edema; Gastrointestinal Agents; Gene Expression; Immunohistochemistry; Male; Pancreatitis; Rats; Rats, Wistar; RNA, Messenger; Transforming Growth Factor beta

Lipid peroxidation, which may be involved in the pathogenesis of acute pancreatitis, is usually assessed in vitro or indirectly using antioxidants or free radical scavengers. We assessed lipid peroxidation in an in vivo model by measuring ethane exhalation in two models of acute pancreatitis. Edematous acute pancreatitis was induced by a supramaximal intraperitoneal injection of cerulein. Necrotizing acute pancreatitis was induced by retrograde infusion of sodium taurocholate into the pancreaticobiliary duct. Rats were placed in closed chambers and ethane exhalation was measured in aliquots. Ethane exhalation was significantly increased (p < 0.002) in cerulein (n = 12)- but not in taurocholate (n = 6)-induced pancreatitis compared to controls (n = 12 and 6, respectively). Our results suggest that free radicals may play a role in the pathogenesis of edematous pancreatitis but do not play an important role in the progression to necrotizing pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Edema; Ethane; Lipase; Lipid Peroxidation; Male; Necrosis; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

Patients with severe acute pancreatitis die of complications closely related to the systemic activation of protease cascades.. To examine the effects of human C1 esterase inhibitor (C1 INH) and antithrombin III (AT III) on two experimental models of acute pancreatitis.. Oedematous pancreatitis was induced by continuous intravenous infusion of caerulein and haemorrhagic pancreatitis by retrograde injection of sodium taurocholate into the biliopancreatic duct. C1 INH and AT III were given intravenously, either before or after the induction of pancreatitis. Treatment with C1 INH and AT III had no beneficial effect on oedematous pancreatitis. On the other hand, combined C1 INH and AT III therapy improved the survival in haemorrhagic pancreatitis compared with treatment with human serum albumin. This reduction in mortality was found regardless of whether the treatment was given prophylactically or therapeutically.. Treatment with C1 INH and AT III represents a promising therapeutic concept for patients with severe haemorrhagic pancreatitis.

Topics: Acute Disease; Amylases; Animals; Antithrombin III; Biomarkers; Ceruletide; Complement C1 Inactivator Proteins; Drug Therapy, Combination; L-Lactate Dehydrogenase; Lactic Acid; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Serum Albumin

Insulin-like growth factors (IGFs) are important peptides involved in the regulation of cell growth and differentiation in many tissues. The ontogeny of IGF-I was examined in pancreata from 19-day rat fetuses, newborns and 5-, 11-, 26- and 70-day-old rats. For the regeneration studies two models were used: (i) 90% pancreatectomy was carried out and the rats were killed at 1, 2, 3 and 6 days after resection; (ii) acute pancreatitis was induced with caerulein (12 micrograms/body weight three times a day every 8 h for 2 days) and the rats were killed at 1, 2, 5, 7 and 9 days after the first injection. Total RNA was extracted by the guanidinium isothiocyanate method and Northern blots were performed using total RNA and labeled cRNA probes. Abundance of the different mRNA transcripts was estimated by densitometric scanning and normalized to the abundance of 18 S rRNA for each time point. Northern blot analysis during ontogeny showed four (0.8-1.2, 1.9, 4.7 and 7.5 kb) major transcripts in the rat pancreas and liver. Total IGF-I mRNA was 40-fold higher in the adult liver than in the adult pancreas. Moreover, in the liver, IGF-I mRNA levels were higher in the adult than in the fetus, whereas in the pancreas, the highest levels were observed around birth. During the first 3 days after pancreatectomy, a peak of maximal expression was observed after the second day. Densitometric analysis of each IGF-I mRNA species showed concomitant increases in all transcripts. After 6 days, all transcripts had returned to near-control values. IGF-I mRNA expression 2 days after pancreatectomy was 3.5-fold higher than in the newborn. During the first 2 days of acute pancreatitis induction, overexpression of IGF-I mRNA was observed. However, soon after the second day of caerulein treatment, the 7.5 kb transcripts remained elevated whereas those of the others regressed toward control values. Our results show that IGF-I mRNA is overexpressed in both models of pancreatic regeneration but downregulated in the normal adult pancreas.

Topics: Acute Disease; Animals; Animals, Newborn; Ceruletide; Down-Regulation; Female; Fetus; Insulin-Like Growth Factor I; Pancreas; Pancreatectomy; Pancreatitis; Pregnancy; Rats; Rats, Sprague-Dawley; Regeneration; RNA, Messenger

Recent studies indicated that expression of the housekeeping gene GAPDH is highly regulated during proliferation and differentiation. The objective of this study was to characterize by Northern blot the GAPDH mRNA expression in rat pancreas development and regeneration following acute pancreatitis induction by caerulein. Pancreatic GAPDH mRNA levels were the highest between fetal day 19 and the 11 postnatal day; they decreased to their lowest level after weaning on day 26. In acute pancreatitis, GAPDH mRNA levels were clearly increased 18 h after its initiation, were maximal during the first two days of induction and then decreased to control values after 9 days. These data demonstrate that overexpression of GAPDH may be implicated in pancreatic development, maturation and pancreas regeneration after acute pancreatitis.

Topics: Acute Disease; Aging; Animals; Animals, Newborn; Ceruletide; Fetus; Gene Expression Regulation, Developmental; Gene Expression Regulation, Enzymologic; Glyceraldehyde-3-Phosphate Dehydrogenases; Liver; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Regeneration; RNA, Messenger; Time Factors; Transcription, Genetic

Regeneration from cerulein-induced pancreatitis is accompanied by a transient synthesis and deposition of extracellular matrix components in the rat pancreas. The pleiotropic transforming growth factor (TGF)-beta1 has been suggested to regulate extracellular matrix remodeling during regeneration from acute pancreatitis. The present study was designed to verify this hypothesis by investigating the effect of TGF-beta1 inhibition.. Experimental acute pancreatitis was induced in rats by supramaximal doses of cerulein. The biological activity of TGF-beta1 was inhibited by injections of neutralizing TGF-beta1 antibody. Changes in the content of extracellular matrix proteins, TGFs, and their messenger RNA concentrations were monitored.. TGF-beta1 expression in pancreatic cells was suppressed after induction of acute pancreatitis by the application of neutralizing TGF-beta1 antibody. Immunochemical analysis showed a clear reduction of extracellular matrix formation during the regeneration of the pancreas in antibody-treated animals. The hydroxyproline content and the concentration of collagen types I and III, fibronectin on protein, and messenger RNA level were significantly reduced in the pancreas of treated animals.. These results provide evidence that TGF-beta1 is involved in the regulation of extracellular matrix remodeling in the rat pancreas during regeneration from acute pancreatitis.

Topics: Acute Disease; Animals; Blotting, Western; Ceruletide; Collagen; Extracellular Matrix; Extracellular Matrix Proteins; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Regeneration; RNA, Messenger; Transforming Growth Factor beta

The present studies were performed to evaluate pancreatic exocrine function in rats during the early stage of acute pancreatitis in two models: one is edematous pancreatitis induced by four subcutaneous injections of 20 micrograms/kg body weight of cerulein at hourly intervals and the other is hemorrhagic pancreatitis induced by retrograde infusion of 0.4 ml/kg body weight of 3% sodium taurocholate (NaTc) into the pancreatic duct. Secretory studies were performed in vivo under urethane anesthesia at various times after induction of acute pancreatitis. Basal pancreatic fluid secretion was significantly elevated after induction of acute pancreatitis in both the cerulein and the NaTc models, reaching the peak level on postpancreatitic days 1 and 3, respectively. In both models of rats, a stepwise increasing dose of cerulein was unable to cause a further increase in fluid secretion above the basal level, whereas it caused a dose-dependent increase in protein output in both models, although the responsiveness and the sensitivity were markedly reduced compared with the controls. In contrast to cerulein, secretin caused a dose-dependent increase in fluid secretion in both models of pancreatitis. In cerulein-induced postpancreatitic rats, secretin also caused a dose-dependent increase in protein output and bicarbonate concentration, but it had only a small effect at certain doses in NaTc-induced postpancreatitic rats. These results indicate that basal pancreatic fluid secretion was greatly increased but the secretory response to cerulein stimulation was reduced in acute pancreatitis early after the onset but was not reduced to secretin stimulation and that protein output and bicarbonate concentration were reduced depending on the severity of pancreatitis (NaTc-pancreatitis > cerulein-pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cholagogues and Choleretics; Gastrointestinal Agents; Male; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Secretin; Taurocholic Acid

Oxidative stress and the inflammatory response may play roles in the pathogenesis of acute pancreatitis. Herein, we characterized pancreatic expression of oxidative stress-responsive genes [c-fos, heme oxygenase-1 (HO-1), and metallothionein-I (MT-I)] and cytokine genes [interleukin-1 beta (IL-1 beta), IL-6, and tumor necrosis factor-alpha (TNF-alpha)] during caerulein-induced acute pancreatitis in the mouse. c-fos, HO-1, and MT-I mRNAs were coordinately and rapidly (3-7 h) upregulated, and HO-1 and MT-I protein levels were increased slightly in the pancreas during acute pancreatitis. In addition, IL-1 beta, IL-6, and TNF-alpha mRNAs were rapidly (7 h) upregulated in the pancreas, and intrapancreatic IL-1 beta and IL-6 protein levels rapidly increased (3-fold and 6.4-fold, respectively) during acute pancreatitis. These studies suggest that oxidative stress and inflammation each occur in the pancreas during the early stages of acute pancreatitis. However, under a limited set of experimental conditions, we found that an insult that causes pancreatic oxidative stress (diethylmaleate) or one that induces an inflammatory response (bacterial lipopolysaccharide), or a combination of these agents, did not cause the changes characteristic of acute pancreatitis. Therefore, simply inducing oxidative stress and/or inflammation may be insufficient to initiate acute pancreatitis.

Topics: Acute Disease; alpha-Amylases; Animals; Ceruletide; Cloning, Molecular; Cytokines; Escherichia coli; Genes, fos; Glutathione; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Interleukin-1; Interleukin-6; Lipopolysaccharides; Male; Membrane Proteins; Metallothionein; Mice; Mice, Inbred Strains; Oxidative Stress; Pancreas; Pancreatitis; Proto-Oncogene Proteins c-fos; RNA, Messenger; Transcription, Genetic; Tumor Necrosis Factor-alpha

The initial pathophysiological events that characterize acute pancreatitis include the formation of pancreatic edema. An interstitial accumulation of fluid, however, is incompatible with the presence of intact intercellular junctions between acinar cells. This study examined the major components of adherens junctions, E-cadherin, alpha-catenin, beta-catenin, and actin, during the initial phase of experimental pancreatitis.. Pancreatitis was induced in rats by 10 micrograms.kg-1.h-1 intravenous cerulein for up to 12 hours. Adherens junction proteins were localized by immunocytochemistry for fluorescence microscopy or electron microscopy, their expression was studied by slot blot analysis, and their association was investigated by immunoprecipitation and Western blot.. During a rapid increase of E-cadherin-encoding RNA, E-cadherin protein declined only moderately and, unlike its cytoskeletal binding partner actin, was not proteolytically cleaved during pancreatitis. Morphologically, E-cadherin and beta-catenin were localized at the basolateral cell membrane from where they rapidly dissociated early in pancreatitis and to where they slowly relocalized during the subsequent course. E-cadherin/beta-catenin complexes disintegrated and reassembled completely in parallel on immunoprecipitation experiments.. The dissociation of adherens junctions and the internalization, relocalization, and reassembly of their major components seem to represent the critical biochemical event at cell-cell contacts during edema formation and resolution in acute pancreatitis.

Topics: Actins; Acute Disease; alpha Catenin; Animals; beta Catenin; Cadherins; Ceruletide; Cytoskeletal Proteins; Intercellular Junctions; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Trans-Activators

Bacterial infectious complications are the most common cause of morbidity and mortality associated with acute pancreatitis. Most pathogens are common gastrointestinal flora, indicating that the gut is the source of pancreatitis-related infections. However, the route whereby the microorganisms reach distant organs remains speculative. We tested the hypothesis that spread of bacteria occurs via a transperitoneal pathway. Acute interstitial pancreatitis (AIP) was induced in antibiotic (gentamicin, bacithracin, neomycin)-decontaminated rats by intravenous infusion of cerulein. Effects of pancreatic necrosis (PN) were studied in rats that received additional injections into the peritoneal cavity of pancreatic tissue obtained from donor rats. The rats were inoculated with Escherichia coli (O2:KN:H18) resistant to the antibiotics used for decontamination either orally (10(12) microorganisms; experiment I) or intraperitoneally (10(8) microorganisms; experiment II). Moreover, the rat peritoneal cavity wash was inoculated with 10(8) E. coli in vitro (experiment III). In rats with AIP and PN, recovery of the bacteria from liver, spleen, pancreas, lung, and blood following oral inoculation demonstrated that acute pancreatitis promotes bacterial translocation from the gut. The absence of E. coli in these organs following intraperitoneal inoculation showed that the bacteria do not spread from the peritoneal cavity. Rats with PN cleared E. coli from the peritoneal cavity in a shorter period than rats with AIP and controls (5 vs. 7 and 8 days; p < 0.05). The multiplication rate of E. coli in peritoneal cavity wash was lower in rats with PN than in rats with AIP and controls (p < 0.01). We conclude that (1) translocation of E. coli from the gut during cerulein-induced acute pancreatitis occurs via nonperitoneal pathways, (2) the peritoneal cavity acts as a trap for the bacteria rather than a source of bacterial seeding, and (3) PN impairs survival of E. coli in the peritoneal cavity via inhibition of the bacterial multiplication in this model.

Topics: Acute Disease; Animals; Ascites; Bacterial Translocation; Ceruletide; Escherichia coli; Male; Pancreatitis; Peritoneal Cavity; Rats; Rats, Sprague-Dawley

The capacity for intercellular communication (IC) via gap junctions is found in normal pancreatic acinar cells. The major role of IC is considered to be the maintenance of tissue homeostasis and the regulation of signal transmissions. Up to now, the participation of IC via gap junctions in acute pancreatitis has not been reported. We investigated the role of IC in cerulein (Cn)-induced acute pancreatitis in rats using irsogladine, an enhancer of IC via gap junction. Acute edematous pancreatitis was induced in rats by two intraperitoneal injections of 40 micrograms/kg Cn. Rats received various doses (25, 50, or 100 mg/kg body weight) of irsogladine orally, 15 and 2 h before the first Cn injection. The normal control group received only vehicle. The severity of pancreatitis was evaluated enzymatically and histologically 5 h after the first Cn injection. In Cn-induced acute pancreatitis, irsogladine significantly lowered the serum amylase level, the pancreatic wet weight, and the pancreatic amylase and DNA contents, in a dose-dependent manner. Particularly, the amylase content improved to the level of the normal controls. Histologically, the severity of pancreatitis was reduced significantly by treatment with irsogladine and no discernible vacuolization was seen in the group with 100 mg/kg irsogladine treatment. By immunofluorostaining pancreata with anti-connexin 32 (Cx32; a gap junction protein) antibody, we found that pancreatic acini were diffusely positive for Cx32 in the control group, but the number of Cx32-positive grains decreased markedly, to 19%, in the pancreatitis group. With 100 mg/kg irsogladine treatment, the number of Cx32 grains recovered to 70% of the normal control value. These findings indicate that IC via gap junction is disturbed in Cn-induced pancreatitis, which may result in the breakdown of tissue homeostasis and the progression of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Connexins; DNA; Fluorescent Antibody Technique; Gap Junction beta-1 Protein; Gap Junctions; Male; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Triazines

The behavior of neutrophils in a rat acute pancreatitis model was observed in the pancreas and liver using fluorescence microscopy with an image analyzing system after labeling with a specific fluorescent reagent. Nonviable cells of both organs were also labeled and quantified. The role of nitric oxide in neutrophil accumulation and organ damage was estimated by administering a relatively selective inhibitor of constitutive nitric oxide synthase, N-nitro-L-arginine (L-NNA). The animal model of acute pancreatitis was induced by cerulein injection (80 mg/kg). Two groups were created, one given and the other not given L-NNA (2.5 mg/kg) prior to the induction of pancreatitis. The number of accumulated neutrophils in the pancreas and liver increased in a time-dependent manner. There was a close relation between the distribution of the neutrophils and inviable acinar cells or hepatocytes. When pretreated with L-NNA, the numbers of accumulated neutrophils and nonviable cells increased significantly in the pancreas. In the liver, a more pronounced accumulation of neutrophils was observed after treatment with L-NNA. Although hepatocyte injury was mild despite the neutrophil accumulation in the control, such injury was marked in the group treated with L-NNA. This suggests that neutrophils serve an important role in exacerbating acute pancreatitis and that nitric oxide provides a defense mechanism against neutrophil accumulation in pancreas and liver.

Topics: Acute Disease; Animals; Capillaries; Cell Adhesion; Ceruletide; Enzyme Inhibitors; Liver; Male; Neutrophils; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pancreas; Pancreatitis; Rats; Rats, Wistar; Venules

Local microcirculatory dysfunction within the pancreatic gland might be an important factor in the conversion of oedematous to necrotizing pancreatitis. Therapeutic agents, improving the pancreatic blood flow, might be valuable in acute pancreatitis treatment. An influence of nitric oxide, heparin and procaine treatment on microcirculatory values in acute pancreatitis (AP) in rats was investigated. Acute pancreatitis was induced by i.p. injection of cerulein in four doses of 15 microg kg-1 each at 1-h intervals. The rats with pancreatitis were divided into five groups, 12 animals each. One group remained without treatment, four groups were treated i.p. either with NO synthase inhibitor L-NNA (2x25 mg kg-1 or heparin 2x2.5 mg kg-1 or L-arginine 2x100 mg kg-1 or procaine 2x25 mg kg-1. Five control groups, ten animals each, received saline, L-NNA, heparin, L-arginine or procaine only. Five hours after the first ceruleine injection microcirculatory values within the pancreas were measured by means of laser Doppler flowmetry. Acute pancreatitis caused a significant drop of microcirculatory value to 37% of the basal value. The L-NNA administration resulted in a further insignificant reduction of the pancreatic blood flow to 34%. An improvement of microcirculation was observed in rats with pancreatitis receiving heparin (76%) and L-arginine (72%). Procaine had no effect on microcirculatory disturbances within the pancreas in rats with pancreatitis. Cn-induced acute pancreatitis (AP) causes microcirculatory deterioration within the pancreas. Heparin and nitric oxide donor, L-arginine, might be considered as therapeutic agents, improving the diminished pancreatic tissue perfusion observed in acute pancreatitis. Procaine does not improve the pancreatic blood flow in acute pancreatitis.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Heparin; Male; Microcirculation; Nitroarginine; Pancreas; Pancreatitis; Procaine; Rats; Rats, Wistar

We have recently reported that preconditioning through hyperthermia induces expression of pancreatic heat shock proteins (HSPs) and protects against caerulein pancreatitis. Here, we investigate caerulein-mediated effects on pancreatic HSPs without prior hyperthermia. Caerulein time and dose dependently increased pancreatic mRNA levels of the constitutive isoform of HSP70 (HSC70). However, pancreatic HSC70 protein levels were decreased, as were HSP60 protein levels. Also, in contrast to hyperthermia preconditioning, caerulein did not induce measurable levels of mRNA or protein of the inducible isoform of HSP70. Thus the pancreas reacts to different kinds of stress (hyperthermia vs. hyperstimulation) with differential induction of HSP mRNAs. Clearly, hyperthermia leads to induction of HSP protein expression, whereas caerulein treatment does not. Therefore, our current study further supports the idea that hyperthermia-induced protection against caerulein pancreatitis may be mediated through increased protein levels of pancreatic HSPs. It is further tempting to hypothesize that failure to appropriately increase HSP protein levels in response to high doses of caerulein might be a factor in the development of pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Chaperonin 60; Gene Expression Regulation; Heat-Shock Proteins; HSP70 Heat-Shock Proteins; Pancreas; Pancreatitis; Protein Biosynthesis; Rats; Reference Values; RNA, Messenger; Time Factors; Transcription, Genetic

Acute pancreatitis associated with hypercalcaemia has been described in humans and experimental animals. It has been demonstrated that calcium dose dependently accelerates trypsinogen activation, and it is generally believed that ectopic activation of digestive enzymes is an early event in the pathophysiology of acute pancreatitis.. Trypsinogen activation peptide (TAP) was measured in isolated rat pancreatic acini exposed to elevated extracellular calcium in order to investigate the association between calcium and trypsinogen activation in living cells. TAP was determined in the culture medium either before (extracellular compartment) or after (intracellular compartment) cell homogenisation.. Neither secretory stimulation nor elevated calcium alone caused an increase in TAP levels. Maximal cerulein or carbachol stimulation superimposed on high medium calcium, however, significantly increased intracellular trypsinogen activation twofold. This increase was inhibited by either NG-monomethyl-L-arginine (L-NMMA) or verapamil. Acinar cell morphology and function remained intact as demonstrated by electron microscopy and secretagogue dose-response studies.. These results support the hypothesis that increased intracellular trypsinogen activation is an early step in the pathogenesis of hypercalcaemia induced pancreatitis. The model may have a bearing on other types of pancreatitis as elevated cytosolic calcium is thought to be an early event in the pathogenesis of acute pancreatitis in general.

Topics: Acute Disease; Animals; Calcium; Carbachol; Ceruletide; Culture Techniques; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Gastrointestinal Agents; Male; Oligopeptides; Pancreas; Pancreatitis; Rats; Rats, Wistar; Trypsinogen

We investigated the role of neutrophils and the involvement of apoptosis in cerulein-induced acute pancreatitis. Male Sprague-Dawley rats were divided into 2 groups. In the control group, acute pancreatitis was induced by subcutaneous injections of cerulein. In methotrexate-treated group, the rats received intraperitoneal injections of methotrexate to produce neutrophil depletion before the injections of cerulein. The rats were sacrificed at the indicated time points until 72 h after the first injection of cerulein. Neutrophil depletion ameliorated pancreatic edema and vacuole formation in acinar cells during the early stages of cerulein-induced acute pancreatitis. Electron microscopy, DNA gel electrophoresis and in situ nick end-labeling revealed the involvement of apoptosis in acinar cells in cerulein-induced acute pancreatitis. Furthermore, the number of apoptotic acinar cells in neutrophil-depleted rats showed an about 2-fold increase during the late stages when compared with those in the control rats. Our results suggest that neutrophil depletion in cerulein-induced pancreatitis leads to amelioration of pancreatic injury during the early stage, and enhancement of apoptosis by neutrophil depletion occurs during the late stage.

Topics: Acute Disease; Amylases; Animals; Apoptosis; Ceruletide; Lipase; Male; Methotrexate; Microscopy, Electron; Neutropenia; Neutrophils; Pancreas; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley

Interleukin 1beta (IL-1beta) is produced in large amounts during acute pancreatitis and is believed to play a primary role in determining pancreatitis severity and the degree of pancreatic tissue destruction. This study was undertaken to characterize intrapancreatic production of IL-1beta and the remainder of the IL-1 family of genes during sterile acute pancreatitis. Moderate or severe necrotizing pancreatitis was induced by the intraperitoneal injection of a cholecystokinin analogue or the feeding of a choline deficient diet, respectively. Animals were killed during the progression of pancreatitis with severity scored by histological grading and serum amylase concentration. The expression of IL-1beta, IL-1 Receptor 1 (IL-1R1), Il-1R2, IL-1R antagonist (IL-1Ra), and ICE mRNA within the pancreas was examined by quantitative differential RT-PCR. Corresponding intrapancreatic and serum proteins were measured by enzyme-linked immunosorbent assay (ELISA). There was constitutive expression of pancreatic IL-1R1, IL-1R2, IL-1Ra, and ICE but not IL-1beta. As pancreatitis developed, mRNA for IL-1beta, IL-1Ra, and ICE increased in parallel with the degree of pancreatitis severity (all P<0.001 vs baseline) while mRNA for both receptors remained stable (P=NS). Intrapancreatic and systemic IL-1beta and IL-1Ra protein also increased as pancreatitis developed (both P<0.001) with tissue levels being continuously greater than serum. This study demonstrated that sterile, endotoxin-free acute pancreatitis induces the upregulation of specific members of the IL-1 family of genes including production of large amounts of IL-1beta and its receptor antagonist within the pancreatic parenchyma. These changes are indicative of pancreatitis severity and are not model dependent.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; DNA Primers; Interleukin-1; Male; Mice; Multigene Family; Pancreas; Pancreatitis; Polymerase Chain Reaction; Receptors, Interleukin-1; RNA, Messenger; Transcription, Genetic

This study aimed to investigate the protein peroxidation process in cerulein induced acute pancreatitis. Eighteen rats were divided into three equal groups: group 1 acted as control rats had intraperitoneal injection of 0.9% NaCl, in group 2 and 3 rats had injection of cerulein 40 micrograms kg-1 for 3 or 6h of induction period respectively. Protein carbonyls which reflect peroxidative damage were found to be increased after 3h up to 2.53 +/- 0.49 comparing to 1.05 +/- 0.17 in control group and returned to control level 0.95 +/- 0.04 after 6h. These data suggest that during acute pancreatitis free radicals may play an essential role in protein damage. Decrease in protein carbonyls content after 6h suggests an elevated proteolysis of oxidatively damaged proteins.

Topics: Acute Disease; Animals; Ceruletide; Free Radicals; Oxidative Stress; Pancreatitis; Proteins; Rats; Rats, Wistar

The object of the present work was to study the relationship between acute pancreatitis (PA) and hyperlipidic diets. PA was induced by Caerulein (CE) by a single intraperitoneal doses (50 mcg/kg), after feeding the rats during 6 weeks with an hyperlipidic diet (45%). Rats with a normolipidic diet (lipids 5%) were used as control. The increase of serum lipase was similar in both groups treated with CE (control and with hyperlipidic diet). There were increase of interstitial edema, cariorrexis and a specially marked increase in the level of vacuolization of acinar cells with respect to the control group. It was concluded that chronic hyperlipidic diet increases histopathologic lesions in PA induced by CE in rats.

Topics: Acute Disease; Analysis of Variance; Animals; Ceruletide; Dietary Fats; Esterases; Lipid Metabolism; Male; Pancreatitis; Rats; Rats, Wistar

Transforming growth factor beta (TGF-beta) is a putative mediator of fibrosis in several chronic diseases. Recently, chronic pancreatitis was suggested to be related to acute pancreatitis in the so-called necrosis-fibrosis sequence hypothesis. The present study investigated whether TGF-beta is able to promote chronic fibrosis after repeated courses of necrotizing acute pancreatitis induced by cerulein in mice.. Six episodes of acute pancreatitis were repeatedly induced at weekly intervals in mice receiving either recombinant TGF-beta (4 micrograms in 4 days) or excipient alone at each induction. One week after the last induction, pancreatic lesions and collagen deposition were histologically assessed. Expression of pancreatic fibronectin messenger RNA was also examined in both groups.. TGF-beta had no influence on a single course of acute pancreatitis. After six courses of acute pancreatitis, only mild inflammatory changes were observed in the control group. In contrast, important areas of perilobular and intralobular fibrosis were observed adjacent to inflammatory and necrotic foci in the TGF-beta group. Fibronectin messenger RNA expression was significantly higher in this group.. TGF-beta promotes development of pancreatic fibrosis after recurrent episodes of acute pancreatitis. This model of pancreatic fibrosis could be used as a model of chronic pancreatitis consistent with the necrosis-fibrosis sequence hypothesis.

Topics: Acute Disease; Animals; Base Sequence; Ceruletide; Collagen; Disease Models, Animal; Female; Fibronectins; Fibrosis; Mice; Mice, Inbred Strains; Molecular Sequence Data; Necrosis; Pancreas; Pancreatitis; Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor beta

Although several pathophysiological sequences, such as protease activation, free radical generation, and inflammatory mediator release, have been described in acute pancreatitis, the precise mechanism by which acute pancreatitis is initiated is unknown. Cellular calcium, a key physiological signaling element in cell function and also a crucial pathological intracellular messenger in cell injury, appears to be involved in the initiation and development of acute pancreatitis. The present study provides several lines of evidence supporting this suggestion. First, verapamil (a calcium channel blocker) administration was associated with a significant protection of rats from acute pancreatitis induced by high doses of cerulein (50 micrograms/kg/hr, subcutaneously), as evidenced both histologically and biochemically. Second, verapamil was found to minimize the increased tissue levels of calcium, platelet-activating factor, and thromboxane B2 detected during acute pancreatitis. Third, acute pancreatitis could be observed in rats with elevated serum calcium levels at low doses of cerulein (5 micrograms/kg/hr, subcutaneously), but could not be observed in rats with normal serum calcium levels treated with low doses of cerulein. It is proposed that cellular calcium, which is a critical signaling component in the synthesis and release of inflammatory mediators and several other events, may be an important factor in the pathogenesis of cerulein-induced acute pancreatitis.

Topics: Acute Disease; Animals; Calcium; Calcium Channel Blockers; Ceruletide; Drug Synergism; Male; Pancreas; Pancreatitis; Platelet Activating Factor; Rats; Rats, Sprague-Dawley; Thromboxane B2; Verapamil

The authors' aim was to determine the requirement for an active interleukin (IL)-1 receptor during the development and progression of acute pancreatitis.. Interleukin-1 is a pro- inflammatory cytokine that has been shown to be produced during acute pancreatitis. Earlier animal studies of moderate and severe pancreatitis have shown that blockade of this powerful mediator is associated with attenuated pancreatic destruction and dramatic increases in survival. The exact role played by IL-1 and the requirement for activation of its receptor in the initiation and progression of pancreatitis is unknown.. Conventional and IL-1 receptor "knockout" animals were used in parallel experiments of acute pancreatitis induced by intraperitoneal injection of cerulean (50 microg/kg every 1 hour X 4). The conventional mouse strain had the IL-1 receptor blocked prophylactically by means of a recombinant IL-1 receptor antagonist (10 mg/kg injected intraperitoneally every 2 hours). The second mouse strain was genetically engineered by means of gene targeting in murine embryonic stem cells to be devoid of type 1 IL-1 receptor (IL-1 receptor knockout). Animals were killed at 0, 0.5, 1, 2, 4, and 8 hours, with the severity of pancreatitis determined by serum amylase, lipase, and IL-6 levels and blind histologic grading. Strain-specific controls were used for comparison.. The genetic absence of the IL-1 receptor or its pharmacologic blockade resulted in significantly attenuated pancreatic vacuolization, edema, necrosis, inflammation, and enzyme release. Serum IL-6, a marker of inflammation severity, was dramatically decreased in both groups.. Activation of the IL-1 receptor is not required for the development of pancreatitis but apparently is necessary for the maximal propagation of pancreatic injury and its associated inflammation.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Disease Progression; Interleukin 1 Receptor Antagonist Protein; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Pancreatitis; Receptors, Interleukin-1; Recombinant Proteins; Sialoglycoproteins

The recent observation that the severity of pancreatitis is inversely related to the extent of apoptosis in five experimental models of the disease has suggested the possibility that apoptosis might protect against pancreatic injury in pancreatitis. This hypothesis was tested by inducing pancreatitis in mice during a phase of extensive apoptosis. Mice were fed a raw soya diet for five weeks to stimulate pancreatic growth and then switched to a regular chow diet for 27 hrs to permit involution of the hypertrophied gland. That involution is characterized by extensive apoptosis of acinar cells. Pancreatitis was induced, in either control mice or mice undergoing pancreatic involution, by repeated intraperitoneal administration of a supramaximally stimulating dose of caerulein (50 microg/kg given each hr for 12 hrs). The magnitude of hyperamylasemia, degree of inflammation, and extent of necrosis were reduced in the mice receiving caerulein during pancreatic involution. We conclude that induction of apoptosis may protect against acinar cell injury and reduce the severity of pancreatitis.

Topics: Acute Disease; Amylases; Analysis of Variance; Animal Feed; Animals; Apoptosis; Biomarkers; Ceruletide; Female; Glycine max; Hypertrophy; Mice; Mice, Inbred Strains; Necrosis; Pancreatitis

Somatostatin and its analogue octreotide have a profound inhibitory effect on the endocrine and exocrine secretions of the pancreas, stomach, and small intestine. Previous studies have been inconclusive about the possible therapeutic effect of somatostatin and its analogues in the treatment of pancreatitis. This study assessed the effect of the long acting somatostatin analogue, octreotide, in two models of experimental pancreatitis in rats. Necrotizing pancreatitis was induced by pancreatic injection of 5 ml taurocholate, 5% in male Wistar rats. In a second model mild edematous pancreatitis was induced by intravenous injection of caerulein at a supramaximal dose, 6 micrograms/kg/hr, for 5 hr. Compared to untreated rats, treatment with octreotide either prior to or following the induction of necrotizing pancreatitis resulted in less hypocalcemia (P < 0.05) and acidosis (P < 0.05), and prevented the increase in pancreatic weight (P < 0.05). Amylase levels remained high. After 20 days, there was less pancreatic damage, lower mortality rates (P < 0.05), and increase in body weight (P < 0.05). In the model of milder pancreatitis, octreotide treatment attenuated the increase in pancreatic weight (P < 0.05) and pathological damage (P < 0.05). We concluded that the somatostatin analogue octreotide has a beneficial effect in the treatment of experimental acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Hormones; Hydrogen-Ion Concentration; Male; Octreotide; Organ Size; Pancreatitis; Rats; Rats, Wistar; Taurocholic Acid

Interleukin-1 (IL-1) gene expression is selectively induced in tissues involved in multisystem organ failure during acute pancreatitis, suggesting a role in the pathogenesis of distant organ dysfunction. This study was undertaken to investigate the mechanism of pancreatitis-induced end organ cytokine production and to better understand the processes by which IL-1 production is regulated. Seventy adult male transgenic mice in which the type 1 IL-1 receptor had been deleted by gene targeting in embryonic stem cells were utilized (homozygous -/- IL-1R knockout). Acute pancreatitis was induced by one of two methods: (A) IP injections of caerulein (50 microgram/kg/hr x 4) with animals sacrificed at 0, .5, 1, 2, 4, 6, and 8 hr; (B) 48-hr exposure to a choline deficient ethionine supplemented (CDE) diet with animals sacrificed at 0 and 72 hr. Knockout animals were compared to strain-specific control mice expressing the normal wild-type IL-1 receptor gene in which pancreatitis was similarly induced. The severity of pancreatitis was stratified by serum amylase, lipase, and blind histologic grading. IL-1 mRNA production was determined within the pancreas, lungs, liver, and spleen by quantitative differential RT-PCR. Deletion of the IL-1R1 attenuated the severity of pancreatitis, reaching statistical significance in the less severe edematous model. There was little or no constitutive expression of IL-1 mRNA within any of the tissues examined from wild-type animals; however, knockout animals showed elevated steady-state levels in each tissue. IL-1 mRNA became detectable in all tissues of wild-type animals shortly after either form of pancreatitis became apparent and increased significantly with worsening pancreatitis. Despite the attenuated pancreatitis, knockout animals produced significantly greater levels of IL-1 mRNA in each tissue, typically demonstrating a 30-50% increase over time matched IL-1 mRNA production in wild-type animals which was not pancreatitis model dependent. We conclude that genetic deletion of IL-1 receptors results in the overproduction of IL-1 mRNA in organs known to produce cytokines during pancreatitis even when the severity of pancreatitis is lessened. This suggests that a negative feedback loop exists between the IL-1 receptor and IL-1 gene expression.

Topics: Acute Disease; Animals; Base Sequence; Ceruletide; Choline Deficiency; Cytokines; Diet; DNA Primers; Ethionine; Female; Gene Deletion; Gene Expression; Heterozygote; Homozygote; Interleukin-1; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Molecular Sequence Data; Pancreatitis; Polymerase Chain Reaction; Receptors, Interleukin-1; RNA, Messenger; Stem Cells

The importance of interleukin-1beta (IL-1beta) in the pathogenesis of acute pancreatitis has been demonstrated by dramatic attenuation of pancreatic destruction and significant increases in survival when its actions are inhibited. The pancreas has been shown to be a major producer of IL-1beta during pancreatitis but the cell(s) of origin remains unknown. Hypothesizing that infiltrating leukocytes contribute substantially, the intrapancreatic production of IL-1beta was examined after specific leukocyte populations were manipulated in vivo prior to the induction of pancreatitis. Sixty-four adult male Swiss mice were assigned to one of four groups 48 hr prior to induction of pancreatitis: (1) PMN depletion via anti-murine PMN antiserum. [PMN-d], (2) macrophage (Mphi) depletion via anti-macrophage antiserum [Mphi-d], (3) PMN and Mphi depletion [PMN+Mphi-d], and (4) Immunocompetent Pancreatitis. Edematous pancreatitis was induced in all experimental groups by caerulein (50 microg/kg/hr ip X 4). Animals were sacrificed 6 hr after induction of pancreatitis with severity determined by blind histologic grading and serum amylase, lipase, and interleukin-6 (IL-6) levels. Intrapancreatic IL-1beta production was determined by immunohistochemistry and semiquantitative differential RT-PCR. Pancreatitis developed in all animals receiving caerulein; however, leukocyte-depleted animals showed significantly attenuated levels of serum amylase, lipase, and IL-6, as well as lower histologic severity scores. Similarly, pancreatitis induction in immunocompetent mice showed pancreatic infiltration of IL-1beta-producing cells, whereas the leukocyte-depleted animals had significantly decreased numbers (PMN+Mphi-d < Mphi-d < PMN-d). IL-1beta mRNA was upregulated in all animals developing pancreatitis with significantly lower levels seen in the leukocyte-depleted groups. We conclude that infiltrating leukocytes, both neutrophils and macrophages, are responsible for the majority of intrapancreatic IL-1beta production during acute pancreatitis. The elimination of leukocytes and their products, including IL-1beta, significantly decreases the severity of pancreatic destruction.

Topics: Acute Disease; Amylases; Animals; Base Sequence; Ceruletide; DNA Primers; Edema; Gene Expression; Immunohistochemistry; Interleukin-1; Interleukin-6; Leukocytes; Macrophages; Male; Mice; Molecular Sequence Data; Neutrophils; Pancreas; Pancreatitis; Polymerase Chain Reaction; Reference Values; RNA, Messenger; Time Factors

Disruption of pancreatic exocrine secretion is an important feature of acute pancreatitis. Because cytosolic calcium is a key intracellular messenger controlling pancreatic secretion, this study examined patterns of calcium signaling during the early stages of cerulein-induced pancreatitis.. Mice were administered hourly intraperitoneal injections of cerulein (50 micrograms/kg), and paired controls were administered saline. Acini were isolated by collagenase from pancreatic tissue harvested after injections 1, 3, 5, and 7 and were loaded with Fura-2. Individual cellular calcium responses to acetylcholine and cholecystokinin were studied using digital imaging.. The proportion of cells maintaining a normal oscillatory calcium response to physiological secretagogue stimulation diminished progressively after increasing cerulein injections. Also, the normal polarized spatial pattern of calcium Increase within individual acinar cells was progressively lost. A sustained response to high-dose stimulation was maintained but with diminishing amplitude. The characteristic calcium response to the Ca(2+)-adenosine triphosphatase inhibitor thapsigargin was maintained, implying that calcium reuptake and extrusion were not impaired.. Progressive disruption of physiological patterns of pancreatic acinar cell calcium signaling, notably in the secretory pole of the cell, is an early feature of pancreatitis induced by cerulein hyperstimulation. These changes may be important in contributing to the disruption of exocrine secretion in acute pancreatitis.

Topics: Acetylcholine; Acute Disease; Animals; Calcium; Calcium-Transporting ATPases; Ceruletide; Cholecystokinin; Endoplasmic Reticulum; Enzyme Inhibitors; Male; Mice; Mice, Inbred Strains; Pancreas; Pancreatitis; Signal Transduction; Terpenes; Thapsigargin

We investigated the protective and/or therapeutic effects of a new cholecystokinin receptor antagonist, KSG-504, on different types of experimental pancreatitis in the rat and mouse. The intravenous injection of KSG-504 (10, 25, 50 and 100 mg/kg) before caerulein administration to the rat inhibited the increases in plasma amylase, lipase and of pancreatic wet weight in a dose-dependent manner. The histological changes due to caerulein-induced acute pancreatitis were also decreased by KSG-504 when KSG-504 (25, 50 and 100 mg/kg) was administered after the induction of acute pancreatitis; the increases in plasma amylase, lipase and pancreatic wet weight were reduced, but the histological changes of the pancreas were not decreased significantly. In the second experiment, acute pancreatitis was induced in rats by injecting 0.3 ml of 6% sodium taurocholate into the pancreatic interstitial tissue. KSG-504 administered immediately and 1.5 hr after sodium-taurocholate injection at 100 mg/kg reduced the increases of pancreatic enzymes in the plasma, pancreatic wet weight and ascites. Moreover, KSG-504 (50 and 100 mg/kg, i.v., x 2) mitigated the histological changes of taurocholate-induced acute pancreatitis. Another type of acute pancreatitis was induced in mice by dl-ethionine (0.5 g/kg, p.o., x 4) and a choline-deficient diet. KSG-504 (10, 30 and 100 mg/kg) was subcutaneously administered five times every 12 hr during the experiment. KSG-504 elongated the survival of mice in a dose-dependent manner. These findings suggest that KSG-504 has potent protective and/or therapeutic effects against acute pancreatitis and that cholecystokinin may be involved in the development of pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Lipase; Male; Mice; Mice, Inbred ICR; Naphthalenes; Pancreas; Pancreatitis; Pentanoic Acids; Rats; Rats, Wistar; Receptor, Cholecystokinin A; Receptors, Cholecystokinin

Cerulein-induced pancreatitis in rats associated with remote liver and lung dysfunction. Soluble complement receptor 1 (sCR1) does not reduce the local or remote injury. Thus complement activation does not moderate cerulein-induced pancreatitis or the associated liver and injury.. The local and remote injury of pancreatitis resembles other inflammatory events that are mediated by complement. This study examines the effect of complement inhibition with sCR1 in cerulein-induced pancreatitis in rats.. Thirteen Sprague-Dawley rats received five hourly subcutaneous doses of cerulein (100 micrograms initially, then 50 micrograms/kg). Six of these animals received hourly i.v. sCR1 (15 mg/kg, a proven complement-inhibiting dose in rats) and the other seven received i.v. saline. In parallel, 12 additional rats received hourly s.c. and i.v. saline.. Compared to saline controls, rats receiving cerulein showed increased pancreatic wet-to-dry ratio (3.25:8.52), hematocrit (40 to 47%), ascites volume (2.1 to 6.1 mL), serum amylase (1680 to 10,700 U/L), and ascites amylase (32,200 to 167,000 U/L) (all p < 0.05). None of these parameters were modified by treatment with sCR1. Serum SGPT, which increased from 33.4 to 46.6 U/L in cerulein-infused rats, showed a trend toward reduction to 38.8 U/L in rats treated with sCR1. Cerulein-treated rats also had increased lung myeloperoxidase (0.069 to 0.097 U/g) and lung permeability, as assessed by a alveolar lavage to serum ratio of labeled albumen (0.041:0.121) both p < 0.05). Neither were changed by sCR1 treatment.

Topics: Acute Disease; Alanine Transaminase; Amylases; Analysis of Variance; Animals; Biomarkers; Ceruletide; Complement Activation; Complement Inactivator Proteins; Gastrointestinal Agents; Liver; Lung; Male; Organ Size; Pancreatitis; Peroxidase; Rats; Rats, Sprague-Dawley; Receptors, Complement

One of the vasoactive peptides that has been implicated in the progression from edematous to necrotizing pancreatitis is bradykinin. We have investigated the effect of bradykinin administration and bradykinin inhibition on an edematous model of acute pancreatitis in rats (10 micrograms/kg/h of caerulein i.v.). Within six hours i.v. bradykinin reduced circulating serum amylase levels significantly but neither affected tissue edema nor morphology. A bradykinin antagonist (HOE-140), on the other hand, reduced pancreatic edema by 70% and converted edematous pancreatitis into a hemorrhagic and necrotizing variety of the disease. In further experiments we determined the time course and the minimal dosage required for the induction of this severe and non-invasive disease variety. A single dose of caerulein (40 micrograms/kg i.p.) together with a single administration of the bradykinin antagonist HOE-140 (100 micrograms/kg s.c.) consistently resulted in hemorrhagic necrosis of the pancreas within six hours. We conclude that this simple protocol allows for the non-invasive induction of a vascular model of necrotizing pancreatitis and appears ideally suited to study the development of this severe form of the disease.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Bradykinin; Ceruletide; Disease Models, Animal; Edema; Hemorrhage; Male; Necrosis; Pancreatitis; Rats; Rats, Wistar

Exocrine function was studied in anesthetized rats that had received two specific doses of caerulein (maximal stimulation and supramaximal stimulation). Male Wistar rats (body weight, 200-250 g) were divided into three groups: the control group (4-h saline infusion), the maximal stimulation group (0.25 microgram/kg per h caerulein for 4 h), and the caerulein pancreatitis group (10 micrograms/kg per h for 4h). Histologically, interstitial edema and cytoplasmic vacuolization were observed only in the caerulein pancreatitis group, with no abnormal findings in the other groups. The volume of pancreatic juice was significantly increased in both the maximal stimulation group and the caerulein pancreatitis group. The protein output and the amylase output in the 1st h of caerulein infusion were also significantly increased, to 459% and 338% in the maximal stimulation group, and to 925% and 1430% respectively, in the caerulein pancreatitis compared to the baseline values. We also found that the pancreatic juice of the caerulein pancreatitis group contained precipitated protein, and high trypsin activity, and protein degradation was confirmed by electrophoresis. These findings were not observed in the other groups. These results strongly suggest that hypersecretion and the appearance of trypsin activity in pancreatic juice plays an important role in the induction of histological changes in this pancreatitis model in anesthetized rats.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Male; Pancreas; Pancreatic Juice; Pancreatitis; Rats; Rats, Wistar; Trypsin

Recent findings have suggested that oxygen-derived free radicals play an important role in the development and progression of acute pancreatitis. Therefore, the present study was designed to investigate whether metallothionein, a free radical scavenger, can protect against acute pancreatitis. Rats were injected intraperitoneally with zinc, followed by either an infusion of cerulein at 10 micrograms/kg for 4 h or a retrograde injection with 100 microliters/100 g body weight of 5% sodium taurocholate into the pancreaticobiliary duct, in order to induce acute pancreatitis. Zn administration significantly increased the levels of both metallothionein and reduced glutathione in the pancreas; the metallothionein levels reached a peak of 83-fold of normal levels after 24 h. The indications of acute pancreatitis, as well as the mortality, were improved by Zn treatment before the onset of acute pancreatitis. Immunohistochemical studies showed that metallothionein accumulated in the acini of the pancreas in the Zn-treated groups, and with strong staining around the periphery of the vacuoles in the group treated with both Zn and cerulein. These findings suggested that Zn increased both metallothionein and glutathione levels in the pancreas and exerted a beneficial effect against ceruleinor taurocholate-induced acute pancreatitis in rats.

Topics: Acute Disease; Animals; Ceruletide; Free Radical Scavengers; Free Radicals; Glutathione; Immunohistochemistry; Male; Metallothionein; Pancreas; Pancreatitis; Rats; Rats, Wistar; Taurocholic Acid; Thiobarbituric Acid Reactive Substances; Zinc

The role of the hypothalamic-pituitary-adrenal axis (HPA axis) in acute pancreatitis has not yet been clarified. In the present study, the concentrations of serum corticosterone and amylase, the severity of pancreatic edema, and the histology of the pancreas during cerulein-induced pancreatitis were compared in two strains of rats whose HPA axes have been reported to be hyperresponsive (Fischer female) and hyporesponsive (Lewis female) to inflammatory mediators. First, we confirmed that the secretory response of corticosterone to lipopolysaccharide was remarkably blunted in Lewis rats compared with Fischer rats. With a single intraperitoneal injection of cerulein at a dose of 50 micrograms/kg, the serum corticosterone of Fischer rats increased promptly, and their serum levels were significantly higher than those of Lewis rats at all points after the induction of pancreatitis. The edema formation and infiltration of inflammatory cells into the pancreas were more severe in Lewis rats than in Fischer rats. The serum amylase concentration was not significantly different between the two strains, except at 2 h after the induction of pancreatitis. The in vitro study using dispersed pancreatic acini showed that there was no significant difference in cerulein-stimulated amylase secretion between the two strains. These findings suggest that the responsiveness of the HPA axis and the consequent secretion of glucocorticoids might modify the pathological features of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Corticosterone; Female; Hypothalamo-Hypophyseal System; Lipopolysaccharides; Pancreatitis; Pituitary-Adrenal System; Rats; Rats, Inbred F344; Rats, Inbred Lew; Secretory Rate

Pancreatitis is characterized by inflammation and death of acinar cells. Death can occur by either necrosis or apoptosis. The initial injury may cause expression of cytokines that mediate activation and infiltration of neutrophils. The aim of this study was to assess the effect of neutrophils and platelet-activating factor (PAF) in cell death responses.. The effects of neutrophil depletion with antineutrophil serum (ANS) and a PAF antagonist (BN52021) were measured in the cerulein model of pancreatitis. Rats received a 6-hour intravenous infusion of cerulein either alone or after treatment with ANS, BN52021, or both.. Cerulein-induced pancreatitis was characterized by neutrophilic infiltration, vacuolization of acinar cells, and foci of necrosis. Treatment with ANS and BN52021 prevented the inflammatory response caused by cerulein and decreased the cell damage. Treatment with ANS increased apoptosis in cerulein-infused animals. When BN52021 was added, apoptosis was abolished. The measurement of PAF in pancreatic tissue showed a ninefold increase with cerulein treatment alone and a 14-fold increase in cerulein-infused, neutrophil-depleted animals.. The results indicate that cerulein stimulates pancreatic production of PAF. PAF mediates both apoptosis and neutrophil chemotaxis in the pancreas. Neutrophils in turn may convert acinar cells undergoing apoptosis into necrotic cells.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Diterpenes; Ginkgolides; Lactones; Lipase; Male; Neutrophils; Pancreatitis; Platelet Activating Factor; Rats; Rats, Sprague-Dawley

1. We have measured intracellular calcium concentrations in basal conditions and in response to cholecystokinin-octapeptide and acetylcholine in pancreatic acini isolated from rats with caerulein-induced acute pancreatitis and compared them with those in control rats. 2. We also measured amylase secretion in basal conditions and in response to cholecystokinin-octapeptide in both groups. 3. In pancreatic acini from rats with pancreatitis the basal intracellular calcium concentration was significantly increased (134.9 +/- 7.1 nmol/l compared with 71.8 +/- 2.9 nmol/l, P < 0.001). Moreover, the maximum values of intracellular calcium attained during the stimulation period were equivalent in acini from control and pancreatitic rats with no statistically significant differences. 4. In acini from control rats the differences between the resting levels of intracellular calcium and the maximum intracellular calcium values (delta[Ca2+]i) in response to several concentrations of cholecystokinin-octapeptide showed a clear dose-response relationship, with a half-maximal increase at 0.1 nmol/l and a maximal difference (delta[Ca2+]i = 259 +/- 50 nmol/l) at 1 nmol/l. In contrast, a right-shifted response, with a statistically significant smaller increase, was observed in acini from pancreatitic rats. 5. Basal amylase release was significantly higher in acini from rats with pancreatitis (11.7 +/- 1.0% of total compared with 5.9 +/- 1.1% of total, P < 0.001). In contrast, cholecystokinin-octapeptide and acetyl-choline-evoked amylase secretion was reduced by more than 85% in acini from pancreatitic rats. 6. In conclusion, calcium homoeostasis in pancreatic acinar cells from rats with caerulein-induced pancreatitis seems to be impaired. These results suggest excessive release of acinar free ionized calcium, or damage to the integrity of mechanisms that restore low resting levels of intracellular free ionized calcium, and the consequent calcium toxicity could be the key trigger in caerulein-induced acute pancreatitis.

Topics: Acetylcholine; Acute Disease; Amylases; Animals; Calcium; Ceruletide; Culture Techniques; Dose-Response Relationship, Drug; Homeostasis; Intracellular Fluid; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Sincalide

Induction by caerulein of acute pancreatitis with tissue damage and acinar cells loss is followed by recovery. We studied biochemical, histological and functional regeneration of pancreatic tissue after repeated acute pancreatitis. Pancreatitis was evoked in rats by s.c. infusion of caerulein (10 micrograms/kg/h) for 5 h. After infusion, rats were divided into three groups. First group was infused with caerulein one time, in the second group infusion of caerulein was repeated 10 days later. The third groups was infused with caerulein for the 3rd time 10 days after the 2nd infusion. Rats were sacrificed at time sequence of 0, 12, 24, 48, 72 hours and at 5th, and 10th day after last infusion of caerulein. Pancreatic blood flow (PBF) was measured using laser Doppler flowmeter. Plasma and pancreatic amylase, pancreatic weight, RNA and DNA contents, and histological changes were determined. We found that DNA and RNA content, as well, as histological changes in 1st group showed progressive regeneration after 3 days. Regeneration after 1st time caerulein-induced pancreatitis was almost completed within 10 days and amylase content in the tissue and plasma amylase level returned to normal values. Each subsequent infusion of caerulein caused significantly less pronounced destruction of the pancreatic tissue, however, the regeneration occurred progressively later than after the 1st or 2nd infusion. Tissue repair after the 2nd infusion reached peak at 5th day while after 3rd infusion at 10th day. PBF dropped after 1st caerulein induced pancreatitis by about 50% but with repeated caerulein induced pancreatitis lower decreases in PBF were observed and they returned in shorter time back to control value. These results indicate that the pancreas is able to adapt to repeated injury and this is manifested by cumulative decrease of pancreatic damage after each repetition of induction of acute pancreatitis and correlated with the preservation of PBF, however, the pancreatic tissue regeneration is significantly delayed.

Topics: Acute Disease; Adaptation, Physiological; Amylases; Animals; Ceruletide; DNA; Male; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Wistar; Regeneration; RNA

Measurement of local pancreatic blood flow during acute pancreatitis is thought of as being technically difficult in smaller animals. In this study, we first employed an intraductal electrode method of the hydrogen clearance technique for measurement of changes in local pancreatic blood flow in two types of acute pancreatitis in rats, then compared this method with an interstitial electrode method that has been used in rats. There was a very close correlation between these methods (r = 0.998, p < 0.001). The intraductal electrode method was easily performed and caused minimal tissue damage.

Topics: Acute Disease; Amylases; Animals; Blood Flow Velocity; Blood Pressure; Ceruletide; Electrodes; Hematocrit; Hydrogen; Male; Organ Size; Pancreas; Pancreatic Ducts; Pancreatitis; Rats; Rats, Sprague-Dawley

The pancreatitis-associated proteins (PAPs) are major pancreatic secretory proteins during acute pancreatitis. However, mechanisms of regulation of PAP gene expression are poorly understood, and there is a lack of information regarding mouse PAP gene expression. Herein, we employed Northern blotting and RNase protection assays to measure mouse PAP-I mRNA levels in the normal pancreas and intestine, and in the pancreas during caerulein-induced acute pancreatitis. Unexpectedly, we found that mouse PAP-I mRNA levels are constitutively high in the adult pancreas, as well as in the small intestine. Furthermore, mouse pancreatic PAP-I mRNA levels are rapidly and dramatically down-regulated (3 h) after the initiation of caerulein injections, but slowly return to high levels by 72 h. Interestingly, we found that pancreatic PAP-I mRNA levels are also transiently and dramatically down-regulated after L-buthionine-[S,R]-sulfoximine administration. Thus, a correlation between PAP-I mRNA levels and glutathione levels in the mouse pancreas was demonstrated.

Topics: Acute Disease; Acute-Phase Proteins; Animals; Antigens, Neoplasm; Biomarkers, Tumor; Blotting, Northern; Buthionine Sulfoximine; Ceruletide; Down-Regulation; Gene Expression Regulation; Glutamate-Cysteine Ligase; Glutathione; Intestine, Small; Lectins, C-Type; Male; Mice; Pancreas; Pancreatitis; Pancreatitis-Associated Proteins; Proteins; RNA, Messenger

During acute pancreatitis, data obtained in vitro suggest that pancreatic lipase, acting on circulating or tissular triglycerides, might generate nonesterified fatty acids (NEFA) that could promote pancreatic and fat tissue necrosis. This work determined whether NEFA were actually produced in vivo in pancreatic tissue and in blood during cerulein-induced pancreatitis in rats. Intraperitoneal injections of cerulein induced pancreatitis. To promote the possible NEFA release by pancreatic lipase, a venous infusion of human very low density lipoprotein (VLDL) was used to cause hypertriglyceridemia. NEFA were measured in portal and aortic blood and in tissue extracts prepared from pancreas homogenates. NEFA did not increase either in peripheral or in portal blood. In pancreatic tissue, NEFA levels did not differ from controls. The major hypertriglyceridemia produced by human VLDL intravenous infusion neither altered the course of the disease nor promoted plasma NEFA release. The role commonly attributed to NEFA in acute pancreatitis seems questionable.

Topics: Acute Disease; Animals; Ceruletide; Fatty Acids, Nonesterified; Hypertriglyceridemia; Lipase; Lipoproteins, VLDL; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

Clinical as well as experimental studies in insulinopenic diabetes mellitus have demonstrated abnormal pancreatic exocrine responses to cholecystokinin (CCK). In the present study, we examined pancreatic exocrine and endocrine function in the recently developed genetically diabetic Otsuka Long-Evans Tokushima fatty (OLETF) rats and compared them with those in the control Long-Evans Tokushima Otsuka (LETO) rats of the same age. Stepwise increasing doses of CCK octapeptide (CCK-8; 0.027-7.0 nmol.kg-1.h-1) evoked a characteristic biphasic dose-response curve for pancreatic juice and protein output in the LETO rats, whereas the OLETF rats were totally insensitive to CCK-8 stimulation. However, the responsiveness and the sensitivity to both carbamylcholine and secretin were similar in the two groups. Intraduodenal infusion of casein (500 mg/h) failed to stimulate pancreatic exocrine secretion in the OLETF rats despite a greater CCK response than in the LETO rats (peak response: 8.43 +/- 0.97 vs 5.12 +/- 0.30 pmol/l in LETO, P < 0.01). Intravenous infusion of CCK-8 (4.4 nmol.kg-1.20 min-1) caused a significant increase in serum insulin concentrations and a concomitant decrease in glucose levels in the LETO rats but not in the OLETF rats. On the other hand, an intravenous bolus injection of 1.1 mmol/kg glucose caused a greater insulin release in the OLETF rats than in the LETO rats. In contrast, gastric acid secretion in the OLETF rats was significantly high in basal and in response to intravenous infusion of CCK-8 compared with that in the LETO rats. Four subcutaneous injections of 20 micrograms/kg caerulein at hourly intervals over 3 h induced acute pancreatitis in the LETO rats but did not elicit any significant increase in serum amylase or lipase activities and pancreatic wet weight or histological evidence of acute pancreatitis in the OLETF rats. These results indicate that the exocrine and endocrine pancreas of the recently developed genetically diabetic OLETF rats are totally and specifically insensitive to exogenous and endogenous CCK stimulation, whereas parietal cells in these rats are sensitive to CCK stimulation.

Topics: Acute Disease; Animals; Carbachol; Caseins; Ceruletide; Cholecystokinin; Duodenum; Gastric Acid; Glucose; Injections; Insulin; Islets of Langerhans; Male; Obesity; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Secretin; Sincalide

Pancreatic exocrine hypofunction is markedly deteriorated during acute exacerbation in a rat model with chronic pancreatitis.. Little is known about pancreatic exocrine function during acute exacerbation in patients with chronic pancreatitis. We investigated changes in pancreatic exocrine function after inducing acute pancreatitis in an animal model of spontaneous chronic pancreatitis.. WBN/Kob rats with chronic pancreatitis sequentially underwent pancreatic exocrine function test 1-6 d after surgical preparation with external pancreatic fistula. We induced acute pancreatitis in another WBN/Kob rats by i.v. administration of cerulein at a rate of 10 micrograms/kg/h for 4 h 4 d after surgical preparation. Pancreatic exocrine function test was undertaken in a conscious state 1 d before and after cerulein administration.. In WBN/Kob rats not given cerulein, pancreatic exocrine function remained almost constant at 3-6 d after surgery. Marked hyperamylasemia developed immediately after cerulein administration. After its administration, the pancreas microscopically showed prominent interstitial edema and intracellular vacuolization of acinar cells in addition to the finding of pre-existing chronic pancreatitis. Basal and cholecystokinin-stimulated flow rate, bicarbonate output, and protein output, which were substantially impaired 1 d before cerulein administration, were further reduced 1 d after its administration.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Chronic Disease; Disease Models, Animal; Dose-Response Relationship, Drug; Male; Pancreas; Pancreatic Function Tests; Pancreatic Juice; Pancreatitis; Rats; Rats, Inbred Strains; Sincalide

The pancreatic damage initiated via different pathogenetic pathways can be increased by triglycerides. Thus, triglycerides seem to play an important role in the pathogenesis of acute pancreatitis.. Lipolytic enzymes and their substrates may play a role in the pathogenesis of acute necrotizing pancreatitis. We investigated, therefore, whether triglycerides alter the course of acute pancreatitis in three experimental models of rats.. 1. Edematous acute pancreatitis induced by repeated sc injections of cerulein; 2. Necrotizing acute pancreatitis by retrograde duct injection of sodium taurocholate; and 3. Pancreatic edema by ligation of: a. The bile duct at the liver hilus; b. The common bile/pancreatic duct close to the duodenal wall; or c. A combination of a. and b. Six hours later, rats were sacrificed and the isolated perfused pancreas prepared. The pancreases were perfused with either HEPES/Ringer/HAES alone or in combination with various concentrations of triglycerides (1-5% wt/vol). The activities of lipase and amylase in the portal venous effluents were regarded as a marker of pancreatic injury. In addition, the pancreases were evaluated by light microscopy.. In both cerulein and taurocholate acute pancreatitis, amylase/lipase activities were significantly higher compared to controls during 45 min of perfusion. In both models, addition of triglycerides caused a dose-dependent marked elevation of enzymes. Ligation (a) did not cause any rise in enzymes in the venous effluent; triglycerides had no effect. Ligation (b) or (c) caused a significant increase of pancreatic enzymes, which was further increased by triglycerides. Histology showed various degrees of severity of tissue damage depending on the model used. The additional damaging effect of a 45-min perfusion with triglycerides, however, could not be detected by histology.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Gastrointestinal Agents; Hypertriglyceridemia; In Vitro Techniques; Lipase; Male; Pancreas; Pancreatitis; Pancreatitis, Acute Necrotizing; Rats; Rats, Sprague-Dawley; Taurocholic Acid; Triglycerides

Microcirculatory derangements are important early features in many organs during the process of acute pancreatitis. However, dynamic evaluation of these factors has been difficult. Antiprotease has long been used for the treatment of acute pancreatitis, although its effects and mechanism have not been fully elucidated. The involvement of proteases and microcirculatory derangement early in the course of acute pancreatitis are the main concern of this study. A severe acute pancreatitis model in male Sprague-Dawley rats (225-275 g) was established by adding caerulein (15 microg/kg/ hr) in intravenous infusion fluid and intraductal injection of 0.1 ml glycodeoxycholic acid (5 mM). Gabexate mesilate [GM; ethyl-4-(6-guanidinohexanoyloxy)benzoate methanesulfonate], a synthetic antiprotease, was infused intravenously in doses of 0.01, 0.1, 1, and 10 mg as a therapeutic intervention in this model. Pathology hematocrit, serum amylase level, and glutamic-oxaloacetic transaminase (GOT) levels were used to confirm the severity of disease and effect of therapy. In vivo microscopic technique was used as a investigating tool in this study of microcirculatory derangement in pancreas and liver, 8 hr after induction of acute pancreatitis. GM can significantly improve pathologic criteria and changes of serum amylase levels in the range of 1-10 mg/kg/hr. The severity of changes of hematocrit and GOT was significantly lessened with GM in the range of 0.1-10 mg/kg/hr. This agent also could improve the microcirculatory environment in pancreas and liver after induction of acute pancreatitis according to the parameters, such as flow velocity and rolling leukocyte phenomenon, in the range of 1-10 mg/kg/hr. According to our observation, severity of hyperpermeability had not changed with the treatment of GM. These results indicated the beneficial effects of GM on pancreatic and hepatic microcirculation early in the course of acute pancreatitis. The beneficial effects were noted in serum parameters and hematocrit. The importance of protease activation and remote organ dysfunction is emphasized in the course of acute pancreatitis from this study.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Endothelium, Vascular; Gabexate; Leukocytes; Liver; Male; Microcirculation; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Rheology

In order to clarify the derangement of the pancreatic microcirculation in acute pancreatitis, the pancreatic microcirculation in caerulein pancreatitis was monitored intravitally and the roles of bradykinin and nitric oxide were examined using bradkykinin B2 receptor antagonist, HOE140. Under an intravital microscope, the pancreatic microcirculation was observed 2 or 6 hr after the induction of acute pancreatitis. HOE140 was administered 30 min before the induction of acute pancreatitis. The videoimages were taken into the computer, and the value of grayscale was measured with imaging software to quantify the degree of extravasation. Extravasation in the postcapillary venules was remarkable and the velocity in pancreatic terminal arterioles decreased significantly. However, the adherence of leukocytes was not observed until 6 hr after the induction. Both the extension of extravasation and the decrease of velocity were prevented by HOE140. The levels of nitric oxides in the pancreatic tissue declined and this decline was not influenced by HOE140. Bradykinin participates mainly in the regulation of vascular permeability in the early stage of caerulein pancreatitis. Further, the impairment of pancreatic microcirculation may play a key role in the onset and development of acute pancreatitis.

Topics: Acute Disease; Animals; Arterioles; Body Water; Ceruletide; Leukocytes; Male; Microcirculation; Microscopy, Fluorescence; Microscopy, Video; Nitrates; Nitrites; Pancreas; Pancreatitis; Rats; Rats, Wistar

Prostacyclin (PGI2) and its stable analogue iloprost (I) exert beneficial effect in acute pancreatitis (AP). The aim of the study was to evaluate the role of iloprost in pancreas regeneration after AP in rats.. AP was induced in male Wistar rats by s.c. injections of caerulein 12 micrograms/kg t.i.d. for 2 days. Rats were divided into four groups: control + saline (C); control + iloprost (C/I); AP + saline (AP); AP + I (AP/I). Rats were treated for 7 or 14 days. I (Schering AG) was given at the dose I microgram/kg b.w., i.p., t.i.d. After the rats were killed, the pancreata were weighed and their protein, DNA, RNA, chymotrypsin, alpha-amylase contents were evaluated. Light microscopic examination of representative pieces of pancreas was performed.. Acute pancreatitis resulted in pancreas destruction observed even 7 days after the onset of the disease. The significant decrease of pancreatic weight, RNA and chymotrypsin contents were observed in AP rats when compared to C. The improvement of pancreatic histology and significant increase of DNA content were found in I treated (during 7 days) AP rats in comparison to untreated AP group. Two weeks after pancreatitis induction the pancreas regeneration occurred in both pancreatitis groups and it was connected with pancreas hypertrophy. Treatment with I resulted in slight not significant increase of some of cellular hypertrophy indices when compared to AP untreated animals. Healthy rats injected with I during 7 days showed significant elevation of DNA content in comparison to C. When treatment with I was prolonged up to 14 days such hyperplastic effect was not observed. Our results suggest, that treatment with iloprost exerts temporary hyperplastic influence on the pancreas of healthy rats and pancreas regenerating after caerulein-induced pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Iloprost; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Regeneration

To study the importance of oxygen-derived free radicals in acute pancreatitis, an experimental study of in vivo detection of superoxide free radicals (O2-) was performed using rats. Using a new chemiluminescence probe (2-methyl-6-[p-methoxyphenyl]-3,7-dihydro-imidazol[1,2-1]pyrazin- 3-one; MCLA; a Cypridina luciferin analogue) and a highly sensitive photon counting system, O2- from the pancreatic surface of rats with experimental acute pancreatitis induced by 180 micrograms cerulein/kg was detected. The time course of MCLA-dependent luminescence suggested that O2- production began 2-3 h after cerulein injection and then decreased gradually. Superoxide free radical production in the pancreas of rats with cerulein-induced acute pancreatitis was confirmed using MCLA-dependent chemiluminescence. This new method allows direct observation of the behavior of oxygen-derived free radicals.

Topics: Acute Disease; Animals; Ceruletide; Free Radicals; Imidazoles; Kinetics; Luminescent Measurements; Male; Pancreas; Pancreatitis; Pyrazines; Rats; Rats, Wistar; Superoxide Dismutase; Superoxides

Stimulation of the exocrine pancreas with cholecystokinin analogues leads to a variety of intraacinar processes, many coupled to energy consumption. It was hypothesized that extensive ATP depletion could play a role in the pathophysiology of acute pancreatitis, especially in the hyperstimulation (cerulein) model. Mice received seven intraperitoneal injections of cerulein at hourly intervals, at doses ranging from physiological (0.1 micrograms/kg) to pharmacological (50 micrograms/kg). A single dose of cerulein induced a 28-33% decrease in ATP, whereas a complete course of injections led to a nadir as low as 45% of the control value. The overall pattern of ATP tissue content during the observed time course was surprisingly similar in all four groups and statistically not different at any time point. Until 12 h, ATP levels in all groups remained below the control value. In contrast, serum amylase and light microscopy reflected a degree of pancreatitis in a close dose-response pattern to the administered cerulein dose. These findings suggest that ATP depletion--although probably facilitating acinar damage--does not seem to play a causal or primary role in the pathophysiology of acute pancreatitis.

Topics: Acute Disease; Adenosine Triphosphate; Amylases; Animals; Ceruletide; Energy Metabolism; Female; Injections, Intraperitoneal; Kinetics; Mice; Pancreas; Pancreatitis; Vacuoles

Silver staining of nucleolar organizer regions (AgNORs) demonstrates loops of DNA that transcribe ribosomal RNA. Their number and size have been attributed to rDNA transcription activity involved in protein synthesis and thus associated with proliferation. The exact relationship among proliferation, protein synthesis, and expression of AgNORs is, however, not yet well established. We therefore investigated AgNORs in an experimental model of cerulein-induced rat pancreatitis. During secretory stimulation with maximal doses of cerulein (0.25 micrograms/kg/h) for 12 h, AgNOR number and size per nucleus as well as 3H-thymidine label index did not change, although there was a marked increase in pancreatic volume flow, up to 150%, and of protein synthesis rate, up to 180% of the control levels. In contrast, after infusion of supramaximal doses of cerulein (5.0 micrograms/kg/h), AgNOR and 3H-thymidine label values rose significantly, with a distinct peak at day 3 after induction of pancreatitis. Most interestingly, AgNOR number and size were elevated 12 h before DNA replication started as determined by 3H-thymidine incorporation. At the same time intracellular protein synthesis proved to be decreased approximately 30-50% compared to controls. Our data confirm that AgNOR is a marker of proliferation that reflects regulatory events in the cell cycle earlier than 3H-thymidine incorporation. Here we demonstrate for the first time that this phenomenon is independent of the total intracellular protein synthesis rate.

Topics: Acute Disease; Animals; Cell Division; Ceruletide; DNA; Male; Nucleolus Organizer Region; Pancreas; Pancreatitis; Rats; Rats, Wistar; Regeneration; RNA, Ribosomal; Silver Staining; Transcription, Genetic

The effects of chronic administration of hydrocortisone (10 mg/kg/day) on the development and evolution of acute pancreatitis induced by supramaximal stimulation with cerulein were examined in the rat. In these circumstances the potentially therapeutic effect of L-364,718, a CCK-receptor antagonist, was assayed. Administration of hydrocortisone over 7 days did not increase the severity of edematous acute pancreatitis induced by cerulein, since the reduction in pancreatic secretion, the hyperamylasemia and the increase in the levels of hematocrit and fluid in the pancreatic tissue were similar in rats with acute pancreatitis treated and untreated with hydrocortisone previously. When hydrocortisone was administered chronically, before administration of supramaximal doses of cerulein, a spontaneous regression of acute pancreatitis occurred. However, when hydrocortisone administration was continued after inducing pancreatitis, pancreatic recovery was prevented, observing a significantly depressed acinar secretion and elevated values of hematocrit and tissue fluid (edema). L-364,718 administration proved to be detrimental in the evolution of edematous acute pancreatitis when the rats had been treated chronically with hydrocortisone because the blockade exerted on secretion prevented the draining of enzymes stored in excess by hydrocortisone administration.

Topics: Acute Disease; Amylases; Animals; Benzodiazepinones; Ceruletide; Devazepide; Hematocrit; Hydrocortisone; Male; Pancreas; Pancreatic Juice; Pancreatitis; Rats; Rats, Wistar; Receptors, Cholecystokinin

Dextrans improve pancreatic microcirculation in acute experimental pancreatitis. They could therefore facilitate the transport of a protease inhibitor to ischemic areas of tissue injury and be of additional benefit.. To compare the effects of dextrans with and without aprotinin, necrotizing pancreatitis was induced in 33 male dextran-resistant Wistar rats by intraductal infusion of low-dose glycodeoxycholic acid (10 mmol/l) followed by intravenous cerulein (5 micrograms/kg/h) for 6 h. Three and four hours after the start of the cerulein infusion the animals received infusions of either Ringer's lactate (RL) (12 ml/kg), 70,000 Da dextran (10%) (DEX-70) (4 ml/kg) alone, or DEX-70 (4 ml/kg) with aprotinin (5000 IU/kg) (DEX-70/A).. The death rate was 60% within 9 h in the RL group (6 of 10) but only 10% in the DEX-70 group (1 of 10) (p < 0.03; Fisher's exact test) and 23% in the DEX-70/A group (3 of 13). Histomorphometry demonstrated a significant reduction of acinar necrosis in both treatment groups compared with control animals (p < 0.014; t test). Total amounts of trypsinogen activation peptides (TAP) in ascites were also significantly lower in these groups (p < 0.05; t test).. DEX-70 given 3 h and 4 h after induction of pancreatitis significantly reduced the levels of TAP, limited acinar necrosis, and improved survival rate in acute necrotizing rodent pancreatitis. There was no additional benefit from the combination with aprotinin.

Topics: Acute Disease; Animals; Aprotinin; Ceruletide; Dextrans; Glycodeoxycholic Acid; Hemodilution; Male; Necrosis; Oligopeptides; Pancreas; Pancreatitis; Plasma Substitutes; Rats; Rats, Wistar; Serine Proteinase Inhibitors; Time Factors; Trypsinogen

In an effort to elucidate factors that determine the severity of an attack of acute pancreatitis, we have quantitated the extent of necrosis and of apoptosis in five different models of experimental acute pancreatitis. Severe pancreatitis was induced by obstructing the opossum common bile-pancreatic duct, by administering to mice 12 hourly injections of a supramaximally stimulating dose of caerulein, and by feeding young female mice a choline-deficient, ethionine-supplemented diet. In each of these models of severe pancreatitis, marked necrosis but very little apoptosis was found. Mild pancreatitis was induced by obstructing the rat common bile-pancreatic duct and by infusing rats with a supramaximally stimulating dose of caerulein. In contrast to our findings in severe pancreatitis, mild pancreatitis was characterized by very little necrosis but a high degree of apoptosis. Our finding that the severity of acute pancreatitis is inversely related to the degree of apoptosis suggests that apoptosis may be a teleologically beneficial response to acinar cell injury in general and especially in acute pancreatitis.

Topics: Acute Disease; Animals; Apoptosis; Ceruletide; Cholestasis; Choline Deficiency; Constriction, Pathologic; Diet; Ethionine; Female; Male; Mice; Mice, Inbred Strains; Necrosis; Opossums; Pancreatic Ducts; Pancreatitis; Rats; Rats, Wistar

To use mice to examine the effects of cyclosporine and tacrolimus (FK 506) on two forms of acute pancreatitis often seen after clinical organ transplantation.. In the first experiment, male CD-1 mice received cyclosporine (10 mg/kg), tacrolimus (0.32 mg/kg), or saline solution (control) subcutaneously once a day for 10 days. On the 11th day, acute edematous pancreatitis was induced by ceruletide (cerulein). In the second experiment, female ICR mice were fed with a choline-deficient, ethionine-supplemented (CDE) diet for 72 hours to induce necrotizing pancreatitis. After 30 hours on the CDE diet, the mice received cyclosporine (10 mg/kg), tacrolimus (0.32 mg/kg), or saline solution (control) subcutaneously twice daily for 3 days.. The pancreatic dry-to-wet weight ratios after ceruletide injections significantly decreased in mice treated with cyclosporine but did not with tacrolimus. Cyclosporine also significantly increased serum amylase levels, but tacrolimus did not. Cyclosporine or tacrolimus alone did not produce pancreatitis. In the CDE diet groups there was a significant difference in survival among the cyclosporine-treated, the tacrolimus-treated, and the control groups.. Cyclosporine or tacrolimus given alone does not induce acute pancreatitis. In contrast, cyclosporine can adversely affect the course of acute edematous pancreatitis, and both immunosuppressants may worsen the survival of mice with acute hemorrhagic necrotizing pancreatitis. This study also demonstrated that the deteriorating effect of tacrolimus is less potent than that of cyclosporine.

Topics: Acute Disease; Animals; Ceruletide; Choline Deficiency; Cyclosporine; Edema; Ethionine; Female; Male; Mice; Necrosis; Pancreatitis; Tacrolimus

The efficacy of long acting somatostatin analogue, octreotide acetate (SMS 201-995) on the caerulein-induced acute pancreatitis and on the regeneration of the gland was examined. The effect of the drug on the acute injury was examined at 6 and 24 hours following the intervention, while the regeneration was examined on Day 3 and Day 5 in all cases by determination of plasma amylase levels and by analysis of the pancreatic tissue. The use of octreotide could not counteract the occurrence of acute pancreatitis, however, it has some benefit as seen by it's ability to moderate the increases of serum amylase levels. During the examination of pancreatic regeneration it was found that the weight of the pancrease decreased and this was not affected by octreotide. As a matter of fact, the octreotide coadministered with caerulein counteracted the caerulein-induced increase of pancreatic DNA content and therefore acted against the reactive pancreatic hyperplasia. Thus long term administration of octreotide in acute pancreatic injury may not be rational.

Topics: Acute Disease; Amylases; Animals; Ceruletide; DNA Replication; Male; Octreotide; Organ Size; Pancreas; Pancreatic Function Tests; Pancreatitis; Rats; Rats, Wistar; Regeneration

The present study was undertaken to investigate the involvement of PAF in acute pancreatitis induced by cerulein in rats. Cerulein (two doses of 20 micrograms/rat, the first s.c. and the second i.v., 1 h apart) induced a significant increase in vascular permeability in the pancreas, evaluated by the Evans blue (EB) extravasation method. Plasma amylase levels were also significantly increased in this group. The PAF antagonists, BN-52021 (5 mg/kg) and WEB-2170 (1 and 10 mg/kg), both significantly reduced the extravasation of EB in the pancrease induced by i.v. injection of PAF (1 microgram/kg). At these concentrations, BN-52021 was effective at inhibiting cerulein-induced pancreatitis (60-70% of inhibition) whereas WEB-2170 had no significant effect. Although the inhibition induced by BN-52021 suggests the involvement of PAF in cerulein-pancreatitis, the lack of effect of WEB-2170 reported here does not allow a definite conclusion. Further studies are needed to elucidate the differential effect of the PAF antagonists.

Topics: Acute Disease; Amylases; Animals; Azepines; Capillary Permeability; Ceruletide; Diterpenes; Evans Blue; Ginkgolides; Lactones; Male; Pancreas; Pancreatitis; Platelet Activating Factor; Rats; Rats, Wistar; Triazoles

Nitric oxide synthase activity is detected in the pancreas, but the role of NO on pancreatic function has not been fully characterized. The aim of this study was to evaluate the role of NO in normal and diseased pancreatic function.. Amylase and NO secretion were measured in vivo in rats and in vitro in dispersed acini, with and without NO synthesis blockade, by NG-nitro-L-arginine methyl ester (L-NAME). Rats were subjected to cerulein-induced pancreatitis, and the effects of L-NAME or NO donors were assessed.. L-NAME reduced amylase output to 60% of basal. This effect was reversed by L-arginine. The secretory response to optimal doses of cerulein induced a poor amylase secretion and a marked release of NO. High doses of cerulein in combination with L-NAME inhibited NO formation and amylase secretion. In dispersed acini, supramaximal cerulein concentrations induced NO release, but the amylase dose-response curve was not modified by NO inhibition. In acute pancreatitis, L-NAME increased amylasemia and tissue myeloperoxidase activities, whereas NO donors reduced amylasemia, lipasemia, and the histological damage score.. The L-arginine/NO pathway facilitates basal and stimulated pancreatic secretion in vivo. NO donor drugs may improve the course of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Arginine; Ceruletide; Free Radicals; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitroprusside; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Superoxide Dismutase

It has been hypothesized that microcirculatory disturbance plays an important role in the development of severe pancreatitis. In this study we investigated the effects of exogenous endothelin-1 on the development of severe pancreatitis in rats.. Acute pancreatitis was induced by two intraperitoneal injections of cerulein (10 micrograms/kg body weight). Endothelin-1 was administered via an abdominal aortic catheter as a bolus of 250-750 pmol/kg BW every hour for 4 h.. Remarkable morphologic changes in the pancreas, including hemorrhage, and increases in serum amylase level and active elastase content in pancreatic tissue were observed in rats treated with cerulein plus endothelin-1 in a dose-dependent manner 5 h after the first cerulein injection. Local pancreatic blood flow decreased significantly, and microcirculatory disturbances in the pancreas were demonstrated.. These results suggest that endothelin-1 causes pancreatic microcirculatory disturbance and might be a contributing factor in the aggravation of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Endothelins; Hemorrhage; Male; Microcirculation; Pancreas; Pancreatic Elastase; Pancreatitis; Rats; Rats, Sprague-Dawley; Regional Blood Flow

Fulminant acute pancreatitis is a disease of complex origin that results in activation of several of the proinflammatory cytokines. Because interleukin-1 (IL-1) is an integral early component of the acute inflammatory process, the use of an IL-1 receptor antagonist (IL-1ra) was investigated in experimental acute pancreatitis to determine the therapeutic potential of proximal cytokine blockade and to further establish the role of inflammatory cytokines in the pathogenesis of acute pancreatitis.. IL-1ra was administered in escalating doses either before or after acute edematous, necrotizing pancreatitis was induced in adult male mice by injection of cerulein. The severity of pancreatitis was quantified by serum amylase, lipase, interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) levels, pancreatic wet weight, and blinded histologic grading.. Administration of medium (10 mg/kg) and high (100 mg/kg) doses of IL-1ra either before or after the induction of pancreatitis significantly decreased the expected rise in pancreatic wet weight, lipase, IL-6, and TNF-alpha (all, p < 0.01). Serum amylase was significantly reduced when IL-1ra was administered in either dosage before (p < 0.05), but not after, induction of pancreatitis. Pancreatic edema, necrosis, and inflammatory cell infiltrate were significantly diminished (p < 0.05) by histologic grading in all animals receiving medium or high doses of IL-1ra. Low doses of IL-1ra (1.0 mg/kg) had modest effects if given before, but no effect if given after, induction of pancreatitis.. The proinflammatory cytokines IL-6 and TNF-alpha are elevated during experimental acute pancreatitis and correlate well with the severity of local pancreatic destruction. Blockade of the cytokine cascade at the level of the IL-1 receptor before or soon after induction of pancreatitis significantly attenuates the rise in these cytokines and is associated with decreased severity of pancreatitis and reduced intrinsic pancreatic damage.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Edema; Interleukin 1 Receptor Antagonist Protein; Interleukin-1; Interleukin-6; Lipase; Male; Mice; Necrosis; Organ Size; Pancreas; Pancreatitis; Receptors, Interleukin-1; Recombinant Proteins; Sialoglycoproteins; Single-Blind Method; Tumor Necrosis Factor-alpha

Endothelin-1, a 21-residue peptide isolated from vascular endothelial cells, has a broad spectrum of actions. To clarify the involvement of endothelin-1 in acute pancreatitis, we examined the effects of endothelin-1 and its receptor antagonist BQ-123 on cerulein-induced pancreatitis in rats. Rats were infused intravenously with heparin-saline (control), endothelin-1 (100 pmol/kg/hr), cerulein (5 micrograms/kg/hr), or cerulein plus endothelin-1 for 3.5 hr. In another experiment, cerulein or cerulein plus BQ-123 (3 mg/kg/hr) was infused. Infusion of cerulein caused hyperamylasemia and pancreatic edema. Endothelin-1, when infused with cerulein, decreased the extent of pancreatic edema with a significant increase in the pancreatic dry- to wet-weight ratio. Histological changes induced by cerulein were markedly attenuated when endothelin-1 was given with cerulein. In contrast, endothelin-receptor blockade with BQ-123 further augmented pancreatic edema caused by cerulein. The extent of inflammatory cell infiltration was greater than BQ-123 was given with cerulein. Endothelin-1 or BQ-123 had no influence on hyperamylasemia. This study suggests that endothelin-1 has protective effects on experimental acute pancreatitis.

Topics: Acute Disease; Amylases; Analysis of Variance; Animals; Ceruletide; Disease Models, Animal; Drug Evaluation, Preclinical; Endothelin Receptor Antagonists; Endothelins; Male; Pancreas; Pancreatitis; Peptides, Cyclic; Rats; Rats, Wistar

Trypsinogen activation peptide (TAP) concentration and alpha 2-macroglobulin-trypsin complex (alpha 2M-T) activity were measured in two experimental models of acute pancreatitis in rats to evaluate the significance of activation of trypsinogen in acute pancreatitis. TAP concentration and alpha 2M-T activity in serum rose significantly in trypsin-taurocholate-induced hemorrhagic acute pancreatitis, while in cerulein-induced edematous acute pancreatitis they did not rise in spite of a similar increase in immunoreactive trypsin. When rats in trypsin-taurocholate-induced pancreatitis were treated by protease inhibitor (FUT-175; nafamostat mesilate; FUT group), alpha 2M-T activity in serum was significantly lower than that in nontreated controls (mean +/- SEM, 20.8 +/- 1.43 U/L in the FUT group vs 79.1 +/- 24.5 in controls; p < 0.01). The survival rate at 24 h was significantly improved in the FUT group compared with the controls (70 vs 43%; p < 0.05). The increase in TAP concentration in the FUT group was similar to that in controls. The TAP concentration in pancreatic tissue at 24 h was significantly (p < 0.01) lower in the survival group (7.8 +/- 0.8 ng/ml) than in the lethal group (25.9 +/- 3.7 ng/ml). Activation of trypsinogen and its subsequent enzyme activity play an important role in the evolution of severe acute pancreatitis.

Topics: Acute Disease; alpha-Macroglobulins; Animals; Benzamidines; Ceruletide; Disease Models, Animal; Guanidines; Male; Oligopeptides; Pancreatitis; Proglumide; Protease Inhibitors; Rats; Rats, Wistar; Receptors, Cholecystokinin; Taurocholic Acid; Trypsin; Trypsinogen

Studies in animal models suggest that oxygen radicals are important in the pathogenesis of acute pancreatitis. Cerulein, a decapeptide isolated from the skin of the frog, Hyla caerula, is closely related to the C-terminus of cholecystokinin and it is a potent stimulant of pancreatic exocrine secretion. The aim of the present study was to measure the activity of endogenous scavengers, superoxide dismutase, catalase and glutathione levels in cerulein-induced acute pancreatitis in rats. We found that the plasma amylase and ribonuclease levels in the pancreatitis group were both significantly high (p < 0.01, p < 0.05, respectively) when compared with the control group. Although superoxide dismutase and glutathione levels of pancreatic tissue were decreased significantly (p < 0.01, p < 0.01 respectively), we observed a significant increase (p < 0.01) in catalase activity in the cerulein treated group compared to the control group. Therefore, we concluded that the profound alteration of the activities of endogenous scavengers (superoxide dismutase, catalase) and glutathione depletion occurring after cerulein-induced pancreatitis seemed to be important in tissue injury and may provide the basis for successful therapy of the disease.

Topics: Acute Disease; Amylases; Animals; Anura; Catalase; Ceruletide; Disease Models, Animal; Free Radical Scavengers; Glutathione; Pancreatitis; Rats; Ribonucleases; Superoxide Dismutase

Studies of experimental pancreatitis have generally focussed on early time points rather than the stages of healing and resolution or scarring. We recently characterized a new pancreatitis model of moderate severity by combining intraductal infusion of very low concentrations of glycodeoxycholic acid with intravenous caerulein. This study evaluates late histopathologic changes and gross complications in this pancreatitis model compared to the traditionally used high-dose bile salt model in rats. After 14 days, histopathologic features of caerulein pancreatitis were not different from saline controls. High-dose intraductal bile salt infusion resulted in widespread chronic inflammation, acinar dilation and atrophy, marked reactive stromal proliferation, and ductular budding with periductal fibrosis. In contrast, animals receiving low-dose intraductal bile salt infusion combined with intravenous caerulein demonstrated a moderate degree of chronic inflammation and acinar atrophy along with an intermediate degree of periductal fibrosis and stromal reaction. We conclude that due to its moderate degree of injury, this model may be useful for the study of tissue injury and repair following acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Glycodeoxycholic Acid; Necrosis; Pancreas; Pancreatitis; Random Allocation; Rats; Rats, Sprague-Dawley; Time Factors

Cytokines activate the hypothalamic-pituitary-adrenal axis and suppress inflammation by stimulating glucocorticoid secretion. The state of adrenocortical function during acute pancreatitis and its role in this disease were determined.. Cerulein-induced pancreatitis or closed duodenal loop pancreatitis was produced in rats that had undergone adrenalectomy or sham adrenalectomy, and the serum corticosterone and interleukin 8 levels and the intensity of the pancreatitis were examined.. Serum corticosterone levels were significantly higher than basal levels in both models of experimental pancreatitis. In both models, adrenalectomy increased serum amylase and pancreatic edema and produced more severe inflammation. Adrenalectomy significantly increased mortality in animals with closed duodenal loop pancreatitis. Exogenous hydrocortisone administered to adrenalectomized animals suppressed the elevation of serum interleukin 8 levels and decreased both the severity of pancreatitis and mortality.. These results suggest that the adrenocortical function is stimulated during acute pancreatitis and that the secretion of endogenous glucocorticoids may play an important role in mitigating the progress of this disease, probably by inhibiting cytokine production.

Topics: Acute Disease; Adrenal Cortex; Adrenalectomy; Amylases; Animals; Ceruletide; Corticosterone; Disease Models, Animal; Duodenum; Glucocorticoids; Hydrocortisone; Interleukin-8; Least-Squares Analysis; Male; Pancreatitis; Rats; Rats, Wistar

Nitric oxide (NO) has been implicated to regulate pancreatic circulation, promote capillary integrity, and inhibit leukocyte adhesion. We investigated the role of NO in the development of pancreatitis. Nitro-L-arginine, an inhibitor of NO synthase, in total dose of 35 mg/kg body wt was infused in the rats with edematous pancreatitis induced by two intraperitoneal injections of cerulein (20 micrograms/kg). L-Arginine (125 or 250 mg/kg), a NO donor was intravenously administered twice in the rats with hemorrhagic pancreatitis induced by water-immersion stress plus two intraperitoneal injections of cerulein (40 micrograms/kg). The degree of pancreas edema, serum amylase levels, and histologic alterations were investigated. Nitro-L-arginine exacerbated cerulein-induced pancreatitis and caused a decrease in pancreatic blood flow. L-Arginine ameliorated the severity of hemorrhagic pancreatitis dose dependently and improved the pancreatic blood flow. These findings suggest that NO could confer protection against the development of hemorrhagic pancreatitis, probably through improvement of the pancreatic microcirculation.

Topics: Acute Disease; Animals; Arginine; Ceruletide; Drug Interactions; Edema; Enzyme Inhibitors; Gastrointestinal Hemorrhage; Male; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pancreas; Pancreatitis; Random Allocation; Rats; Rats, Sprague-Dawley; Time Factors

Extraintestinal trypsinogen activation peptides (TAP) have been shown to correlate with severity of acute pancreatitis in humans as well as in various animal models. Ischemia superimposed on experimental pancreatitis, however, increases acinar cell injury without increasing TAP in plasma. We speculated that TAP generated in the pancreas might not reach the circulation in necrotizing pancreatitis due to decreased pancreatic perfusion. To test the hypothesis that generation of TAP in plasma is related to pancreatic perfusion and that plasma TAP may therefore underestimate acinar cell injury in necrotizing disease, we correlated TAP in pancreatic tissue and body fluids with capillary pancreatic blood flow in necrotizing and edematous pancreatitis. The ratio between necrosis and TAP in tissue was similar in both models; the ratio between TAP in plasma and tissue, however, was significantly lower in necrotizing pancreatitis, indicating that a certain amount of TAP generated in the pancreas did not reach the circulation. Decreased pancreatic perfusion found in necrotizing pancreatitis was consistent with this finding. Our data suggest that TAP in tissue is most reliable to indicate severity of acute pancreatitis, whereas plasma TAP may underestimate pancreatic injury in necrotizing disease due to decreased pancreatic perfusion.

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Edema; Glycodeoxycholic Acid; Male; Microcirculation; Necrosis; Oligopeptides; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Trypsinogen

Little is known about the pathophysiological factors that determine the clinical severity of acute pancreatitis. Because impairment of pancreatic circulation and oxygenation is associated with greater disease severity and morphological damage in experimental pancreatitis it has been suggested that various vasoactive mediators might participate in the progression from the oedematous to the necrotising variety of the disease. This study used an animal model of acute pancreatitis induced by intravenous caeruleint (10 micrograms/kg/h for up to six hours), which does not entail either haemorrhage or significant necrosis of the pancreas. This study considered whether the administration or the inhibition of either nitric oxide, bradykinin, or adrenergic mediators can convert this mild variety into haemorrhagic and necrotising pancreatitis. Neither nitric oxide nor catecholamines were involved in the progression from oedematous to haemorrhagic pancreatitis. Their substitution, activation, and inhibition all failed to change the severity of the disease process. Bradykinin alone seemed to be critically involved in the pathogenesis of pancreatic haemorrhage and necrosis. However, the inhibition of bradykinin and not its activation or substitution increased the severity of the disease.

Topics: Acute Disease; Animals; Arginine; Bradykinin; Catecholamines; Ceruletide; Edema; Labetalol; Male; Microcirculation; Microscopy, Electron, Scanning; Necrosis; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitroglycerin; Pancreas; Pancreatitis; Phenylephrine; Rats; Rats, Wistar

The present studies were done to evaluate the therapeutic potential of C1-esterase inhibitor in three different models of acute pancreatitis: (1) Edematous pancreatitis with acinar cell necrosis was induced by 7-h ip injections of 50 micrograms/kg cerulein in mice; (2) Hemorrhagic pancreatitis was induced by feeding a choline-deficient, ethionine-supplemented (CDE) diet in mice; and (3) Hemorrhagic pancreatitis was induced by retrograde infusion of 0.6 mL 5% sodium-taurocholate into the pancreatic duct in rats. C1-esterase inhibitor was given at 100 mg/kg iv before the onset of pancreatitis and at certain intervals thereafter. The severity of pancreatitis was assessed at various times after its onset by determination of serum amylase, by grading of histological alterations, and by determination of survival (survival determined only in models of hemorrhagic pancreatitis). In some of the models, C1-esterase inhibitor slightly ameliorated the degree of histological alterations; the increase in serum amylase was reduced by C1-esterase inhibitor only in CDE diet-induced pancreatitis. In all three models, C1-esterase inhibitor, however, failed to cause major beneficial effects and also failed to improve survival in taurocholate- and diet-induced pancreatitis. Additional studies in 12 patients with acute pancreatitis showed that C1-esterase inhibitor activity was markedly increased in serum of all patients during the first 9 d of the disease, suggesting that C1-esterase inhibitor behaves like an acute phase protein. Taken together the results from the animal and the human studies, C1-esterase inhibitor appears to only have a limited potential for treatment of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Complement C1 Inactivator Proteins; Diet; Disease Models, Animal; Female; Male; Mice; Pancreatitis; Rats; Rats, Wistar; Taurocholic Acid

Redistribution of vitamin E in the rat body was studied during acute pancreatitis induced by two intraperitoneal doses of cerulein 40 micrograms/kg of body weight at 1-hr intervals. Hyperamylasemia (2064 +/- 521 vs 6419 +/- 129 U/dL) and pancreatic edema (pancreatic water content, 71 +/- 1.2% vs 78 +/- 2%) were observed. In this model the increased level of lipid soluble fluorophore was also observed (274 +/- 18 vs 120 +/- 9.0 relative fluorescence per g dry wt). Parallel with these changes was a decrease in the level of vitamin E in the serum and an increase in the pancreas. The concentration of vitamin E in the pancreas after 6 h was 162 +/- 8.5 ng/mg dry mass vs 128.1 +/- 6.1 ng/mg dry mass in control animals. The effect of heparin on vitamin E redistribution induced by acute pancreatitis was also investigated. It was found that heparin at a dose of 100 U/kg body mass prevents the drop of the vitamin E level in the serum as well as the increases in the concentration in the pancreas tissue. It was concluded that acute pancreatitis induced redistribution of vitamin E in the rat body. Moreover, we studied the effects of heparin treatment on oxidative stress in the pancreas tissue. Acute pancreatitis caused an increase in lipofuscin accumulation, and a decrease in protein sulfhydryl groups in citrate synthetase (CS) and in malate dehydrogenase (MDH) activity. Heparin treatment that protected vitamin E accumulation in the pancreas tissue did not influence the changes in the level of lipofuscin and proteins sulfhydryl.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Animals; Ceruletide; Citrate (si)-Synthase; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Vitamin E

Topics: Acute Disease; Animals; Blood Flow Velocity; Ceruletide; Disease Models, Animal; Endothelin Receptor Antagonists; Endothelins; Hemorrhage; Male; Pancreatitis; Peptides, Cyclic; Rats; Rats, Sprague-Dawley; Stress, Physiological

Acute pancreatitis can be induced in the rat by high doses of cerulein, a cholecystokinin analogue. Regeneration of the pancreatic gland after this aggression can be accelerated by endogenous or exogenous cholecystokinin. However, the biochemical and molecular events associated with the cholecystokinin-induced regeneration process have not yet been identified. This study was therefore undertaken to determine the potential involvement of particulate and crude cytosolic tyrosine kinases as well as phospholipase D (PLD) in the course of pancreatitis induction and during regeneration. Acute pancreatitis was induced by cerulein, 12 micrograms kg-1, every 8 h for 2 days; this treatment was followed by 3 days of rest, and the regeneration treatment was started on the morning of the sixth day, with cerulein given at 1 microgram kg-1 every 8 h for 1-4 days. Animals were sacrificed 1, 3, 6, 12, 24, and 48 h after the first cerulein injection (high dose), on the morning of the sixth day (end of the rest period), and on the morning of the seventh and tenth days (low dose, regeneration period). After sacrifice, pancreata were excised and prepared for tyrosine kinase and PLD assays. Parallel increases in tyrosine kinase and PLD activities were observed from 6 to 48 h during pancreatitis induction and at the end of the resting period. Activities returned to control values during the regeneration period in the untreated cerulein-pancreatitis groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Animals; Ceruletide; Male; Pancreas; Pancreatitis; Phospholipase D; Protein-Tyrosine Kinases; Rats; Rats, Sprague-Dawley; Regeneration

During acute pancreatitis, experimental data obtained in vitro suggested that pancreatic lipase generates nonesterified fatty acids (NEFA), noxious for acinar cells, by hydrolysis of pancreatic or circulating triglycerides. The purpose of this work was to determine whether experimentally induced high plasma NEFA levels do indeed aggravate in vivo cerulein-induced pancreatitis. Anesthetized Sprague-Dawley rats received cerulein and were simultaneously infused intravenously with either saline or a triglyceride + heparin mixture (TGH) in order to increase the amount of circulating NEFA. Plasma NEFA increased about fourfold (3.02 +/- 0.28 mumol/liter) in animals infused with TGH with respect to controls (0.75 +/- 0.05 mumol/liter). In rats receiving cerulein + TGH, pancreatic enzyme levels in plasma, ascites, and histological alterations of the pancreas did not differ from those observed in the rats receiving cerulein + saline. There was less macroscopic pancreatic edema (P < 0.01) in the cerulein + TGH group than in the cerulein + saline group. Separate infusion of either heparin alone or of triglycerides alone had no effect. We conclude that high levels of circulating NEFA do not aggravate cerulein pancreatitis in rats and may even induce a protective effect.

Topics: Acute Disease; Animals; Ceruletide; Edema; Fatty Acids, Nonesterified; Heparin; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Triglycerides

A previous report has shown that undernutrition reduces the mortality of acute experimental pancreatitis probably by decreasing pancreatic enzyme content. Cerulein in physiological doses reduces the enzyme content of the pancreas without any harmful effect on the organ. The aim of the present study was to asses the effect of acute reduction of pancreatic enzyme content on the outcome of acute pancreatitis. Two groups of male Wistar rats weighing 230-250 g were studied: group I, 12-h fasted animals, and group II, ad libitum-fed animals who received cerulein at the inframaximal dose (0.2 microgram kg-1 h-1) for 2 h. Cerulein administration resulted in the reduction of the pancreatic contents of chymotrypsinogen (71%), trypsinogen (55%), proelastase (60%), amylase (62%) and cathepsin B (45%) (P < 0.05). However, no significant reduction in pancreatic phospholipase content was observed. Acute pancreatitis was induced in group I after 12-h fasting and in group II at the end of cerulein infusion by retrograde injection o 0.5 ml of 2.5% Na+ taurocholate into the pancreatic duct. Ascites volume and the degree of histologically observed lesions were similar in both groups, but 72-h mortality was 56% in the control group (10/ 18) and 23% (5/22) in the cerulein group (P < 0.05). We speculate that the reduction of pancreatic enzyme content may exert its beneficial effect in acute pancreatitis by decreasing the quantity of pancreatic enzymes reaching the circulation and consequently their pathogenic effects.

Topics: Acute Disease; Animals; Ceruletide; Gastrointestinal Agents; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar

This study was designed to investigate the role of nitric oxide (NO) in the formation of pancreatic edema in caerulein-induced pancreatitis in rats. Pancreatitis was produced by two intraperitoneal injections of caerulein, and plasma amylase concentration, pancreatic edema index (pancreatic wet weight/body weight), and Evans blue extravasation (as a measure of vascular permeability) were evaluated 5 h after the first injection. Four doses (1, 2.5, 5, and 10 mg/kg) of NG-nitro-L-arginine (L-NNA), an NO synthase inhibitor, were subcutaneously administered at -0.5, 0.5, 1.5, 2.5, and 3.5 h after the first injection of caerulein. L-NNA significantly lowered the edema index, the wet/dry weight ratio of the pancreas, and Evans blue extravasation in the rats with pancreatitis. The maximal effect was obtained by L-NNA at a dose of 2.5 mg/kg; this inhibited the increase in pancreatic edema formation from the control value by 60%-70%. Intraperitoneal injections (20 mg/kg, five times) of L-arginine, a substrate for NO production, partly reversed the L-NNA-induced inhibition of pancreatic edema formation, but D-arginine, an enantiomer of L-arginine, did not show any effect. Plasma amylase concentrations were not significantly affected by any dose of L-NNA, nor were they affected by L- or D-arginine. These findings strongly suggest that endogenous NO plays an important role in the formation of pancreatic edema in caerulein-induced pancreatitis in rats, probably by increasing vascular permeability and protein extravasation.

Topics: Acute Disease; Amylases; Animals; Arginine; Capillary Permeability; Ceruletide; Dose-Response Relationship, Drug; Edema; Enzyme Inhibitors; Evans Blue; Male; Nitric Oxide; Nitroarginine; Pancreatitis; Rats; Rats, Wistar

Effects of colchicine, a microtubule-disrupting agent, on rate exocrine pancreas were examined in comparison with the microtubule stabilizer Taxol for the purpose of analyzing the pathogenesis of cerulein-induced acute pancreatitis. Taxol ameliorated the inhibition of pancreatic secretion, elevation of serum amylase level, pancreatic edema, and histological alterations induced by supramaximal cerulein stimulation. In contrast, colchicine by itself and colchicine followed by cerulein stimulation (maximal and supramaximal) inhibited pancreatic secretion but did not induce the hyperamylasemia, pancreatic edema, or formation of large vacuoles, which characterized cerulein-induced pancreatitis. Electron microscopic studies in the colchicine-treated rats revealed that transport vesicles were accumulated in the supranuclear region and that no large vacuoles were observed in the apical lesion. Immunofluorescence studies confirmed that colchicine inhibited pancreatic secretion and disrupted the arrangement of microtubules. Posttreatment of colchicine did not prevent the development of cerulein-induced pancreatitis. Vinblastine, another microtubule-disrupting drug, as well as colchicine, inhibited pancreatic secretion but did not induce acute pancreatitis. The results obtained in this study suggest that microtubule disorganization at a specific step in the process of intracellular vesicular transport causes cerulein-induced pancreatitis and that this step is more apical than that at which colchicine inhibits secretion in the pancreatic acinar cell.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Colchicine; Disease Models, Animal; Gastrointestinal Agents; Male; Microscopy, Electron; Microscopy, Fluorescence; Microtubules; Paclitaxel; Pancreatic Juice; Pancreatitis; Rats; Rats, Wistar; Trypsin; Tubulin

The effect of a potent protease inhibitor, sepimostat mesilate (CAS 103926-82-5, FUT-187), on acute interstitial edematous pancreatitis induced by a supramaximal dose of cerulein, a cholecystokinin (CCK) analogue, was evaluated. The serum amylase activity increased 18-fold over normal control after the infusion of cerulein at 5 micrograms/kg/h for 6 h. The serum lipase activity showed a 235-fold increase. An elevated pancreatic water content, pancreatic interstitial edema, inflammatory infiltration and vacuolization of the acinar cells were found. Redistribution of cathepsin B shifted from the lysosomal pellet fraction to the zymogen granule pellet fraction was noted in the early stages. All these parameters of pancreatitis mentioned above were inhibited by FUT-187 pretreatment at doses of 30 to 300 mg/kg. These observations suggest that FUT-187 inhibits the redistribution of cathepsin B shift from the lysosomal fraction to the zymogen fraction in cerulein-induced acute pancreatitis and improves the parameters of acute pancreatitis.

Topics: Acute Disease; Administration, Oral; Amylases; Animals; Body Water; Cathepsin B; Ceruletide; Gastrointestinal Agents; Imidazoles; Lipase; Male; Pancreas; Pancreatitis; Protease Inhibitors; Rats; Rats, Sprague-Dawley

It has been found earlier that the bradykinin antagonist, icatibant (Hoe 140), prevents the pancreatic oedema and the ensuing hypotension and haemoconcentration, and facilitates the removal of activated enzymes form the tissue during caerulein-induced acute pancreatitis. For a potential therapeutic use of the compound in clinical situations it is essential to investigate whether the associated increase in enzyme activities in the blood serum has any adverse effects on the pancreas itself or on other organs. Normal amylase secretion into the biliopancreatic duct stimulated by a low dose of caerulein (0.4 nmol kg-1 h-1, i.v.) was not affect by icatibant (100 nmol kg-1, s.c.) Acute pancreatitis, induced by a high dose of caerulein (4 nmol kg-1 h-1 for 2 h, i.v.), resulted in elevations in the activities of amylase and lipase in the pancreatic tissue and in the blood serum lasting for at least 4 h after the end of the caerulein infusion. While the rise in enzyme activities in the blood serum was augmented in icatibant-treated rats only at the end of the caerulein-infusion, the enzyme accumulation in the pancreas was significantly reduced by icatibant for at least 4 h after the end of the caerulein infusion. The secretion of amylase and lipase into the biliopancreatic duct was significantly increased only during the first 20 min of acute pancreatitis; in rats pre-treated with icatibant, no significant increase could be observed. Twenty-four hours after induction of pancreatitis, a low-dose caerulein stimulation of the exocrine function of the pancreas led to a reduced but sustained secretion of amylase regardless of whether the animals had received icatibant or not. During the first 45 min of pancreatitis, blood glucose concentrations were significantly reduced, but returned to values not different from those obtained in saline-infused controls. This effect was not affected by icatibant. No changes in the response to an i.v. glucose tolerance test were found on the day after induction of acute pancreatitis. The serum activities of glutamic pyruvic transaminase and gamma-glutamyl transpeptidase determined up to 24 h after induction of pancreatitis were not different from saline controls. icatibant had no effect on the activities of these enzymes. It is concluded that during caerulein-induced acute pancreatitis normal exocrine secretion of pancreatic enzymes into the pancreatic duct ceases almost immediately. Pre-treatment with icatibant significantly reduces th

Topics: Acute Disease; Animals; Blood Glucose; Bradykinin; Ceruletide; Female; Lipase; Liver; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

The prophylactic and therapeutic effects of a potent cholecystokinin (CCK) receptor antagonist, L-364,718, on acute pancreatitis induced by caerulein were evaluated, analyzing morphologic and functional pancreatic parameters jointly. Edematous pancreatitis was induced by four subcutaneous injections of caerulein (20 micrograms/kg) in rats at 1-h intervals. Prophylactic administration of L-364,718 (0.1 mg/kg) prevented rise in serum amylase levels, interstitial edema, vacuolization, and impairment of pancreatic enzyme secretion that accompany caerulein-induced acute pancreatitis. After 7 days, a spontaneous regression of the morphologic alterations caused by caerulein-induced acute pancreatitis occurs; however, recovery of the secretory function of the pancreas was only reached after this period of time when L-364,718 was administered therapeutically (0.1 mg/kg/day). Prophylactically or therapeutically administered, L-364,718 exerts a beneficial effect on caerulein-induced acute pancreatitis, indicating that CCK (exogenous or endogenous) plays an important role in the development of this pathology.

Topics: Acute Disease; Amylases; Animals; Benzodiazepinones; Ceruletide; Devazepide; Injections, Subcutaneous; Male; Pancreatitis; Rats; Rats, Wistar; Receptors, Cholecystokinin; Trypsin

1. We have assessed the role of platelet-activating factor in caerulein-induced acute pancreatitis (four subcutaneous injections of caerulein at a dose of 20 micrograms/kg) by measuring platelet-activating factor levels in portal blood, pancreatic tissue and peritoneal exudate in rats with and without pancreatitis. 2. We have also observed the effect of the platelet-activating factor antagonist, BN-52021, on the hyperamylasaemia and exocrine pancreatic secretion impairment associated with pancreatitis. 3. In rats with pancreatitis the basal pancreatic flow rate was increased (1.63 +/- 0.41 versus 0.25 +/- 0.03 microliters/min). Total protein output was similar in both untreated (5.98 +/- 1.93 micrograms/min) and caerulein-injected (6.5 +/- 2.0 micrograms/min) animals. Amylase output was lower in rats with pancreatitis (19.6 +/- 4.8 mu-units/min) than in controls (39.4 +/- 16.6 mu-units/min). 4. Caerulein-treated animals had significantly higher serum amylase levels than untreated animals. BN-52021 significantly reduced the caerulein-induced hyperamylasaemia. 5. Portal blood platelet-activating factor levels increased in rats with pancreatitis and in rats infused with cholecystokinin. Rats injected with caerulein and BN-52021 had portal blood levels of platelet-activating factor that were lower than those with pancreatitis. 6. Morphological derangements associated with pancreatitis (inflammatory infiltration and cell vacuolization) were also markedly reduced in BN-52021-treated animals. 7. The results of this study suggest that platelet-activating factor is involved in the development of caerulein-induced acute pancreatitis in rats.

Topics: Acute Disease; Amylases; Animals; Ascitic Fluid; Ceruletide; Cholecystokinin; Diterpenes; Ginkgolides; Lactones; Male; Pancreas; Pancreatitis; Platelet Activating Factor; Rats; Rats, Wistar

Platelet activating factor (PAF) was administered to anesthetized rabbits with cerulein-induced acute pancreatitis to investigate the role of PAF in the development of acute pancreatitis. In acute edematous pancreatitis, induced with cerulein 20 micrograms/kg/h i.v. for 5 h, blood flow in the gastroduodenal and superior mesenteric arteries (GDAF and SMAF) had decreased significantly by 30 min and the serum amylase and lipase levels were significantly increased in the early phase. In the cerulein+PAF group, in which PAF was injected 100 ng/kg/min i.v. for 20 min simultaneously with cerulein, GDAF and SMAF declined significantly to 52 +/- 4 and 47 +/- 3% (p < 0.05), serum amylase and lipase levels rose significantly to 1,110 +/- 150 and 1,370 +/- 190% (p < 0.01) at 300 min, much higher than in the cerulein group. Furthermore, scattered hemorrhages and more marked inflammatory cell infiltration were observed histologically. These findings suggest that PAF has an additive role in the aggravation of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Hemodynamics; Lipase; Male; Pancreas; Pancreatitis; Platelet Activating Factor; Rabbits

Pancreatic enzyme secretion is inhibited during acute pancreatitis, resulting in an increase in acinar zymogen content. Since the premature activation of zymogens has been assigned a central role in the pathogenesis of acute pancreatitis, minimizing the amount of stored zymogens might lead to less severe acute pancreatitis. Inhibition of enzyme synthesis or stimulation of enzyme secretion would result in reduction of zymogen stores. Opiates have a varying effect on pancreatic secretion, depending on the dosage, site of administration, and presence of pancreatic stimulants. The effect of opiates and acute pancreatitis on individual pancreatic enzyme synthesis is unknown. The following study was undertaken in order to examine the effects of an opiate on pancreatic enzyme secretion and synthesis during experimental acute pancreatitis. Four groups of rats were studied. Group I received cerulein (25 micrograms/kg); group II received an opiate, buprenorphine (BPN, 0.5 mg/kg); and group III received cerulein and BPN. Drugs were dissolved in gelatin/saline and injected subcutaneously. A control group (group IV) received only gelatin/saline. Rats were sacrificed 4 hr after injection, and pancreatic mass was measured. Pancreatic acini were prepared and assayed for amylase and DNA content. Amylase, trypsinogen, chymotrypsinogen and lipase synthesis, and amylase secretion were measured for 2 hr. Results showed that, compared to controls, acini of rats with AP had increased amylase content, a finding consistent with decreased in vivo amylase secretion. Total protein and individual enzyme synthesis rates were significantly lower in the acini of the rats with AP than in those of the controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Amylases; Animals; Buprenorphine; Ceruletide; Chymotrypsinogen; Edema; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Trypsinogen

The decompartmentation of lysosomal compartment in pancreatic acinar cells with consecutive activation of zymogens might play an important role as a "trigger mechanism" in acute pancreatitis. The admixture of lysosomal hydrolases to secretory enzymes in pancreatic juice was found, but their role in pancreatic secretion remains obscure. The aim of the present study was to assess the fragility of pancreatic lysosomal structure after maximal (optimal) or supramaximal stimulation of rats with cerulein during 3, 6, 12 h, and after recovery. In the mitochondrial-lysosomal (M-L) and in the supernatant (S) of pancreases free (F) total (T), and fractional free (%F/T) activities of beta-glucuronidase (beta G), acid phosphatase (AcP), cathepsins (Cs), and beta-N-acetyl-hexosaminidase (NAH) were estimated. In edematous pancreatitis following supramaximal stimulation with cerulein, a significant increase of %F/T of beta G in whole homogenate began at 6 h of hyperstimulation in comparison to the control (93 vs 42% p < 0.01). This increment persisted until 12 h of hyperstimulation and declined after 24 and 48 h of recovery to 67-69%. The changes of %F/T of beta G in M-L followed those in whole homogenate, and additionally the increase free activity in S after 6 h of hyperstimulation and after 24 h recovery occurred. The respective activities of other hydrolases showed a similar pattern of changes. It is of interest that fragility of lysosomal membranes increases significantly also after maximal stimulation when inflammatory changes were absent. Our results suggest that the increase of lysosomal fragility of the pancreas is most unlikely pathological in itself, but also occurs during stimulated pancreatic secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Animals; Ceruletide; Glucuronidase; Hydrolases; Lysosomes; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar

To evaluate the factors thought to be involved in the pathogenesis of acute pancreatitis associated with alcohol.. The mechanism of alcohol-induced pancreatitis is believed to involve synergistic effects of various pathogenetic factors. The present study was designed to evaluate the possible contribution of pancreatic duct obstruction, physiologic exocrine stimulation, or secretory hyperstimulation to alcohol-induced pancreatic injury.. Wistar rats were allocated randomly to a control group (group 1), or to a group with pancreatic duct obstruction (group 2), physiologic exocrine stimulation (group 3), ductal obstruction and exocrine stimulation (group 4), or exocrine hyperstimulation with the cholecystokinin analogue cerulein (group 5). Three hours after this pretreatment, animals in each experimental group were randomly divided into two subgroups for intragastric administration of either water (groups 1A through 5A) or beer (groups 1B through 5B). Test solutions were instilled over 9 hours (total amount of alcohol administered, 4.8 g/kg). Twenty-four hours after beginning the test infusion, animals were killed for histologic evaluation of pancreatic edema and determination of an acinar cell necrosis score. Serum amylase levels were determined at 3, 9, and 24 hours.. No increase in amylase levels or significant morphologic changes were found in control animals (group 1A) or in animals subjected to physiologic exocrine stimulation (group 2A). Pancreatic duct obstruction, with or without physiologic exocrine hyperstimulation (groups 3A and 4A), and exocrine hyperstimulation (group 5A) induced pancreatitis of similar severity with minor acinar cell damage. Alcohol superimposed on exocrine hyperstimulation (group 5B) increased acinar cell injury (group 5A, 0.4 +/- 0.1 points vs 5B, 1.0 +/- 0.2 points; P < .05) and serum amylase levels at 24 hours (group 5a, 41 +/- 6 U/L vs group 5B, 72 +/- 11 U/L; P < .05), whereas no differences between subgroups A and B (water vs beer) were found in groups 1 through 4.. Our findings suggest that the pathogenesis of acute alcoholic pancreatitis may require a state of exocrine hyperstimulation, perhaps via cholecystokinin, but do not support a role for constriction or obstruction of Oddi's sphincter.

Topics: Acute Disease; Animals; Beer; Ceruletide; Constriction, Pathologic; Disease Susceptibility; Ethanol; Female; Pancreas; Pancreatic Ducts; Pancreatitis; Rats; Rats, Wistar

Pancreatic exocrine and endocrine function in postpancreatitic rats treated with cholecystokinin (CCK) receptor antagonist loxiglumide was compared with that treated with saline and CCK octapeptide (CCK-8) or with that in normal control rats. Treatment with loxiglumide (50 mg/kg body weight), CCK-8 (2.5 micrograms/kg body weight), or saline (2.5 ml/kg body weight) was given three times a day for 6 days starting 1 day after the induction of acute pancreatitis by a 4-h subcutaneous infusion of 20 micrograms/kg body weight/h of caerulein. On day 8, pancreatic exocrine and endocrine function was simultaneously determined following an intravenous injection of a mixed solution of 0.2 g/kg body weight glucose plus 100 ng/kg body weight caerulein. Basal pancreatic juice flow was significantly increased in all of the postpancreatitic rats irrespective of the treatment, whereas the maximal juice flow in the loxiglumide- and saline-treated rats was significantly low compared with the CCK-8-treated and the control rats. Basal and the peak protein outputs in the loxiglumide-treated rats were comparable to those in saline-treated rats, but were lower than those in the control or the CCK-8-treated rats. Although serum glucose concentrations in all of the postpancreatitic rats were similar to those in the control rats, stimulated as well as basal insulin release tended to be high compared with the control rats. In particular, loxiglumide-treated rats showed the exaggerated insulin response compared with other groups of rats. These present observations indicate that administration of high dose of loxiglumide for a long period decreases pancreatic enzyme output and causes insulin resistance.

Topics: Acute Disease; Animals; Ceruletide; Glucose; Injections, Intravenous; Injections, Subcutaneous; Insulin; Insulin Resistance; Insulin Secretion; Islets of Langerhans; Male; Pancreatitis; Proglumide; Rats; Rats, Wistar; Receptors, Cholecystokinin; Sincalide

To investigate the benefit of pentoxifylline in severe experimental pancreatitis.. Prospective, randomized, controlled study.. Experimental animal laboratory in a University hospital.. Forty-two adult male Sprague-Dawley rats.. Acute pancreatitis was induced by supramaximal stimulation with cerulein plus a pressure and volume controlled 10 min intraductal infusion of 10-mM glycodeoxycholic acid. Thirty minutes after pancreatitis was induced, animals were randomized to receive pentoxifylline (60 mg/kg over 2.5 hrs), or saline. All animals received fluid resuscitation with lactated Ringer's solution (8 mL/kg/hr), and surviving animals were killed at 24 hrs.. There was a progressively significant decrease in mean arterial pressure after pancreatitis was induced, with no difference between pentoxifylline-treated rats and controls. Hematocrit increased significantly in both groups at 6 hrs, and returned to baseline values at 24 hrs. Ascites volume and levels of trypsinogen activation peptide in plasma and ascites were similar in both groups. Twenty-four hour mortality was 47% for the pentoxifylline group and 52% for the control group. Histologic scores for necrosis, edema, inflammation, and hemorrhage showed no significant differences between the two groups.. Treatment with pentoxifylline failed to improve outcome in a model of severe acute pancreatitis in the rat.

Topics: Acute Disease; Analysis of Variance; Animals; Ceruletide; Chi-Square Distribution; Disease Models, Animal; Drug Evaluation, Preclinical; Glycodeoxycholic Acid; Male; Pancreas; Pancreatitis; Pentoxifylline; Prospective Studies; Random Allocation; Rats; Rats, Sprague-Dawley

Impairment of pancreatic microcirculation has often been advocated as one pathogenic mechanism in necrotizing pancreatitis. In contrast, data on pancreatic capillary perfusion in edematous pancreatitis are scarce. It was the aim of this experimental study to compare changes in pancreatic microcirculation in edematous and necrotizing pancreatitis. Twelve rabbits were allocated to two groups. Two different models of acute pancreatitis were used. Edematous pancreatitis was elicited by intravenous administration of cerulein (25 micrograms/kg/hr) (N = 6). Necrotizing pancreatitis of the biliary type was induced by pressure-controlled intraductal infusion of a mixture of taurocholate, trypsin, and blood (N = 6). Pancreatic microcirculation was quantified by means of intravital microscopy assessing functional capillary density, blood cell velocity, and distribution of the plasma marker FITC-dextran 70. Systemic hemodynamics were maintained at baseline values by fluid administration. Regardless of edema or necrosis, pronounced extravasation of FITC-dextran was recorded in the early stage of pancreatitis. In cerulein-induced pancreatitis, hyperemia developed as indicated by an increase in blood cell velocity in the presence of homogeneous capillary perfusion. In contrast, a progressive reduction of the number of perfused capillaries was detected in necrotizing pancreatitis. In conclusion, pancreatic microvascular perfusion may be regarded as an important pathogenetic factor for the determination of acute pancreatitis.

Topics: Acute Disease; Animals; Capillary Permeability; Ceruletide; Edema; Ischemia; Microcirculation; Necrosis; Pancreas; Pancreatitis; Rabbits

Contrast-enhanced computed tomography is widely used to evaluate severe acute necrotizing pancreatitis (ANP) by demonstrating areas of malperfusion, which might indicate irreversible necrosis. Because of our prior finding that the intravenous contrast medium (CM) accentuates the severity of ANP by promoting further necrosis and higher mortality, we sought to investigate the mechanism by which this injury is mediated.. Mild acute pancreatitis was induced in Sprague-Dawley rats with intravenous caerulein hyperstimulation; and severe ANP, with intravenous caerulein plus intraductal glycodeoxycholic acid. Control animals and rats with pancreatitis were randomized to be given intravenous CM or saline.. Diffuse reflectance spectroscopy was used to measure the index of hemoglobin content and oxygen saturation in pancreatic tissues in vivo.. Oxygen saturation of hemoglobin was increased in animals with mild acute pancreatitis (AP) (mean [+/- SEM], 58.7% +/- 1.2% vs 55.2% +/- 1.5% in control animals; P < .05) and was decreased in animals with ANP (51.2% +/- 1.2% vs 55.2% +/- 1.5%; P < .05). Fifteen minutes after the infusion of CM, oxygen saturation of hemoglobin significantly decreased further in animals with ANP (51.4% +/- 1.8% before infusion of CM vs 46.1% +/- 1.7% at 15 minutes; P < .05) and remained significantly below the comparable group receiving intravenous saline for the entire 60-minute test. This decrement was not observed in animals with ANP given saline or in animals with mild AP or in control animals after infusion of saline or CM. The index of hemoglobin content remained unchanged throughout the experiment in all groups.. The prolonged reduction of oxygen saturation of hemoglobin in the pancreas following the administration of intravenous CM in rats with severe ANP indicates that CM impairs the pancreatic microcirculation in necrotizing forms of AP. This may explain our previous finding that CM increases pancreatic injury and mortality in rodents with ANP, and it underlines our concern that the use of contrast-enhanced computed tomography early in human AP may promote the evolution of pancreatic necrosis.

Topics: Acute Disease; Animals; Blood Gas Analysis; Ceruletide; Contrast Media; Glycodeoxycholic Acid; Hemoglobins; Infusions, Intravenous; Male; Microcirculation; Necrosis; Oxyhemoglobins; Pancreas; Pancreatitis; Random Allocation; Rats; Rats, Sprague-Dawley; Severity of Illness Index; Sodium Chloride; Tomography, X-Ray Computed

Bacterial infection of pancreatic necrosis is thought to be a major determinant of outcome in acute necrotizing pancreatitis. The determinants and possibilities for prophylaxis are unknown and difficult to study in humans.. The time course of bacterial infection of the pancreas in a rodent model of acute necrotizing pancreatitis was characterized. The authors ascertained if there is a correlation with the degree of necrosis.. Acute pancreatitis (AP) of graded severity was induced under sterile conditions by an intravenous infusion of cerulein (5 micrograms/kg/hr) for 6 hours (mild AP), or a combination of intravenous cerulein with an intraductal infusion of 10-mM glycodeoxycholic acid (0.2 mL for 2 min for moderate AP, 0.5 mL for 10 min for severe AP). Sham-operated animals (intravenous and intraductal NaCl 0.9%) served as controls. Ninety-six hours after induction, animals were killed for quantitative bacterial examination and histologic scoring of necrosis. In addition, groups of animals with severe AP were investigated at 12, 24, 48, 96, and 144 hours.. No significant pancreatic necrosis was found in control animals (0.3 +/- 0.1) or animals with mild AP (0.6 +/- 0.1) killed at 96 hours. Necrosis scores were 1.1 +/- 0.2 for animals with moderate AP and 1.9 +/- 0.2 for animals with severe AP. Control animals did not develop significant bacterial infection of the pancreas (> or = 10(3) CFU/g). At 96 hours, the prevalence of infection was 37.5% in animals with mild AP and 50% in animals with moderate AP. In animals with severe AP, infection of the pancreas increased from 33% in the first 24 hours to 75% between 48 and 96 hours (p < 0.05). The bacterial counts and the number of different species increased with time and was maximal (> 10(11) CFU/g) at 96 hours.. Bacterial infection of the pancreas in rodent AP increases during the first several days, and its likelihood correlates with the severity of the disease. This model, which closely mimics the features of human acute pancreatitis, provides a unique opportunity to study the pathogenesis of infected necrosis and test therapeutic strategies.

Topics: Acute Disease; Animals; Bacterial Infections; Ceruletide; Colony Count, Microbial; Disease Models, Animal; Edema; Enterococcus; Escherichia coli Infections; Glycodeoxycholic Acid; Gram-Positive Bacterial Infections; Leukocytes; Male; Necrosis; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Staphylococcal Infections; Survival Rate; Time Factors

The intracellular distribution and action of a new synthetic protease inhibitor, E3123, were studied in cerulein-induced acute pancreatitis in rats. Acute pancreatitis was induced by a 4-h iv infusion of a supramaximal dose of cerulein, and was treated by prophylactic (pretreatment) or therapeutic (posttreatment) continuous administration of E3123. Pancreatic edema and hyperamylasemia were ameriolated only by prophylactic treatment. A subcellular fractionation study showed that the activities of cathepsin-B and trypsin in the zymogen granule-enriched fraction of the cerulein-pancreatitis group were remarkably increased. Both prophylactic and therapeutic treatment significantly prevented the elevation of these enzyme activities. These effects were accompanied by amelioration of pancreatic histopathological features, including intracellular vacuolization and fat necrosis. A microscopic autoradiographic study using 3H-labeled E3123 showed diffuse intracellular distribution of E3123, and the radioactivity of 3H-E3123 in the posttreatment group was three times greater than that in the pretreatment group. This study provides the first experimental evidence that, even when administered therapeutically, exogenous protease inhibitors are transported into pancreatic acinar cells, thereby reducing the severity of early intracellular alterations in cerulein-induced acute pancreatitis.

Topics: Acute Disease; alpha-Amylases; Animals; Cathepsin B; Ceruletide; Edema; Guanidines; Male; Pancreatitis; Rats; Rats, Wistar; Serine Proteinase Inhibitors; Subcellular Fractions; Trypsin; Vacuoles

Topics: Acute Disease; Animals; Ceruletide; Cysteine; Glutathione; Pancreas; Pancreatitis; Pyrrolidonecarboxylic Acid; Sulfhydryl Compounds; Thiazoles; Thiazolidines

Topics: Acute Disease; Animals; Bile Acids and Salts; Cathepsin B; Ceruletide; Male; Pancreatitis; Rats; Rats, Sprague-Dawley

The incidence of acute acalculous cholecystitis (AAC) is increasing and associated mortality is high. Biliary stasis and sludge formation are probably important factors in the pathogenesis of this disease. No data concerning the dynamics of these changes in the early phase of intensive care therapy are available. The gallbladders of 20 patients treated after major abdominal surgery in the surgical intensive care unit (SICU) with mechanical ventilation and without enteral feedings were therefore observed sonographically during the first 5 postoperative days in a prospective observational study. 20 patients treated on a regular ward after major abdominal surgery also not receiving any enteral nutrition served as control group. 24 hours after admission to the intensive care unit and on all subsequent days of observation the gallbladders of the patients in the SICU-group were significantly larger than in the control group. Sludge also appeared earlier and more frequently in the gallbladders of the SICU-patients. Lack of enteral feedings alone cannot explain these results. Positive-pressure ventilation and medications used in SICU are most likely responsible for the observed differences. Besides the necessity to make the diagnosis of AAC as early as possible, it appears to be worthwhile to investigate measures of prophylaxis. Since gallbladder distension in patients treated in SICU can be already observed on the first postoperative day it seems to be reasonable to initiate a regimen of prophylactic measures (e.g. with cholecystokinin or ceruletide) early in the course of ICU-therapy.

Topics: Abdomen; Acute Disease; Aged; Biliary Dyskinesia; Ceruletide; Cholecystitis; Cholecystokinin; Critical Care; Female; Humans; Male; Middle Aged; Parenteral Nutrition, Total; Positive-Pressure Respiration; Postoperative Complications; Prospective Studies; Risk Factors; Ultrasonography

We studied morphologic changes in a rat model of acute hemorrhagic pancreatitis in order to investigate the mechanism by which water immersion stress injures the pancreas. Acute hemorrhagic pancreatitis was induced by two intraperitoneal injections of 40 micrograms/kg body weight of caerulein at 1-h intervals under water immersion stress for 5 h at 23 degrees C. Light microscopy showed interstitial edema with inflammatory cell infiltration, degeneration and necrosis of acinar cells, and bleeding. Electron microscopy showed large autophagic vacuoles, decreased zymogen granules, and dilated rough endoplasmic reticulum in acinar cells. Basolateral exocytosis of large vacuoles and phagocytosis of the degenerated acinar cells were observed. In addition, microvascular damage, including the destruction of the capillary endothelial cells, capillary thrombosis, and the extravasation of blood cells, was seen. In contrast, in a pancreatitis model induced by caerulein injection alone, there was no bleeding, no remarkable vascular change, and no thrombosis. Degeneration and necrosis of acinar cells were less severe. In the pancreas under stress alone, microvascular damage and degeneration of acinar cells were observed. These findings demonstrate that stress injures the pancreas and worsens the pancreatitis by causing microcirculatory disturbances, such as vascular damage, thrombosis, increased vascular permeability, and bleeding. These results suggest that chemical mediators, such as free radicals and platelet-activating factor (PAF), which are produced by vascular damage and thrombosis, may accelerate the activation of zymogen proteases in acinar cells in caerulein-induced pancreatitis, leading to hemorrhagic pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Hemorrhage; Immersion; Male; Microscopy, Electron; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Stress, Physiological

Contrast-enhanced computed tomography (CECT) is used to show areas of decreased pancreatic perfusion in severe acute pancreatitis (AP). To evaluate possible adverse effects of the contrast medium (CM) on the course of AP, the impact of intravenous CM in AP of graded severity in the rat was studied.. Pancreatitis of three levels of severity was induced in Sprague-Dawley rats with intravenous cerulein hyperstimulation plus time- and pressure-controlled intraductal infusion of saline or glycodeoxycholic acid. At 7 hours, control and pancreatitis animals received intravenous ionic CM, nonionic CM, or saline. The principal outcome measures were 24-hour survival, trypsinogen activation peptides (TAP) in ascites, and histological acinar necrosis score.. There was no measurable effect of CM on the index features in control animals or animals with mild or moderate AP. In severe AP, CM caused a significant increase in mortality, ascites TAP, and necrosis score.. Intravenous CM increases pancreatic injury when administered early in the course of severe experimental AP. Because CM may convert borderline ischemia to irreversible necrosis, CECT performed early in pancreatitis to show poor perfusion and predict areas of necrosis may depict a self-fulfilling prophecy. Early CECT should be reconsidered and perhaps avoided.

Topics: Acute Disease; Animals; Ascites; Ceruletide; Contrast Media; Glycodeoxycholic Acid; Injections; Injections, Intravenous; Male; Necrosis; Pancreatic Ducts; Pancreatitis; Peptides; Rats; Rats, Sprague-Dawley; Sodium Chloride; Survival Analysis; Time Factors; Trypsinogen

Topics: Acute Disease; Aminopyrine; Animals; Ceruletide; Liver; Pancreatitis; Perfusion; Rats; Rats, Wistar

The protective effects of human urinary trypsin inhibitor against pancreatic injuries in multifactor-related experimental model of acute pancreatitis were evaluated.. Experimental study.. Acute pancreatitis was induced by short-termed (1-hour) pancreatico-biliary duct obstruction with cerulein stimulation (30 minutes; 0.2 microgram/kg per hour) and systemic hypotension (30 minutes; 30% reduction of mean arterial pressure) in rats. In this model, the protective effects of UTI against pancreatic injuries were evaluated at a dose of 10,000 U/kg per hour.. In this model, significant increases in portal serum amylase, cathepsin B and malate dehydrogenase levels were observed as compared with the control rats. The redistribution of cathepsin B from the lysosomal to the zymogen fraction and activation of trypsinogen were also observed. Moreover, the increased lysosomal and mitochondrial fragility as well as impaired pancreatic adenylate energy metabolism were noted. The therapeutic administration of human urinary trypsin inhibitor had significant protective effects against these pancreatic injuries. Furthermore, the combined prophylactic and therapeutic administration of human urinary trypsin inhibitor had more significant protective effects than only therapeutic treatment.. These results suggest the importance of timing and of selecting a pertinent protease inhibitor, such as urinary trypsin inhibitor, in the treatment of pancreatitis.

Topics: Acute Disease; Adenosine Diphosphate; Adenosine Monophosphate; Adenosine Triphosphate; Amylases; Animals; Cathepsin B; Ceruletide; Cholestasis; Disease Models, Animal; Drug Evaluation, Preclinical; Glycoproteins; Hypotension; Infusions, Intravenous; Malate Dehydrogenase; Male; Pancreatitis; Rats; Rats, Wistar; Trypsin; Trypsin Inhibitors

Our purpose was to investigate enzymatically and morphologically the acute effect of the immunosuppressive agent cyclosporin A (CsA) on the exocrine pancreas of rats. The intravenous injection of CsA 10 and 20 mg/kg body weight (BW) increased the content of pancreatic amylase and protein and decreased the content of pancreatic DNA. Histologically, we observed intraacinar vacuolization and individual cell necrosis. Under stimulation of the pancreas by two intraperitoneal injections of caerulein 5 micrograms/kg BW at 1-h intervals (which did not induce any evident change in the pancreas), CsA induced a significant increase in serum amylase and in pancreatic wet weight in a dose-dependent manner. CsA at doses of 10 and 20 mg/kg BW produced a significant increase in the content of pancreatic amylase and protein. Macroscopically, we observed marked pancreatic edema, venous dilatation, and patchy hemorrhage. Histologically, there were significant differences in the severity of intra-acinar vacuolization, interstitial edema, neutrophil infiltration, individual cell necrosis, and hemorrhage, severity of which was dose dependent. Pancreatic ductal erosion was particularly marked following treatment with CsA 20 mg/kg BW. These findings indicate that CsA accelerates abnormal pancreatic enzyme secretion and suggest that the therapeutically recommended doses of CsA can induce acute pancreatitis under stimulation of the pancreas.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cyclosporine; DNA; Male; Necrosis; Organ Size; Pancreas; Pancreatitis; Proteins; Rats; Rats, Wistar; Vacuoles

The possible role of cathepsin B in the pathogenesis of two forms of acute pancreatitis was studied using the cathepsin B inhibitor known as E-64. In an edematous, nonfatal pancreatitis induced by supramaximal doses of cerulein, increases in the serum amylase and lipase levels were less pronounced in rats pretreated with E-64. Other parameters of pancreatic injury were unaffected by inhibition of cathepsin B. In a necrohemorrhagic type of pancreatic injury induced by retrograde infusion of bile salts into the pancreatic duct, E-64 partially attenuated increases in serum levels of amylase and lipase, and in addition, reduced the activation of trypsinogen. However, the high mortality in this model of pancreatitis was not modified.

Topics: Acute Disease; Amylases; Animals; Cathepsin B; Ceruletide; Disease Models, Animal; Leucine; Lipase; Male; Pancreatitis; Rats; Rats, Sprague-Dawley

Systemic prostacyclin and thromboxane A2 production in rat experimental acute pancreatitis has been evaluated by measuring the urinary excretion of the 2,3-dinor 6-keto prostaglandin F1 alpha and 2,3-dinor thromboxane B2, respectively. Acute pancreatitis was induced by intraductal administration of 4.5% sodium taurocholate (0.1 ml/100 mg body weight) and intravenous cerulein perfusion (5 micrograms/kg/hr) for 6 hr, respectively. Urinary excretion of 2,3-dinor 6-keto prostaglandin F1 alpha and 2,3-dinor thromboxane B2 were much more important in sodium taurocholate- than in cerulein-induced acute pancreatitis. These data confirm an altered prostacyclin and thromboxane metabolism occurring in experimental acute pancreatitis. Phospholipase A2 activity and the effect of gabexate mesilate on the arachidonate metabolism were also evaluated.

Topics: 6-Ketoprostaglandin F1 alpha; Acute Disease; Amylases; Animals; Ceruletide; Epoprostenol; Gabexate; Lipase; Male; Pancreatitis; Phospholipases A; Phospholipases A2; Rats; Rats, Sprague-Dawley; Taurocholic Acid; Thromboxane A2; Thromboxane B2

The effects of single and repeated short-term (4 hr) obstruction of pancreaticobiliary duct (PBDO), with or without exocrine stimulation (intraductal hypertension) by cerulein infusion (0.2 micrograms/kg.hr), on the exocrine pancreas were evaluated in the rat. Single blockage of pancreaticobiliary duct for 4 hr caused a significant rise in serum amylase levels, pancreatic water content, and redistribution of lysosomal enzyme, cathepsin B from the lysosomal fraction to the zymogen fraction, which was considered to mean the colocalization of lysosomal enzymes with pancreatic digestive enzymes in the same subcellular compartment in acinar cells. In addition, the accelerated lysosomal and mitochondrial fragility was observed in the single pancreaticobiliary-duct-obstructed animals. Moreover, the repeated PBDO for 4 hr (2 hr in each obstruction and 1 hr of free flowing of pancreaticobiliary juice between two obstructions) caused more marked changes in almost the all parameters, and the repeated PBDO with intraductal hypertension caused an activation of trypsinogen in the pancreas, making more marked changes in almost the all parameters than the repeated PBDO only group. These results indicate that the present model of repeated PBDO with exocrine stimulation seems to be a pertinent model for gallstone pancreatitis in humans, and that redistribution of lysosomal enzymes and subcellular organellar fragility seem to play an important role in the pathogenesis of pancreatic injuries induced by PBDO, particularly by repeated PBDO with exocrine stimulation, probably via activation of trypsinogen to trypsin by lysosomal enzyme, cathepsin B.

Topics: Acute Disease; Amylases; Animals; Body Water; Cathepsin B; Ceruletide; Cholecystitis; Cholelithiasis; Male; Pancreas; Pancreatic Ducts; Pancreatitis; Rats; Rats, Wistar; Trypsin; Trypsinogen

The plasma bradykinin (BK) and serum amylase levels and histological changes in rats during the course of acute pancreatitis induced by a large dose of cerulein were examined. Animals were given four intraperitoneal injections of 20 micrograms/kg body wt of cerulein at hourly intervals. The plasma concentration of BK-like immunoreactivity (BK-LI), measured by a highly sensitive and specific radioimmunoassay established in this study, was found to reach a peak 6 h after the first injection of cerulein and then to remain elevated. On the other hand, the serum amylase and the histological alterations (i.e., interstitial edema, vacuolization, and inflammatory infiltration) were maximal 9 h after the first injection and returned to nearly normal after 24 h. These observations suggest that the BK generation is indicative of the participation of the kallikrein-kinin system in the pathophysiological change and that the plasma BK-LI level is a good marker of cellular damage and inflammation within the pancreas during the course of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Bradykinin; Ceruletide; Immune Sera; Male; Pancreatitis; Radioimmunoassay; Rats; Rats, Wistar; Sensitivity and Specificity

We evaluated the protective effect and the mechanism of action of the trypsin inhibitor, urinastatin, extracted from human urine, in experimental acute pancreatitis induced by a supramaximal dose of cerulein (5 micrograms/kg/hr for 3.5 hr). Urinastatin in a dose of 10,000 units/kg/hr was given by three different methods of continuous infusion: (1) 2 hr before and during cerulein infusion, (2) only during cerulein infusion, and (3) starting 1 hr after the beginning of cerulein infusion and continued for 3.5 hr. In protocol 1 and 2 urinastatin was significantly more protective than in 3. In protocol 1 urinastatin was very protective in all parameters tested (serum amylase level, pancreatic water and amylase content, distribution of lysosomal enzymes, cellular and lysosomal fragility). These results suggest that the administration of urinastatin before and during cerulein infusion may suppress the pathogenesis and evolution of pancreatitis by inhibiting the chain reaction of pancreatic enzyme activation closely related to redistribution of lysosomal enzyme and lysosomal fragility.

Topics: Acute Disease; Amylases; Animals; Cathepsin B; Ceruletide; Drug Evaluation, Preclinical; Extracellular Space; Glycoproteins; L-Lactate Dehydrogenase; Lysosomes; Male; Pancreas; Pancreatitis; Rats; Rats, Wistar; Time Factors; Trypsin Inhibitors

In rats we studied the effects of lactoferrin on experimental acute pancreatitis caused by a single subcutaneous injection of 100 micrograms/kg body weight of caerulein. Lactoferrin isolated from human milk was given intraperitoneally to rats immediately after the injection of caerulein. The injection of 100 mg/kg body weight of lactoferrin significantly reduced the elevation of the serum amylase level and pancreatic wet weight; histological alterations of the pancreas were markedly suppressed. These results suggested that lactoferrin had a protective effect on the biochemical and histological alterations in this model.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Female; Humans; Injections, Intraperitoneal; Iron; Lactoferrin; Male; Milk, Human; Pancreas; Pancreatitis; Rats; Rats, Wistar

This study was designed to investigate the role of bradykinin in the aggravation of acute pancreatitis. After injection of bradykinin 2 micrograms/kg to anesthetized rabbits with cerulein-induced acute pancreatitis, the pancreatic blood flow through gastroduodenal and superior mesenteric arteries (GDAF and SMAF) was determined with electromagnetic blood flow meters, the serum amylase level was measured, and pancreatic tissue was observed histologically. In rabbits treated with a supramaximal dose of cerulein alone (20 micrograms/kg/h), pancreatic blood flow was decreased and the serum amylase level was increased significantly by the early phase, and histological examination showed acute edematous pancreatitis. In rabbits treated with cerulein and bradykinin, GDAF and SMAF were significantly diminished at 300 min (51 +/- 5% and 50 +/- 4%, respectively, p < 0.05), and the serum amylase level rose significantly at 180 and 300 min (730 +/- 130% and 1,190 +/- 200%, respectively, p < 0.01) compared with rabbits treated with cerulein alone, and histological examination revealed pancreatic necrosis and greater inflammatory cell infiltration. These findings suggest that bradykinin has an additive role in the aggravation of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Bradykinin; Ceruletide; Kallikrein-Kinin System; Male; Pancreas; Pancreatic Juice; Pancreatitis; Rabbits; Regional Blood Flow

We investigated the concentration of immunoreactive pancreatic phospholipase A2 (pan-PLA2) and the catalytic activity of phospholipase A2 (CA-PLA2) in plasma and pancreases of rats with cerulein-induced acute pancreatitis. Edematous pancreatitis with ascites and fat necroses in the abdominal cavity developed after 8 h infusion of cerulein (5 micrograms/kg/h). Large vacuoles were found in acinar cells and there were small areas of acinar cell necrosis. No pathological changes were seen in saline-infused control animals. Pancreatic PLA2 was localized by immunohistochemistry in pancreatic acinar cells in both groups of animals and in the proximal tubular cells of the kidney in cerulein-infused animals. The lungs and kidneys appeared normal by light microscopy in all animals. The pan-PLA2 values increased markedly, whereas the CA-PLA2 values did not change during the cerulein-infusion. The CA-PLA2 values in the homogenates of pancreatic tissue of cerulein-infused animals did not differ significantly from those of saline-infused controls. The results indicate that the CA-PLA2 in plasma is independent from the concentration of pan-PLA2 in cerulein-induced acute pancreatitis in rat.

Topics: Acute Disease; Animals; Catalysis; Ceruletide; Hematocrit; Immunoenzyme Techniques; Immunohistochemistry; Male; Osmolar Concentration; Pancreas; Pancreatitis; Phospholipases A; Phospholipases A2; Rats; Rats, Wistar

The effects of prostaglandin E2 on the fragility of cellular and subcellular organelles in caerulein-induced acute pancreatitis were investigated in rats. PGE2 at doses of 50 and 100 micrograms/kg/hr infused for 2 hours before and during caerulein (5 micrograms/kg/hr for 3.5 hours) infusion significantly prevented the increased discharge of both amylase and lactate dehydrogenase from dispersed acini, and the leakage of cathepsin B from lysosomes and of malate dehydrogenase from mitochondria in the subcellular fraction in vitro. These results suggest that PGE2 has a cytoprotective effect against caerulein-induced pancreatitis by stabilizing cell and lysosomal and mitochondrial membranes.

Topics: Acute Disease; Animals; Cathepsin B; Cell Membrane; Ceruletide; Dinoprostone; Disease Models, Animal; In Vitro Techniques; Lysosomes; Malate Dehydrogenase; Male; Mitochondria; Pancreatitis; Rats; Rats, Wistar

We examined the stimulatory state of peritoneal macrophages (M phi) in caerulein-induced pancreatitis. Edematous pancreatitis was developed by the intravenous continuous injection of caerulein (5 micrograms/kg/hr) for 4 hr. Thereafter peritoneal M phi were collected and the activity for free radical production was measured by the reduction of nitro blue tetrazorium in the presence of phorbol myristate acetate. The increase in free radical production reached a statistical significance at 12 hr and a maximum at 20 hr after the beginning of caerulein infusion. These results suggested that the peritoneal M phi are activated even in mild edematous pancreatitis, and that their activation is involved into the mechanism of the development of remote organ failure in acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Macrophage Activation; Macrophages, Peritoneal; Male; Pancreatitis; Rats; Rats, Inbred F344

Severe acute pancreatitis is often complicated by intraperitoneal infection, resulting in multiple organ failure (MOF). It is known to elevate serum tumor necrosis factor (TNF-alpha) in patients with sepsis and/or MOF. In order to study the role of TNF-alpha in the aggravation of acute pancreatitis, we investigated TNF-alpha production by peritoneal macrophages in acute pancreatitis rat using the cerulein-induced pancreatitis model. TNF-alpha production by isolated peritoneal macrophages following lipopolysaccharide (LPS) stimulation was significantly increased in pancreatitis rats as compared with nonpancreatitis control rats (p < 0.001). Serum TNF-alpha activity was elevated following intraperitoneal administration of LPS as the septic challenge both in pancreatitis rats and in control rats, being significantly higher in the former (p < 0.05). Histological findings and liver function tests revealed that LPS induced more severe liver damage in pancreatitis rats than in control rats within 24 h after LPS administration. These results indicate that increased TNF-alpha production by peritoneal macrophages in acute pancreatitis augmented LPS-induced liver injury and suggest the possibility that TNF-alpha may play a role in the development of MOF during acute pancreatitis complicated by intraabdominal sepsis.

Topics: Acute Disease; Animals; Ceruletide; Lipopolysaccharides; Macrophages, Peritoneal; Male; Multiple Organ Failure; Pancreatitis; Rats; Rats, Wistar; Tumor Necrosis Factor-alpha

Infections from enteric bacteria are a major cause of morbidity and mortality during acute pancreatitis (AP), but the pathways by which these organisms reach distant organs remains speculative. Experiments were conducted to determine if bacterial translocation could be a mechanism for infection during this disease. AP was induced in Lewis rats by i.v. infusion of caerulein (experiment I) or ligation of the head of the pancreas (experiment II). In a third experiment, rats were gavaged with 1 x 10(8) 14C-radiolabeled Escherichia coli and pancreatitis was induced with caerulein. Results in all three experiments showed that AP increased the number of viable bacteria recovered in peritoneal fluid, mesenteric lymph nodes (MLN), liver, lungs, and pancreas. Radionuclide counting indicated that AP enhanced the gut permeability to 14C E. coli. To estimate the impact of AP on the magnitude of translocation and on the ability of the host to clear bacteria, the nuclide and colony-forming units (CFU) ratios were calculated between animals with and without AP. Blood, peritoneal fluid, and MLN had the highest nuclide ratio. During AP, these tissues may be the principal routes for bacterial spreading from the gut lumen. Peritoneal fluid, pancreas, and lung were the tissues with the highest CFU ratio. Bacterial killing ability of these tissues is likely impaired during AP.

Topics: Acute Disease; Animals; Ascitic Fluid; Ceruletide; Escherichia coli; Escherichia coli Infections; Ligation; Liver; Lung; Lymph Nodes; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Lew

Four models of acute pancreatitis have been previously developed that use the ex vivo perfused isolated canine pancreas preparation. The four models include the intraarterial infusion of oleic acid (FFA) that mimics hyperlipemic pancreatitis, partial obstruction of the pancreatic duct with secretin stimulation (POSS) that mimics gallstone pancreatitis, a 2-hour period of ischemia before perfusion (ISCH 2) that mimics shock pancreatitis, and the infusion of cerulein at supramaximal stimulatory doses (CER), which lacks an obvious clinical counterpart. In the FFA, POSS, and ISCH 2 pancreatitis, but not in the CER pancreatitis, toxic oxygen metabolites, generated by the enzyme xanthine oxidase (XO), have been shown to be important mediators in the early pathogenesis. Ordinarily XO primarily occurs as xanthine dehydrogenase (XD) but can be converted to XO, which is the form that generates toxic oxygen metabolites. This conversion of XD to XO may take place either reversibly by way of sulfhydryl group oxidation or irreversibly by means of proteolytic cleavage of XD. This study was undertaken to investigate the mechanism of conversion of XD to XO in the FFA-, POSS-, and ISCH 2-induced pancreatitis models. CER pancreatitis was studied for comparison. After 4 hours of perfusion, pancreatitis was manifest by edema, weight gain, and hyperamylasemia in all four models. Dithiothreitol, a sulfhydryl group protector, ameliorated the weight gain in the FFA (40 +/- 14 gm to 18 +/- 13 gm; p < 0.05), POSS (28 +/- 10 gm to 9 +/- 3 gm; p < 0.05), and ISCH 2 pancreatitis (30 +/- 13 gm to 15 +/- 3 gm; p < 0.05), and ameliorated the hyperamylasemia in the POSS pancreatitis (12,062 +/- 4304 units/dl to 5877 +/- 2659 units/dl; p < 0.05). The CER pancreatitis was not ameliorated with dithiothreitol. A serine protease inhibitor of low molecular weight, phenylmethylsulfonyl fluoride, ameliorated only the CER pancreatitis (weight gain from 28 +/- 10 gm to 17 +/- 10 gm, p < 0.05; amylase activity from 38,116 +/- 6491 units/dl to 23,372 +/- 11,654 units/dl, p < 0.05), and not the FFA, POSS, or ISCH 2 pancreatitis. We conclude that in the three models of pancreatitis (FFA, POSS, and ISCH 2) that are mediated by toxic oxygen metabolites, XD is converted to XO reversibly by way of sulfhydryl group oxidation rather than irreversibly by way of proteolysis. In the CER pancreatitis, where XO does not play a role in the pathogenesis, proteolytic enzymes may be important mediators in the injury.

Topics: Acute Disease; Animals; Ceruletide; Dithiothreitol; Dogs; In Vitro Techniques; Ischemia; Oleic Acid; Oleic Acids; Pancreas; Pancreatitis; Phenylmethylsulfonyl Fluoride; Secretin; Time Factors; Xanthine Dehydrogenase; Xanthine Oxidase

Phospholipase A2 (PLA2) has been postulated to play an important role in the pathogenesis of acute pancreatitis. To study the mechanism through which PLA2 may cause cellular damage, we used an in vitro model of isolated rat pancreatic acini prepared by collagenase digestion. Newly synthesized proteins were labeled by [35S]methionine. Cellular destruction was measured by the degree of release of radiolabeled proteins. Incubation of pancreatic acini with PLA2 alone caused only minor damage when very high concentrations of this enzyme were used. However, when acini were incubated with PLA2 in combination with its substrate, lecithin, cells were destroyed in a time- and concentration-dependent manner. Incubating cells with pancreatic homogenates and lecithin caused damage only when there had been prior activation of homogenates with either trypsin or enterokinase. The damage could be simulated by incubating acini with pure lysolecithin. Alcohol and cerulein did not further increase the destruction caused by PLA2 and lecithin. When acini were incubated with supernatants from another set of acini to which oleic acid had been added, a similar degree of damage resulted as compared with acini incubated with oleic acid alone. However, adding PLA2 to supernatants from acini preincubated with fatty acids significantly increased the degree of cellular necrosis. The destruction by PLA2 and lecithin was inhibited by albumin but could not be inhibited by gabexate mesilate, nafamostat mesilate, or cytidine diphosphocholine. We conclude that PLA2 could play a role in pancreatic acinar cell damage, especially in the spread of cellular necrosis within the organ, provided that its substrate, lecithin, is present.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Albumins; Animals; Ceruletide; Cholecystokinin; Ethanol; In Vitro Techniques; Male; Oleic Acid; Oleic Acids; Pancreas; Pancreatitis; Phosphatidylcholines; Phospholipases A; Phospholipases A2; Proteins; Rats; Rats, Sprague-Dawley

The current study was done to evaluate the effects of short term (60 minutes) pancreatic biliary duct obstruction (PBDO) with intraductal hypertension (IDH) stimulated by secretin (0.2 clinical unit per kilogram per hour) and caerulein (0.2 microgram per kilogram per hour) plus 30 minutes of temporary pancreatic ischemia (ISCH) produced by ligation of celiac and superior mesenteric artery on the exocrine pancreas and protective effects of a new potent protease inhibitor, ONO3307 in combination with xanthine oxidase inhibitor, allopurinol, in this multifactor related model of acute pancreatitis in rats. Twelve hours after PBDO with IDH plus ISCH, we observed hyperamylasemia (23 +/- 3 units per milliliter) (p < 0.01); moderate pancreatic histologic changes; pancreatic edema (water content--81 +/- 2 percent) (p < 0.02), as well as the impaired amylase (2,889 +/- 328 units per kilogram per hour) (p < 0.01) and cathepsin B output (7 +/- 3 units per kilogram per hour) (p < 0.01) into the pancreatic juice of rats stimulated by caerulein (control group--serum amylase levels, 6 +/- 1 units per milliliter; pancreatic water content, 74 +/- 1 percent. Furthermore, PBDO with IDH plus ISCH caused the redistribution of lysosomal enzyme from lysosomal fraction (12 kilo times gravity pellet; 40 +/- 3 percent; p < 0.01) to zymogen fraction (1.3 kilo times gravity pellet; 38 +/- 3 percent; p < 0.01) (control group--12 kilo times gravity pellet, 59 +/- 2 percent; 1.3 kilo times gravity pellet, 24 +/- 2 percent) and the impaired pancreatic adenylate energy metabolism (0.79 +/- 0.02, p < 0.02) (control group--energy charge equals 0.88 +/- 0.01). Only PBDO with IDH caused no significant changes. Although only ONO3307 or allopurinol therapy showed the partial significant protective effects against pancreatic injuries, improving serum amylase levels, the administration of ONO3307 in combination therapy with allopurinol showed almost complete protective effects against the pancreatic injuries induced by PBDO with IDH plus ISCH (serum amylase levels, 9 +/- 2 units per milliliter; pancreatic water content, 76 +/- 2 percent; amylase and cathepsin B output, 7,127 +/- 946 and 18 +/- 3 units per kilogram per hour; 1.3 kilo times gravity pellet, 28 +/- 2 percent; 12 kilo times gravity pellet, 54 +/- 2 percent, and energy charge equals 0.85 +/- 0.02).(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Acute Disease; Allopurinol; Animals; Ceruletide; Cholestasis, Extrahepatic; Drug Therapy, Combination; Guanidines; Ischemia; Male; Pancreas; Pancreatic Ducts; Pancreatitis; Rats; Rats, Wistar; Secretin; Serine Proteinase Inhibitors

The redistribution of cathepsin B, a lysosomal enzyme, from the lysosomal pellet to the zymogen pellet in the subcellular fractionation, the colocalization of cathepsin B with digestive enzyme, and increased cellular, lysosomal, and mitochondrial fragility within acinar cells have been found during the early stages of caerulein-induced acute pancreatitis in rats. In the present study, the authors investigated the protective effects of prostaglandin E1 and E2, a combined therapy of these prostaglandins, and a new, synthetic, low molecular weight protease inhibitor, ONO3307, on the exocrine pancreas in this noninvasive model of experimental pancreatitis in vivo and in vitro. Prostaglandin E2, but not E1, prevented hyperamylasemia, congestion of amylase and trypsinogen in the acinar cells, redistribution of cathepsin B, and amylase and lactate dehydrogenase discharge from the dispersed acini. It also prevented cathepsin B leakage from the lysosomes and malate dehydrogenase leakage from the mitochondria in an almost dose-dependent manner, particularly at the dose of 100 micrograms/kg/hr continuous infusion. Furthermore, the combined therapy of prostaglandin E2 with ONO3307 strongly inhibited all the parameters tested in this study. This combination therapy seems to be the most effective against secretagogue-induced pancreatic injuries. These results indicate that cellular and subcellular organellar fragility seem to be closely involved in the pathogenesis of acute pancreatitis. Prostaglandin E2 seems to have important cytoprotective effects on the biologic membranes, such as a stabilizer of lysosomal or mitochondrial membranes. In addition, these findings also suggest the crucial roles of some unknown proteases in the etiology of acute pancreatitis, and indicate the clinical effectiveness of prostaglandins and this type of low molecular weight protease inhibitor for acute pancreatitis.

Topics: Acute Disease; Alprostadil; Amylases; Animals; Body Water; Cathepsin B; Ceruletide; Dinoprostone; Guanidines; L-Lactate Dehydrogenase; Lysosomes; Male; Microsomes; Pancreas; Pancreatitis; Rats; Rats, Wistar; Serine Proteinase Inhibitors; Subcellular Fractions; Trypsinogen

Caerulein-induced acute pancreatitis was studied in rats. Consistent with this type of acute pancreatitis morphological (edema, leukocytic infiltration and acinar cell vaculization) and biochemical (increase in pancreatic protein content. PAF release and serum amylase) changes developed 5 hours after caerulein administration. In addition increase in pancreatic weight and decrease in pancreatic blood flow were noticed. PAF administration caused pancreatic damage similar in some parameters to caerulein-induced pancreatitis, along with reduction of pancreatic blood flow, increase in pancreatic protein content, and serum amylase. TCV-309, a selective PAF antagonist, administered prior to caerulein and/or PAF, reduced caerulein-induced pancreatitis and prevented PAF-induced pancreatitis. Results of our present studies indicate the crucial role of PAF in pathogenesis of experimental acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Isoquinolines; Male; Pancreas; Pancreatitis; Platelet Activating Factor; Pyridinium Compounds; Rats; Rats, Wistar; Regional Blood Flow; Tetrahydroisoquinolines

A supramaximal dose of caerulein (5 micrograms/kg.hr for 3.5 hours) caused an acute pancreatitis with marked hyperamylasemia and intense interstitial edema in rats. In this model of pancreatitis, the redistribution of lysosomal enzyme in acinar cells as well as the increased lysosomal and mitochondrial fragility were also observed. The combined therapy of a low molecular weight protease inhibitor, FOY, a synthetic platelet activating factor (PAF) antagonist, CV 6209, and a xanthine oxidase inhibitor, allopurinol produced more significant improvements in all the parameters examined than the therapy of any only one of these three agents, each only one therapy exerting a partial significant protective effect. These results indicate that several factors, such as unknown proteases activities, PAF and oxygen-derived free radicals may be involved in the pathogenesis of pancreatic injuries in this caerulein-induced pancreatitis. These results also suggest that such a combined therapy of different kinds of agents, whose therapeutic mechanisms are also different, is useful in the clinical treatment of acute pancreatitis.

Topics: Acute Disease; Allopurinol; Animals; Ceruletide; Drug Therapy, Combination; Gabexate; Male; Pancreatitis; Platelet Activating Factor; Pyridinium Compounds; Rats; Rats, Wistar; Xanthine Oxidase

The effect of taxol, which is a microtubule stabilizer, was examined in a model of acute edematous pancreatitis induced in rat by the administration of caerulein. Prophylactic administration of taxol ameliorated inhibition of pancreatic secretion, increased level of serum amylase, pancreatic edema, and histological alterations in this model. Immunofluorescence studies revealed that taxol stabilized the arrangement of microtubules by the action of promoting tubulin polymerization and prevented inhibition of pancreatic digestive enzyme secretion. In isolated rat pancreatic acini, taxol reversed the inhibition of amylase secretion induced by supramaximal concentrations of cholecystokinin octapeptide and did not affect the binding of cholecystokinin octapeptide to its receptor. The results obtained in this study suggest that microtubule disorganization is the initiating event in caerulein-induced pancreatitis and that the inhibition of pancreatic digestive enzyme secretion by interfering with intracellular vesicular transport due to microtubule disorganization causes caerulein-induced pancreatitis.

Topics: Acute Disease; Alkaloids; Amylases; Animals; Cell Separation; Ceruletide; Edema; Fluorescent Antibody Technique; Male; Microtubules; Paclitaxel; Pancreas; Pancreatitis; Rats; Receptors, Cholecystokinin; Sincalide; Trypsin; Tubulin

The role of exogenous and endogenous cholecystokinin has been studied in the process of pancreatic regeneration after acute pancreatitis. A mild form of pancreatitis was induced in rats by subcutaneous cerulein at 12 micrograms.kg-1, three times a day for 2 days. After 3 days of rest, the cerulein-treated rats were divided into four groups: rats with acute pancreatitis fed 20% casein, who received no treatment; rats fed 50% casein; rats fed 20% casein supplemented with 1% soybean trypsin inhibitor (SBTI); and rats fed 20% casein who received 1 microgram.kg-1 of subcutaneous cerulein, three times a day. Controls were fed 20% casein plus saline subcutaneously. Rats were killed after 5, 10, or 20 days of treatment. Pancreatitis resulted in significant decreases in pancreatic weight and contents of protein, amylase, chymotrypsin, RNA and DNA. During the regenerative process, 1 microgram.kg-1 of cerulein increased all parameters to control values within 5 days and induced pancreatic growth thereafter. SBTI restored the pancreas to normal after 10 days with cellular hypertrophy; the 50% casein diet gave a response similar to SBTI without hypertrophy. It can be concluded that cerulein and SBTI can accelerate pancreatic regeneration after an attack of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Body Weight; Ceruletide; Cholecystokinin; Chymotrypsin; Drug Therapy, Combination; Glycine max; Hypertrophy; Male; Nucleic Acids; Organ Size; Pancreas; Pancreatitis; Proteins; Rats; Rats, Inbred Strains; Regeneration; Trypsin Inhibitors

To define the role of free radicals and of lipid peroxide involvement during the progress of cerulein-induced acute pancreatitis in mice, we evaluated the effect of a novel free radical scavenger, 2-octadecylascorbic acid (CV-3611), on pancreatic edema formation, and the levels of serum enzymes (amylase, lipase) and of lipid peroxide in pancreatic tissue. Mice were divided into three groups: control group, intraperitoneal injection of saline only; pancreatitis group, cerulein 50 micrograms/kg injected intraperitoneally six times at 1-hr intervals; treatment groups, CV-3611 10 mg/kg subcutaneously just after intraperitoneal cerulein injection. After the cerulein injection, the degree of pancreatic edema formation, serum amylase and lipase levels, and the amount of lipid peroxide in pancreatic tissue increased significantly during the observation period of 12 hr. Treatment with CV-3611 resulted in significant reduction in pancreatic edema formation at 3.5 hr (P less than 0.05) and 9 hr (P less than 0.05), serum amylase and lipase levels at 3.5 hr (P less than 0.05) and 12 hr (P less than 0.05), and lipid peroxide levels at 3.5 hr (P less than 0.05), 6 hr (P less than 0.05) and 12 hr (P less than 0.05). These results indicate that a novel free radical scavenger, CV-3611, has a strong therapeutic effect during the development of acute pancreatitis and suggest that oxygen-derived free radicals play an important role in the pathogenesis of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ascorbic Acid; Ceruletide; Female; Free Radical Scavengers; Lipase; Mice; Mice, Inbred BALB C; Pancreatitis; Peroxidases

A study was made with different doses of cerulein (2, 4, 10 and 20 micrograms/kg) administered subcutaneously to rats by four injections at intervals of 1 hr; the aim of this work was to study exocrine pancreatic secretion of the rat under cerulein-induced acute pancreatitis, analyzing enzyme and hydroelectrolyte secretion of pancreatic juice. A further aim was to study the relationship between the dose of cerulein and the plasma levels of peptides controlling hydroelectrolyte secretion of the pancreas, like secretin and vasoactive intestinal peptide (VIP). At the lowest dose schedule, the amounts of total protein and enzymes (amylase and trypsin) in pancreatic juice decreased significantly, plasma amylase increased, and the pancreas became edematous. Higher doses magnified these effects. By contrast, ductular function (flow and HCO3-) was well preserved in cerulein-treated rats, and this was probably due to the significant increase in plasma levels of immunoreactive secretin whereas VIP levels were unchanged. The secretin released by treatment with cerulein is able to palliate the lack of flow from acinar origin that is affected in the process of acute pancreatitis, being a beneficial response to the cerulein treatment.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Drug Administration Schedule; Injections, Subcutaneous; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Reproducibility of Results; Secretin; Vasoactive Intestinal Peptide

We investigated the effects of ketanserin, a S 2 (5-hydroxytryptamine 2; 5-HT 2)-serotonergic receptor antagonist, on cerulein-induced pancreatitis in the rat. Large pharmacological doses of cerulein induced acute pancreatitis in the rat. Ketanserin reduced the cerulein-induced increase in serum amylase concentration in a dose-dependent manner. Treatment with 10 mg/kg of ketanserin per os markedly improved cerulein-induced pancreatitis and was associated with a significant reduction of the increase in serum amylase concentration. In addition, a very specific serotonin S 2 antagonist, ritanserin which has no antihypertensive effect, also reduced the cerulein-induced increase in the serum amylase concentration. These results suggest that S 2 (5-HT 2) may play a role in pathophysiology of cerulein-induced pancreatitis in the rat.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Dose-Response Relationship, Drug; Ketanserin; Male; Pancreatitis; Rats; Rats, Inbred Strains; Ritanserin; Serotonin

Ischemia has been considered to play a role in the development of acute pancreatitis. The aim of this study was to investigate the effect of ischemia, caused by hemorrhagic shock, on cerulein-induced acute pancreatitis in rats. Acute pancreatitis was induced by the intravenous infusion of a supramaximally stimulating dose of cerulein (10 micrograms/kg/hr) for 6 hr. Hemorrhagic shock was induced by the removal of blood until the mean arterial blood pressure reached 35 mm Hg. This level was maintained for 30 min, after which time all the blood was reinfused. Hemorrhagic shock alone induced no morphological change in the pancreas. However, after the induction of hemorrhagic shock in animals treated with cerulein, hemorrhage and parenchymal necrosis were frequently observed in the pancreas. Seven of 20 rats (35%) receiving cerulein plus hemorrhagic shock had died by 48 hr after the start of cerulein infusion, whereas none of the rats in the cerulein or shock group died during this experiment. Cathepsin B activity in the pancreas of the cerulein plus shock group was significantly higher than in the other groups at 48 hr. These results suggest that ischemia may be a contributing factor in the pathogenesis of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Cathepsin B; Ceruletide; Ischemia; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Shock, Hemorrhagic

The association of serum amylase activity with the extent of pancreatic injury in acute pancreatitis is unclear. To clarify this relationship, we induced acute pancreatitis ranging from mild to lethal in 118 Sprague-Dawley rats (350-450 g). This was achieved by controlled intraductal infusion of low- or high-dose bile salt, with or without enterokinase, followed by intravenous cerulein or saline for 6 hr. Serum amylase was measured at baseline and 6 hr. Pancreatic histopathology was evaluated by two blinded pathologists employing total surface scoring (N = 118) and morphometric 20-field documentation (N = 22). Serum amylase correlated best with edema (r = 0.61) and fat necrosis (r = 0.58), less well with acinar necrosis (r = 0.53) and inflammation (r = 0.50), and poorly with hemorrhage (r = 0.33) and perivascular infiltrate (r = 0.31). Inasmuch as edema and fat necrosis are not important determinants of severity, these observations could explain the poor prognostic value of serum amylase activity in patients with acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Glycodeoxycholic Acid; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

The present study investigated the protective effects of the new potent synthetic protease inhibitors, ONO3307 (4-sulfamoyl phenyl-4-guanidinobenzoate methanesulfonate) and FOY007 (gabexate misilate), on the exocrine pancreas in rat caerulein-induced acute pancreatitis in both in vitro and in vivo experiments. These protease inhibitors prevented hyperamylasemia, pancreatic edema, congestion of amylase, redistribution of lysosomal enzyme in acinar cells, and lactic dehydrogenase (LDH) discharge from dispersed acini. They also inhibited the cathepsin B leakage from the lysosomes in a dose-dependent manner in doses of 2-10 mg/kg.h of ONO3307 and 20-50 mg/kg.h of FOY007. These results indicate that both ONO3307 and FOY007 exert protective effects against pancreatitis at subcellular levels in lysosomes and cellular or organelle membranes. Proteases appear to be important in the pathogenesis and development of acute pancreatitis, and low-molecular-weight protease inhibitors may be of clinical use in the treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Body Water; Cathepsin B; Ceruletide; Edema; Gabexate; Guanidines; L-Lactate Dehydrogenase; Lysosomes; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Serine Proteinase Inhibitors

The study investigated the protective effect of a new synthetic protease inhibitor, E-3123, a 4-guanidinobenzoate methanesulphonate, on the exocrine pancreas in caerulein-induced pancreatitis of rats both in vivo and in vitro. Hyperamylasaemia, pancreatic oedema and congestion of amylase, as well as cathepsin B leakage from lysosomes and malate dehydrogenase leakage from mitochondria, were prevented by infusion of 5 mg/kg.h E-3123 particularly when infused for 2 h before and during 5 micrograms/kg.h caerulein infusion for 3.5 h. The results indicate that E-3123 plays its protective roles against pancreatitis in the subcellular compartments such as lysosomes and mitochondria, and that such a low molecular weight protease inhibitor as E-3123 may be clinically useful in the treatment of acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Body Water; Ceruletide; Guanidines; Lysosomes; Male; Mitochondria; Pancreas; Pancreatitis; Rats; Rats, Wistar; Serine Proteinase Inhibitors

1. The novel bradykinin antagonist, HOE 140, completely blocked the fall in rabbit blood pressure caused, not only by i.v. bradykinin, but also by i.v. kallikrein. This shows that both the effects of exogenously administered bradykinin and those of endogenously released kinins are antagonized by HOE 140. 2. Acute pancreatitis was induced in rats by i.v. infusion of the cholecystokinin analogue, caerulein. This treatment resulted in massive oedema of the pancreas, increased activities of amylase and lipase in serum and a characteristic, biphasic fall in blood pressure. 3. HOE 140 prevented the caerulein-induced pancreatic oedema and the second phase of hypotension whereas NPC 349, a widely used, but short-acting, bradykinin antagonist did not show a significant inhibition. HOE 140, in contrast to its inhibitory effects on caerulein-induced pancreatic oedema and hypotension, significantly augmented the increases in amylase and lipase activities in serum. 4. It is concluded that in this model of acute pancreatitis, the release of kinins induces pancreatic oedema and hypotension. Prevention by HOE 140 of the kinin-induced oedema allows the pancreatic enzymes to leave the tissue without hindrance and thus will diminish subsequent pathological events. It is suggested that the results obtained with the highly potent and long-acting bradykinin antagonist, HOE 140, provide a pharmacological basis for a clinical trial in acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Anti-Inflammatory Agents, Non-Steroidal; Blood Pressure; Bradykinin; Ceruletide; Female; Kallikreins; Lipase; Oligopeptides; Pancreatitis; Rats; Rats, Sprague-Dawley

The importance of platelet activating factor in acute pancreatitis was examined by determining the tissue content of endogenous platelet activating factor and the protective effects of TCV-309, a highly selective platelet activating factor blocker, against caerulein induced pancreatitis in rats. Infusion of caerulein (10 micrograms/kg/h) for five hours resulted in about 70% increase in pancreatic weight, 22% rise in protein content, 50% reduction in tissue blood flow, nine fold increase in tissue level of platelet activating factor and 165% rise in plasma amylase as well as histological evidence of acute pancreatitis. Such infusion of caerulein in chronic pancreatic fistula rats caused a marked increase in protein output from basal secretion of 10 mg/30 minutes to 40 mg/30 minutes in the first hour of infusion followed by a decline in protein output to 15-20 mg/30 minutes in the following hours of the experiment. Exogenous platelet activating factor (50 micrograms/kg) injected ip produced similar alterations in weight, protein content, blood flow, and histology of the pancreas but the increment in serum amylase was significantly smaller and pancreatic secretion was reduced below the basal level. TCV-309 (50 micrograms/kg) given ip before caerulein or platelet activating factor administration significantly reduced the biochemical and morphological alterations caused by caerulein and abolished those induced by exogenous platelet activating factor. These results indicate that platelet activating factor plays an important role in the pathogenesis of acute pancreatitis probably by reducing the blood flow and increasing vascular permeability in the pancreas.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Isoquinolines; Male; Pancreatic Juice; Pancreatitis; Platelet Activating Factor; Pyridinium Compounds; Rats; Rats, Inbred Strains; Tetrahydroisoquinolines

In order to clarify the effect of prostaglandin E2 (PGE2) on cerulein-induced rat pancreatitis, we investigated the interaction of PGE2 with cerulein or secretin. Intravenous infusion of 10 micrograms/kg.h cerulein inhibited external secretion of the pancreas from one hour and caused macroscopic edema at 3 hours. Administration of PGE2 relieved the inhibitory effect of supramaximal dose of cerulein and decreased the pancreatic edema. The 100 micrograms/kg.hr PGE2 had no significant effect on the pancreatic juice volume and amylase secretion stimulated with 0.2 micrograms/kg.hr of cerulein. Intravenous injection of 100 micrograms/kg PGE2 inhibited both the volume and amylase secretion of pancreatic juice stimulated with 1 U/kg.h of secretin. The protective effect of PGE2 on cerulein-induced pancreatitis was not the stimulation on secretion but caused the cytoprotective effect of PG such as stabilization of cytoplasmic and lysosomal membrane.

Topics: Acute Disease; Animals; Ceruletide; Dinoprostone; Drug Interactions; Male; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

Extreme maldistribution and immediate establishment of severe cellular injury are typical features of traditional bile salt models of acute pancreatitis; both factors complicate assessment and interpretation of therapeutic benefits in trials of experimental therapy. Even more important, both are indications not of the desired induction of pancreatitis but rather of local injury by barotrauma and noxious chemicals. This study contrasts the severity and regional variability of cellular injury in traditional high-dose bile salt models with that seen in a new preparation employing the combination of intravenous (iv) caerulein (CAE) and intraductal (id) low-dose glycodeoxycholic acid (GDOC). Thirty-six male Sprague-Dawley rats (350-450 g) were induced with (group A) high-dose GDOC id (34 mmol/L), low-dose GDOC id (10 mmol/L) (group B), or low-dose GDOC id combined with caerulein iv for 6 h (group C). The regional distribution of histopathologic injury within the pancreas was assessed in 20 fields/organ by two pathologists unaware of the induction technique used. High-dose GDOC id (group A) resulted in extremely heterogenous distribution of injury for all variables (edema, p = 0.001; acinar necrosis, p = 0.0001; inflammation, p = 0.0001; and hemorrhage p = 0.001). The lesions were confined to the head of the pancreas, which showed large areas of necrosis involving entire lobules, whereas adjacent areas were unaffected. Low-dose GDOC id (group B) was more homogenously distributed, but the injury was mild and regional variability (edema, p = 0.0001; acinar necrosis, p less than 0.04; inflammation, p = 0.0001; and hemorrhage p less than 0.05) was still demonstrable. In contrast, low-dose GDOC id combined with CAE iv (group C) produced moderately severe pancreatitis, which equally affected all areas of the gland. There were no geographical differences in acinar necrosis or inflammation. This feature of the new model provides a desirable prerequisite for accurate and reproducible assessment of histopathology in studies aimed at detecting effects of therapy. We suggest that it replace traditional id bile salt infusion models.

Topics: Acute Disease; Animals; Bile Acids and Salts; Ceruletide; Computer Simulation; Disease Models, Animal; Infusions, Intravenous; Infusions, Parenteral; Male; Pancreatitis; Rats; Rats, Inbred Strains

Rats were chronically fed either an ethanol-containing diet (36% of total calories derived from alcohol) or a pair-fed, control diet (no alcohol) for 8 wk, and acute pancreatitis (AP) was subsequently induced by a 3-h i.v. infusion of caerulein (CR) at a dose of 5 micrograms/kg/hr. CR-induced AP in control rats (no alcohol) was characterized by a significant elevation in serum lipase content, pancreatic interstitial edema, infrequent occurrences of karyorrhexis, and the appearance of vacuoles in acinar cells. Chronic feeding of the ethanol diet followed by treatment with CR resulted in increases in serum lipase content, interstitial edema, karyorrhexis, and acinar vacuolization that were significantly greater than that seen in rats fed the control diet and treated with CR. It is concluded that chronic ethanol intake in the rat intensifies AP that is subsequently induced by CR.

Topics: Acute Disease; Alcoholism; Animals; Ceruletide; Disease Models, Animal; Edema; Lipase; Lysosomes; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Vacuoles

Topics: Acute Disease; Animals; Carbachol; Ceruletide; Culture Techniques; Enzyme Precursors; Pancreas; Pancreatitis; Rats

This study was undertaken to determine the involvement of endogenous cholecystokinin (CCK) in the regeneration of pancreatic tissue after cerulein-induced acute pancreatitis treated by the CCK receptor antagonist L364,718. Acute pancreatitis was induced in rats by s.c. injections of cerulein in gelatin (12 micrograms/kg) three times a day for 2 days with controls receiving saline in gelatin. Rats were then divided into four treatment groups: saline-dimethyl sulfoxide (DMSO) (SD), saline-L364,718 (SA), cerulein-pancreatitis-DMSO (CD), and cerulein-pancreatitis-L364,718 (CA). In the first experiment, rats were treated for 3 or 10 days with DMSO or L364,718 (0.1 mg/kg, twice a day). In the second experiment, rats were treated for 13 days with DMSO or L364,718 (1.0 mg/kg, twice a day). After the rats were killed, pancreata were weighed and evaluated for their total protein, amylase, chymotrypsin, RNA, and DNA. We found that destruction of the pancreatic tissue occurred after cerulein-induced pancreatitis and that regeneration of the tissue was in progress but incomplete after 10 days; the low dose of L364,718 did not prevent regeneration. After 13 days, regeneration was still incomplete but the 1-mg dose of L364,718 strongly inhibited spontaneous regeneration. These data suggest that endogenous CCK is an important and potent trophic factor in the regeneration process of pancreatic tissue following an episode of acute pancreatitis.

Topics: Acute Disease; Animals; Benzodiazepinones; Ceruletide; Cholecystokinin; Devazepide; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Receptors, Cholecystokinin; Regeneration

The present studies were done to evaluate the therapeutic potential of several antioxidants and free radical scavengers in three different models of acute pancreatitis. (a) Edematous pancreatitis with acinar cells necrosis was induced by seven hourly intraperitoneal injections of 50 micrograms of caerulein per kg in mice. (b) Hemorrhagic pancreatitis was induced by feeding a choline-deficient, ethionine-supplemented (CDE) diet in mice. (c) Hemorrhagic pancreatitis was induced by retrograde infusion of 0.6 ml of 5% sodium taurocholate into the pancreatic duct in rats. The following antioxidants and free radical scavengers were given at various doses intravenously, subcutaneously, or intraperitoneally before the onset of pancreatitis: Ebselen [2-phenyl-1,2-benzisoselenazol-3(2H)-one], superoxide dismutase, catalase, deferoxamine (Desferal), dimethyl sulfoxide, or allopurinol. The severity of pancreatitis was assessed at various times after its onset by determination of serum amylase and pancreatic weight (edema), by grading of histological alterations, and by determination of survival (survival determined in models of hemorrhagic pancreatitis). In general, free radical scavengers and antioxidants ameliorated edema and inflammation to a greater degree than necrosis and the increase in serum amylase. Superoxide dismutase (as did Ebselen in previous studies) exerted beneficial effects on survival in diet-induced pancreatitis in the absence of marked effects on pancreatic necrosis, suggesting that these beneficial effects are due to amelioration of extrapancreatic complications that often contribute to mortality in acute pancreatitis. None of the antioxidants had major beneficial effects in taurocholate-induced hemorrhagic pancreatitis. Thus, formation of free radicals may be important for progression and outcome in diet-induced and, to a lesser degree, in caerulein-induced pancreatitis but not at all in taurocholate-induced pancreatitis. Different models of pancreatitis may, therefore, involve different degrees and mechanisms of free radical formation. Despite the amelioration of edema and the beneficial effects on mortality seen for some antioxidants in some of the models, antioxidants and free radical scavengers appear to have only a limited potential for treatment of acute pancreatitis.

Topics: Acute Disease; Administration, Oral; Allopurinol; Animals; Antioxidants; Catalase; Ceruletide; Choline Deficiency; Deferoxamine; Diet; Dimethyl Sulfoxide; Disease Models, Animal; Ethionine; Female; Free Radical Scavengers; Injections, Intraperitoneal; Injections, Intravenous; Injections, Subcutaneous; Male; Mice; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Severity of Illness Index; Superoxide Dismutase; Taurocholic Acid

Existing models of acute pancreatitis have limitations to studying novel therapy. Whereas some produce mild self-limited pancreatitis, others result in sudden necrotizing injury. The authors developed an improved model providing homogeneous moderately severe injury by superimposing secretory hyperstimulation on minimal intraductal bile acid exposure. Sprague-Dawley rats (n = 231) received low-pressure intraductal glycodeoxycholic acid (GDOC) at very low (5 or 10 mmol/L) concentrations followed by intravenous cerulein. Cerulein or GDOC alone caused only very mild inflammation. However, GDOC combined with cerulein was uniformly associated with more edema (p less than 0.0005), acinar necrosis (p less than 0.01), inflammation (p less than 0.006), and hemorrhage (p less than 0.01). Pancreatic injury was further increased and death was potentiated by increasing volume and duration of intraductal low-dose GDOC infusion. There was significant morphologic progression between 6 and 24 hours. The authors conclude that (1) combining minimal intraductal bile acid exposure with intravenous hyperstimulation produces homogeneous pancreatitis of intermediate severity that can be modulated at will; (2) the injury is progressive over at least 24 hours with finite mortality rate; (3) the model provides superior opportunity to study innovative therapy.

Topics: Acute Disease; Animals; Blood Chemical Analysis; Ceruletide; Disease Models, Animal; Glycodeoxycholic Acid; Hemodynamics; Infusions, Intravenous; Male; Pancreas; Pancreatic Ducts; Pancreatitis; Pulmonary Gas Exchange; Rats; Rats, Inbred Strains

In a variety of animal models of acute pancreatitis, cholecystokinin-receptor antagonists have ameliorated the injury response. These results suggest that cholecystokinin may play a primary role in the pathogenesis of pancreatitis initiated by multiple stimuli. In an effort to test this theory, a sensitive and high affinity cholecystokinin-receptor antagonist L364,718 was administered to four different models of acute pancreatitis that were produced in the ex vivo perfused canine pancreas preparation. The four models of pancreatitis were initiated by cerulein infusion, partial duct obstruction with secretin stimulation, oleic acid infusion, and a 2-hour period of ischemia. In each model, pancreatitis was manifest by edema formation, weight gain, and hyperamylasemia during a 4-hour perfusion. In cerulein infusion-induced pancreatitis L364,718 inhibited edema formation and weight gain (31 +/- 5 gm versus 7 +/- 6 gm; p less than 0.05) and significantly decreased plasma amylase activity (36,605 +/- 21,216 U/dl versus 9421 +/- 5149 U/dl; p less than 0.05). The acute pancreatitis induced by the other three stimuli was not ameliorated by L364,718 treatment. We conclude that in the ex vivo-perfused canine pancreas preparation cerulein-induced pancreatitis is mediated at least in part by the cholecystokinin receptor. Early blockade of the cholecystokinin receptor was of no benefit in treating the other models of pancreatitis, suggesting that cholecystokinin is not involved in the early pathogenesis.

Topics: Acute Disease; Amylases; Animals; Benzodiazepinones; Ceruletide; Cholecystokinin; Devazepide; Dogs; In Vitro Techniques; Pancreas; Pancreatic Ducts; Pancreatitis; Perfusion; Receptors, Cholecystokinin; Reference Values

This study evaluated the effects of the seleno-organic substance Ebselen [2-phenyl-1,2-benzisoselenazol-3(2H)-one] in two models of acute hemorrhagic and acute edematous pancreatitis. Ebselen is known to catalyze glutathione peroxidase-like reactions and to inhibit lipid peroxidation. Hemorrhagic pancreatitis was induced by feeding a choline-deficient, ethionine-supplemented (CDE) diet to mice for 66 h. Edematous pancreatitis was induced by 7-h subcutaneous injections of 50 micrograms/kg of cerulein in mice. Ebselen was given from the beginning of the CDE diet either as a subcutaneous injection of 100 mg/kg at 6-h intervals or was mixed in with the CDE diet to yield a daily dose of 100 mg/kg of Ebselen. In further experiments, Ebselen was given at various time intervals after the beginning of the CDE diet as subcutaneous injections of 100 mg/kg at 6-h intervals. In the cerulein model, Ebselen was given 5 min prior to each cerulein injection at doses from 10-500 mg/kg. Prophylactic administration of Ebselen given orally or subcutaneously significantly improved survival from 38.5% in the control group of saline-injected CDE-fed mice to 61.9 and 65.0%, respectively. Ebselen also reduced increases in serum amylase and pancreatic weight in the diet model. Therapeutic administration of Ebselen significantly increased survival only when injections were started 20 h after the beginning of the CDE diet (64%), but not when started after 40 h (44%). Similarly, increases in serum amylase and pancreatic weight due to the CDE diet were significantly reduced by Ebselen only when injections were started after 20 h but not when started after 40 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Amylases; Animals; Azoles; Ceruletide; Choline Deficiency; Diet; Disease Models, Animal; Ethionine; Female; Free Radicals; Isoindoles; Male; Mice; Organ Size; Organoselenium Compounds; Pancreas; Pancreatitis; Selenium

Alpha-adrenergic drugs commonly are used to treat hypotension resulting from severe acute pancreatitis. It was shown previously that although systemic arterial pressure is increased by phenylephrine, pancreatic microcirculatory perfusion is decreased. Because inadequate tissue perfusion may be critical in the progression of edematous pancreatitis to parenchymal necrosis, it was hypothesized that vasoconstrictors might be harmful in pancreatitis. Therefore the effect of phenylephrine on cerulein-induced mild pancreatitis were studied. Sprague-Dawley rats (n = 54) were randomly allocated to 6 experimental groups and subjected to the following infusion regimens: (1) cerulein (cae) + phenylephrine (phe), (2) cae + saline (NS), (3) NS + phe, (4) cae + phenoxybenzamine (pbz) + phe, (5) NS + pbz + phe, and (6) NS. Initial and terminal hematocrit, serum amylase activity, and blood ionized calcium concentration were determined. The animals were killed 9 hours after starting the infusion. Macroscopic and histologic changes were scored by a 'blinded' pathologist. Phenylephrine increased the severity of cerulein-induced pancreatitis as manifested by statistically significant adverse changes in serum amylase, hematocrit, ionized calcium, peripancreatitic soap formation, and acinar cell vacuolization. These changes were antagonized by alpha-adrenergic receptor blockade with phenoxybenzamine. It is concluded that phenylephrine is deleterious in acute experimental pancreatitis, the first demonstration of such an effect by a pharmacologic vasoconstrictor, and suggested that microcirculatory changes may be important in the transition of mild to severe pancreatitis. Caution in the use of vasoconstrictor drugs in patients with acute pancreatitis is recommended.

Topics: Acute Disease; Amylases; Animals; Calcium; Ceruletide; Fat Necrosis; Hematocrit; Hemodynamics; Hypotension; Male; Pancreas; Pancreatitis; Phenylephrine; Random Allocation; Rats; Rats, Inbred Strains; Regional Blood Flow

A supramaximal dose of caerulein (5 micrograms/kg.hr for 3.5 hours) caused edematous acute pancreatitis in rats, characterized by portal hyperamylasemia (32 +/- 3 U/ml) and pancreatic edema (pancreatic water content, 86 +/- 2%) [control group: amylase, 8 +/- 1 U/ml; water content, 74 +/- 2%]. In this model, increased portal levels of malate dehydrogenase (148 +/- 25 U/ml), increased mitochondrial fragility and impaired pancreatic energy charge level (0.77 +/- 0.05) were also observed [control group: malate dehydrogenase, 54 +/- 11 U/ml; energy charge level, 0.94 +/- 0.03]. Administration of gabexate mesilate, FOY, in a dose of 50 mg/kg.hr for 2 hours before and during the caerulein infusion had a significant protective effect against these pancreatic injuries (portal amylase level, 11 +/- 2 U/ml; MDH level, 72 +/- 19 U/ml; E.C., 0.89 +/- 0.02; water content, 76 +/- 2%). FOY in a dose of 20 mg/kg.hr was partially protective. These results indicate that subcellular organelle fragility and malfunction are closely related to the pathogenesis of acute pancreatitis and suggest the usefulness of FOY in the treatment of this disease.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Gabexate; Malate Dehydrogenase; Male; Mitochondria; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

We investigated correlations between serum biochemical changes and pathological alterations in caerulein-induced acute pancreatitis in rats. Seven consecutive doses of caerulein consisting of 50 mu g/kg, were given intraperitoneally, in 40 male Sprague-Dawley rats at hourly intervals. Serum biochemical and histological changes of the rats were studied at hours 7, 9, 12, 18, 24 and 36 after the first injection, as well as day 3 and 5 respectively. Serum amylase in the controlled rats (n = 6) was 2549 +/- 221 U/L (mean +/- SE). It was elevated to twenty times that of the baseline value (52630 +/- 11397 U/L) at the seventh hour, declined to about two times the control (5520 +/- 800 U/L) at the 18th hour, and returned to the baseline level at the 24th hour after the first caerulein injection. Serum lipase, elevated since the 7th hour, reached its peak value (631 +/- 34 U/L) at the 12th hour, then declined abruptly to the baseline value (0 U/L) at the 24th hour, after the first injection. However, severe histological changes of the pancreas were apparent at the 7th hour, reaching maximal destruction at the 18th hour, with severe inflammatory changes at the 24th hour; all after the first injection. The frank inflammation did not subside until 5 days after the first injection. These results suggest that, in the case of acute pancreatitis, normalization of serum biochemistries does not indicate recovery of the pancreas from acute inflammation.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

Acute oedematous pancreatitis was induced in Wistar male rats by intravenous (i.v.) infusion of cerulein at the rate of 5.10(-6)g.kg-1.h-1 for 3 or 6 hrs. The effects of dimethylsulfoxide (DMSO)-hydroxyl radical scavenger in this model of disease were examined. DMSO was injected i.v., 0.75 g.kg-1, at 0 and 3 hrs during cerulein infusion. Treatment with this agent exerted a protective effect on the rat pancreas in the cerulein-induced acute pancreatitis. This effect is expressed by inhibition of lipid peroxidation in pancreatic tissue, less oedema formation, diminution of hyperlipasemia and a significant reduction in acinar cell vacuolisation. Our data suggest that besides the hydroxyl radical, other oxidants (secondary?) may cause cytotoxicity in this model of disease. We suggest that phagocytic inflammatory cells provide such oxidants.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Dimethyl Sulfoxide; Free Radical Scavengers; Lipase; Lipid Peroxidation; Male; Pancreatitis; Rats; Rats, Inbred Strains; Superoxide Dismutase

We evaluated the changes of lysosomal and digestive enzymes in the exocrine pancreas after caerulein induced acute pancreatitis in rats. The serum amylase levels and water content as well as pancreatic amylase and cathepsin B contents increased significantly in the early stage (0-12 h) after caerulein was administered, however, returned to the normal levels at 36 h. In the early stage, colocalization of lysosomal enzyme and digestive enzyme was found. Histologically, in the early stage, there were remarkable changes such as acinar cell vacuolization and interstitial edema, but these changes disappeared at 36 h. Furthermore, amylase and cathepsin B outputs decreased significantly in the early stage (12 h) but at 24 h, these increased significantly. LDH discharge from dispersed acini and cathepsin B leakage from lysosomes also increased in the early stage (0-12 h), but these values returned to the normal levels at 36 h. These results indicate that exocrine pancreas needs about 36 h to recover from the caerulein induced acute pancreatitis, and in this recovering process, secretion of colocalized digestive enzyme and lysosomal enzyme seem to play an important role.

Topics: Acute Disease; Amylases; Animals; Cathepsin B; Ceruletide; Lysosomes; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

The pathogenesis of pancreatitis-related pulmonary injury was studied at the light- and electronmicroscopic level. Experimental pancreatitis was induced in rats by infusion of supramaximal doses of cerulein for 12 h. Investigations were carried out 3, 6, and 12 h after the start of infusion and 12, 48, and 72 h after the end of pancreatitis induction. Initial manifestations of pancreatitis-associated lung injury revealed a pronounced clustering of polymorphonuclear leukocytes in pulmonary microvessels, followed by severe damage of alveolar endothelial cells. Consecutively, the increase in vascular permeability of the lung resulted in interstitial edema formation. Structural changes were maximal after 12 h and reversed completely after 84 h. In conclusion, the structural appearance of pulmonary injury in cerulein-induced pancreatitis was similar to that reported in early stages of the adult respiratory distress syndrome (ARDS). It is suggested that polymorphonuclear granulocytes play a crucial role in the pathogenesis of pancreatitis-related lung injury.

Topics: Acute Disease; Animals; Capillaries; Ceruletide; Endothelium; Granulocytes; Lung; Lysosomes; Male; Microscopy, Electron; Neutrophils; Pancreas; Pancreatitis; Pulmonary Alveoli; Rats; Rats, Inbred Strains; Respiratory Distress Syndrome

In this study we report the functional changes in isolated perfused lungs from rats with cerulein-induced experimental pancreatitis. Rat lungs isolated immediately after the cerulein infusion demonstrated decreased pressor responses to angiotensin II (A II) and acute hypoxia (FIO2: 0.0). The lung wet- to dry-weight ratio was increased, as was the lung-leak index, consistent with high-permeability edema formation in the lung. Neither saline-solution infusion for 12 h nor perfusion with cerulein of rat lungs isolated from untreated animals caused lung injury or functional alterations. The changes in pulmonary vascular reactivity were normalized 48-72 h after induction of pancreatitis. In conclusion, we describe an animal model of pancreatitis and reversible, ARDS-like lung injury.

Topics: Acute Disease; Angiotensin II; Animals; Capillary Permeability; Ceruletide; Hypoxia; Lung; Male; Organ Size; Pancreatitis; Rats; Rats, Inbred Strains; Respiratory Distress Syndrome; Vascular Resistance; Vasoconstriction

The purpose of this study was to assess the involvement of oxygen radicals in acute edematous and hemorrhagic pancreatitis. Acute pancreatitis was induced in rats by the CCK-analogue cerulein (5 micrograms/kg/h) and by retrograde injection of 5% sodium taurocholate for 30 min, 3.5 h, and 12 h. At the end of the infusion and observation time, serum enzymes, conjugated dienes, and malondialdehyde in the tissue were measured. Moreover, the tissue samples underwent light microscopical examination. In cerulein pancreatitis, an interstitial edema and intravascular margination of granulocytes in the pancreatic gland were observed after 3.5 h. After 12 h, the histological evaluation revealed a pronounced zymogen degranulation, extensive tissue necrosis and migration of granulocytes into the tissue. Parallelly, amylase and lipase increased by 15 and 35 times, respectively. In contrast, conjugated dienes and malondialdehyde increased in cerulein pancreatitis and reached their highest level after 3.5 h and decreased to normal levels after 12 h. The development of the histological damages and serum enzyme levels with sodium taurocholate pancreatitis was similar as compared to the cerulein pancreatitis, however, the development was faster and more traumatic. Already after 3.5 h an extensive zymogen degranulation and cell necrosis was observed. Concomitantly, the amylase and lipase levels increased by 90 and 30 times, respectively. Treatment with superoxide dismutase (100,000 U/kg/h) and catalase (400,000 U/kg/h) prevented lipid peroxidation and reduced zymogen degranulation and tissue necrosis. Tissue edema and inflammatory response were not affected in both models of acute pancreatitis.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Animals; Catalase; Ceruletide; Free Radicals; Lipid Peroxidation; Male; Oxygen; Pancreatitis; Rats; Rats, Inbred WKY; Superoxide Dismutase; Taurocholic Acid

The present study investigated the protective effect of a new potent synthetic protease inhibitor, E-3123 (4-guanidinobenzoate methanesulfonate) on the exocrine pancreas in the caerulein induced experimental pancreatitis both in-vivo and in-vitro at 3 different doses (1, 2, and 5 mg/kg.hr). This protease inhibitor prevented hyperamylasemia, pancreatic edema, congestion of amylase, and both amylase and lactic dehydrogenase (LDH) discharge from dispersed acini, as well as cathepsin B leakage from lysosomes and malate dehydrogenase (MDH) leakage from mitochondria in a dose-dependent manner, particularly in doses of 2 and 5 mg/kg.hr. Furthermore, the combined prophylactic and therapeutic use of this agent seems to be very effective in preventing caerulein induced pancreatitis. These results indicate that E-3123 plays its protective roles against pancreatitis in the subcellular compartment: lysosomes, mitochondria, cellular or organella membranes. It is hoped that such a low molecular weight protease inhibitor as E-3123 will be clinically useful in the treatment of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Guanidines; Male; Organelles; Osmotic Fragility; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Serine Proteinase Inhibitors

Microvascular permeability was studied in the isolated perfused rat pancreas using a rapid multiple indicator-dilution technique. Capillary extractions, permeability-surface area products (PS), and extravascular volumes of distribution (EVV) were determined for 22Na+, 51Cr-labeled EDTA, [57Co]-cyanocobalamin (B12), and 125I-labeled insulin at various perfusion flows. Permeability to albumin was negligible. PS for Na+ and EDTA increased with increasing flow, whereas PS for cyanocobalamin and insulin approached diffusion-limited exchange at flows greater than 3 ml.min-1.g-1. Permeability coefficients for Na+, EDTA, B12, and insulin were 36, 22, 11, and 3.48 x 10(-5) cm/s, respectively, and the permeability ratio for B12/insulin (3.16) indicated restricted diffusion to insulin. In the presence of unlabeled B12 and insulin EVV (0.15-0.19 ml/g) for EDTA, B12 and insulin approximated the interstitial volume. Caerulein-induced pancreatitis or treatment with the synthetic protease inhibitor camostate had no significant effects on permeability. In caerulein-treated rats, EVV for B12 was elevated (0.17 +/- 0.01 vs. 0.28 +/- 0.06; P less than 0.01), reflecting the interstitial edema associated with this model of pancreatitis. Permeability of the rat pancreatic microvasculature is similar to that of other fenestrated tissues, but it is 10- to 20-fold greater than that of continuous capillaries. Contrary to previous assumptions, permeability does not appear to be increased after induction of acute interstitial pancreatitis.

Topics: Acute Disease; Animals; Capillary Permeability; Ceruletide; Edetic Acid; Endothelium, Vascular; Esters; Gabexate; Guanidines; Indicator Dilution Techniques; Insulin; Male; Microcirculation; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Serum Albumin

Acute edematous pancreatitis was induced in rats by iv infusion of caerulein (CR) in a supramaximal dose of 7.5 x 10(-6)g x kg-1 x hr-1 during 6 hr. The most important finding of our study was the marked decrease of protein and nonprotein thiol content in pancreatic tissue of rats with CR-induced acute pancreatitis (AP). Oxygen radicals as well as 4-hydroxyalkenals resulting from lipid peroxidation are believed to be at least partly responsible for this phenomenon. Covalent binding of excessive amounts of 4-hydroxyalkenals to pancreatic tissue protein sulfhydryl groups has been documented. Presented data suggest a serious disturbance of sulfhydryl compounds metabolism in pancreatic tissue of rats with CR-induced AP which may be of importance in the pathogenesis of the disease.

Topics: Acute Disease; Animals; Ceruletide; Free Radicals; Lipid Peroxidation; Male; Pancreas; Pancreatitis; Proteins; Rats; Rats, Inbred Strains; Sulfhydryl Compounds

The pathogenesis of acute pancreatitis is incompletely defined, but the outcome is determined in part by an acute inflammatory process. Pancreatitis-associated inflammation appears to play a role in the local retroperitoneal injury as well as in the associated dysfunction of remote organs such as the lung. Tumor necrosis factor (TNF) appears to be a proximal mediator of the inflammatory response. In this study, anti-TNF antibody was administered to rats with caerulein-induced pancreatitis to determine if the observed increases in pancreatic and pulmonary microvascular permeability were related to plasma TNF activity. In contrast to the expected findings, blockade of TNF activity was found to increase the amount of edema formation in both the pulmonary and pancreatic microvascular beds. The mechanism is not known; however, blockade of TNF-induced down regulation of phagocytic cell activity, ablation of TNF-dependent feedback inhibition of other cytokines, failure of induction of endogenous antioxidant systems, or inactivation of the TNF control of microvascular tone are all possible explanations. This is potentially an important observation as clinical strategies are now being developed to modify the inflammatory response in ways presumed advantageous to an injured host.

Topics: Acute Disease; Animals; Antibodies; Ceruletide; Edema; Male; Pancreatic Diseases; Pancreatitis; Pulmonary Edema; Rats; Rats, Inbred Strains; Tumor Necrosis Factor-alpha

Previous studies using the isolated ex vivo perfused canine pancreatitis preparation showed that during a 4-hour perfusion pancreatitis (edema, weight gain, hyperamylasemia) can be induced by four different stimuli. The stimuli include the intra-arterial infusion of oleic acid (FFA), a 2-hour period of ischemia before perfusion (ISCH), partial obstruction of the pancreatic duct with secretin stimulation (POSS), and the intra-arterial infusion of cerulein at supramaximal doses (CER). In the present study, changes in high-energy phosphate metabolism, as determined by nuclear magnetic resonance spectroscopy, and changes in cellular structure, determined by light and electron microscopy, were documented for all four models of acute pancreatitis. The control preparations remained stable for the 4-hour perfusion period, with no decrease in adenosine triphosphate (ATP) levels. In the FFA preparations, ATP decreased to 36% of baseline levels during the 4-hour perfusion (p less than 0.001). In the ISCH preparations, ATP decreased to undetectable levels during the 2-hour period of ischemia, but recovered rapidly and remained at baseline levels during the perfusion. ATP levels remained stable in the remaining two models of pancreatitis (POSS, CER). Microscopy demonstrated that the initial injury was located chiefly in the capillaries (swollen endothelium, intravascular thrombi) in the FFA and ISCH preparations. In the POSS and CER preparations, capillary changes were minimal and the injury was located chiefly in the acinar cells (swollen endoplasmic reticulum, zymogen granule depletion, vacuolization). The POSS preparations also showed striking dilation of centroacinar lumens reflecting duct obstruction. In additional studies it was shown that the ATP decline in the FFA preparations could be significantly reduced by pretreatment with free radical scavengers. The morphologic changes could be reduced by free radical scavengers in the FFA and ISCH preparations. Any amelioration of morphologic injury in the POSS preparations was obscured by dilatation of centroacinar lumens in both treated and untreated groups. The morphologic changes in the CER preparations were reduced by treatment with a cholecystokinin inhibitor.

Topics: Acute Disease; Adenosine Triphosphate; Animals; Ceruletide; Dogs; Free Radicals; In Vitro Techniques; Magnetic Resonance Spectroscopy; Models, Biological; Oleic Acid; Oleic Acids; Pancreas; Pancreatitis; Perfusion; Secretin

Early ultrastructural and immunohistochemical changes caused by supramaximal secretory stimulation with caerulein were studied in the rat pancreas. The morphological basis for the earlier reported decrease of pancreatic juice secretion after supramaximal caerulein was the appearance of swollen and irregular zymogen-like material containing structures with short segments of budding bristle-coated membranes in the apical parts of acinar cells. Images of exocytosis of zymogen granules were only few. Later, marked vacuolization and signs of autophagocytosis are seen in the basal cytoplasm. Immunohistochemistry showed that the large zymogen containing structures were intensively labelled for trypsin at the early stages of the experiment (4-30 min). Later (1-2 h), the vacuoles were empty or contained occasional, small-labelled granules only. The pancreozymin-receptor antagonist proglumide as well as cycloleucine that inhibits protein synthesis by inhibiting the synthesis of S-adenosylmethionine, effectively prevented the caerulein induced acinar cell changes. The irregular zymogen containing structures with coated pits on their surface indicate disturbed zymogen granule formation leading to the accumulation of large lakes of zymogen material and finally to marked autophagocytosis in acinar cells. The effects of caerulein are receptor-mediated and depend on the process of methylation in the formation of zymogen granules.

Topics: Acute Disease; Animals; Ceruletide; Cycloleucine; Cytoplasm; Cytoplasmic Granules; Enzyme Precursors; Immunohistochemistry; Male; Pancreas; Pancreatitis; Proglumide; Rats; Rats, Inbred Strains; Vacuoles

Topics: Acute Disease; Animals; Ceruletide; Edema; Hemorrhage; Pancreas; Pancreatitis; Rats; Stress, Physiological

The therapeutic effect and the mechanism of action of the synthetic trypsin inhibitor camostate were studied in a rat model of acute interstitial pancreatitis induced by four subcutaneous injections of 20 micrograms/kg body weight of cerulein at hourly intervals. Rats with acute pancreatitis were given either 100 mg/kg body weight camostate or volume- and pH-adjusted water via an orogastric tube 30 min after the last cerulein injection. The elevation of serum amylase activity was significantly reduced by camostate treatment and the peak value was seen 1 hr earlier than that observed in the rats that did not receive camostate. Camostate also inhibited the reduction in pancreatic content of lipase and amylase seen during experimental pancreatitis. These effects were accompanied by alleviation of the histologic signs of acute pancreatitis such as cellular infiltration and acinar cell vacuolization. After oral administration, camostate and its metabolite were absorbed from the intestine and were detectable in plasma for more than 6 hr in concentrations high enough to have antiprotease activity. In addition, camostate in the duodenum was able to increase pancreatic juice flow and protein output and to stimulate endogenous secretin release. These results suggest that oral administration of camostate reduces the severity of cerulein-induced acute pancreatitis by releasing endogenous secretin and by its antiprotease activity.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Esters; Gabexate; Guanidines; Male; Organ Size; Pancreas; Pancreatitis; Proglumide; Rats; Rats, Inbred Strains; Secretin; Trypsin Inhibitors

There are few observations of in vivo pancreatic secretory changes that accompany acute pancreatitis. We hypothesized that acute pancreatitis impairs pancreatic exocrine function. We developed a conscious-rat experimental preparation with gastric, duodenal, bile, and pancreatic fistulas. We studied cholecystokinin-stimulated pancreatic secretion in conscious rats before and after inducing acute pancreatitis with supramaximal administration of caerulein--5 micrograms/kg/hr intravenously for 6 hours. Marked hyperamylasemia developed in all rats immediately after administration of caerulein. Basal and cholecystokinin-stimulated pancreatic juice flow and protein (enzyme) secretion decreased significantly 24 hours after acute pancreatitis was induced even though plasma amylase returned to basal levels. We conclude that acute pancreatitis markedly impairs pancreatic secretion.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Edema; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Sincalide

This study was performed to assess the effects of misoprostol, a synthetic prostaglandin E1 analog, on cerulein-induced pancreatitis. Per group of 10 each, male Wistar rats received either cerulein (2.5 micrograms/kg/h subcutaneously), cerulein and misoprostol (500 micrograms/kg intraperitoneally at 0 and 4 h), or saline. Rats were killed 6 h after the first injection. Misoprostol treatment significantly reduced interstitial edema and acinar cell lesions induced by hyperstimulation. Pancreatic amylase and chymotrypsin contents were increased by cerulein and returned towards control levels in the misoprostol-treated group. The lysosomal volume density and the pancreatic beta-D-glucuronidase activity were significantly increased after hyperstimulation. The two parameters were significantly reduced by misoprostol. A protective effect of misoprostol against lesions induced by cerulein hyperstimulation would be a consequence of a lysosomal stabilizating effect.

Topics: Acid Phosphatase; Acute Disease; Alprostadil; Amylases; Animals; Ceruletide; Chymotrypsin; Edema; Glucuronidase; Male; Microscopy, Electron; Misoprostol; Organ Size; Pancreatic Diseases; Pancreatitis; Prostaglandins E, Synthetic; Rats; Rats, Inbred Strains

Bile salts in the intestinal lumen act to inhibit the release of cholecystokinin (CCK). Recent studies have shown that CCK may play a permissive role in the development of acute pancreatitis. In this study, the amount of luminal bile salts in female Swiss Webster mice was either decreased by feeding 4% (wt/wt) cholestyramine or increased by feeding 0.5% sodium taurocholate for 1 wk. Plasma levels of CCK were stimulated by cholestyramine and inhibited by taurocholate. Then, acute pancreatitis was induced either by caerulein injections, or by feeding a choline-deficient, ethionine-supplemented (CDE) diet. Feeding of cholestyramine significantly decreased survival from 25% to 0% in the CDE pancreatitis, and increased the magnitude of elevation of serum amylase levels and the extent of pancreatic necrosis in both models of pancreatitis; CCK-receptor blockade with CR-1409 completely abolished the adverse effects of cholestyramine. In contrast, feeding of taurocholate significantly increased survival to 100% and decreased the elevation of serum amylase and pancreatic necrosis; CCK-8 antagonized these actions of taurocholate. Luminal bile salts appear to provide a physiologic protection against necrotizing pancreatitis, at least in part, both by inhibiting the release of CCK and by promoting resistance of the pancreas to CCK excessive stimulation in vivo.

Topics: Acute Disease; Amylases; Animals; Bile Acids and Salts; Ceruletide; Cholecystokinin; Cholestyramine Resin; Choline Deficiency; Feedback; Female; Mice; Pancreatitis; Taurocholic Acid

Supramaximal cerulein administration induces acute pancreatitis, which markedly impairs pancreatic secretion in conscious rats. We hypothesized that pretreatment with the potent cholecystokinin antagonist, L-364,718, improves the pancreatic secretory impairment associated with cerulein-induced acute pancreatitis. Rats were surgically prepared with gastric, duodenal, bile, and pancreatic fistulas and jugular vein catheters. On postoperative day 4, groups of rats were administered (a) L-364,718 1 mg/kg intraduodenally, (b) cerulein 5 micrograms/kg/h for 6 h intravenously, (c) L-364,718 1 mg/kg intraduodenally followed by cerulein 5 micrograms/kg/h for 6 h intravenously, and (d) safflower oil carrier intraduodenally. On postoperative day 5, we studied cholecystokinin (CCK)-stimulated pancreatic secretion. Plasma amylase was measured at the time of surgery and at the conclusion of experiments on postoperative days 4 and 5. The duodenally administered CCK antagonist had no effect, 24 h later, on CCK-evoked protein secretion and prevented the pancreatic exocrine impairment and hyperamylasemia caused by supramaximal cerulein administration. These observations suggest that cerulein-induced acute pancreatitis is mediated by a CCK-receptor mechanism.

Topics: Acute Disease; Amylases; Animals; Benzodiazepinones; Bicarbonates; Ceruletide; Cholecystokinin; Devazepide; Duodenum; Pancreas; Pancreatic Juice; Pancreatitis; Rats; Rats, Inbred Strains; Sincalide

In order to clarify the role of free radicals in the pathogenesis of acute pancreatitis, we observed the effect of a new synthetic free radical scavenger (CV-3611) on the pathological state in the models of both caerulein and CDE-diet induced acute pancreatitis in mice. In both models of acute pancreatitis, the levels of serum amylase activity were reduced significantly by the treatment of CV-3611. Pancreatic edema formation was also reduced significantly at 3.5 and 9 h after the first caerulein i.p. injection. The 4 days survival rate in CDE-diet induced pancreatitis was significantly elevated from 41.2% to 81.3% by the treatment of CV-3611. These results indicate that this synthetic scavenger, which has a long circulation half life, high affinity to biomembrane and good cell penetration ability, is effective on the development of both severe and mild pancreatitis. The main pathogenesis of both models is suggested to be radical reactions on the biomembrane which is caused by the interreaction between endothelium and neurophile in caerulein induced pancreatitis, and by the lipid peroxidation on the biomembrane of the organella in the cell in CDE-diet induced pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ascorbic Acid; Ceruletide; Endothelium; Ethionine; Female; Free Radicals; Lipid Peroxidation; Mice; Mice, Inbred BALB C; Neutrophils; Pancreatitis

Hypocalcemia and lipid abnormalities commonly occur in acute pancreatitis. Experimentally, increased plasma concentrations of free fatty acids (NEFA) can lower the serum calcium (Ca). We hypothesized that changes in blood-ionized calcium might parallel changes in NEFA concentration in pancreatitis. This hypothesis was tested in a model of severe necrotizing pancreatitis and a model of mild edematous pancreatitis. Adult male Sprague-Dawley rats (300-400 g) were randomized to receive: 100 microL sodium glycodeoxycholic acid (GDOC 34 mmol/L) infused into the pancreatic duct to produce severe necrotizing pancreatitis (Group 1); 100 microL 0.9% NaCl (NS) infused into the pancreatic duct (Group 2); Sham laparotomy (Group 3); A 6 h IV infusion of cerulein (5 mucg/kg/h) to produce mild edematous pancreatitis (Group 4); and a 6 h IV infusion of NS (Group 5). A significant time dependent decrease in blood-ionized Ca concentration, compared to normal rats, was observed in both GDOC-pancreatitis (0.836 +/- .057 vs 1.069 +/- .038 mmol/L p less than 0.001) and cerulein pancreatitis (0.988 +/- .028 vs 1.069 +/- .038 p less than 0.05), which was maximal 24 h after induction of pancreatitis. The degree of hypocalcemia correlated with the severity of pancreatitis (GDOC 0.836 +/- .057 vs cerulein 0.988 +/- .028 p less than .001). Hypocalcemia was not observed in any of the control groups. All experimental and control groups had significantly increased baseline NEFA concentrations compared with normal rats (p less than 0.001); however, no further increase in NEFA concentration occurred in conjunction with the observed time-dependent decline in ionized calcium concentrations. Although the NEFA concentrations observed in these experiments were comparable to those measured in human acute pancreatitis (exclusive of hyperlipemic pancreatitis), the time course of the changes suggests that increases in serum NEFA concentrations in experimental pancreatitis are not the primary factor mediating hypocalcemia.

Topics: Acute Disease; Amylases; Animals; Calcium; Ceruletide; Disease Models, Animal; Edema; Fatty Acids, Nonesterified; Glycodeoxycholic Acid; Hypocalcemia; Male; Pancreatic Ducts; Pancreatitis; Random Allocation; Rats; Rats, Inbred Strains

This study aimed to assess the role of oxygen free radicals in acute pancreatitis. Acute pancreatitis was induced in rats by infusion of the CCK-analogue cerulein (5 micrograms/kg per hour) for 30 minutes, 3.5 hours, and 12 hours. After the infusion, serum enzymes and conjugated tissue dienes and malondialdehyde were measured and tissue samples were subjected to electron and light microscopy. Electron microscopy after 30 minutes showed moderate intracellular alterations. After 3.5 hours of cerulein infusion interstitial oedema and intravascular margination of granulocytes in the pancreatic gland were seen. After 12 hours histological evaluation showed pronounced zymogen degranulation, extensive tissue necrosis, and migration of granulocytes into the tissue. Amylase and lipase activities increased 15 and 35-fold respectively during this time. After 30 minutes of cerulein infusion conjugated dienes and malondialdehyde increased, they reached their peak after 3.5 hours and decreased to normal values after 12 hours. Treatment with superoxide dismutase (100,000 U/kg/hour) and catalase (400,000 U/kg/hour) either before or after the start of the cerulein infusion prevented lipid peroxidation and reduced zymogen degranulation and tissue necrosis. Tissue oedema and inflammatory response, however, were not affected in any of the treated rats. Oxygen free radicals are instrumental in the development of acute pancreatitis. Even after its onset, scavenger treatment reduced the tissue damage normally observed.

Topics: Acute Disease; Amylases; Animals; Catalase; Ceruletide; Disease Models, Animal; Free Radical Scavengers; Free Radicals; Lipase; Lipid Peroxidation; Male; Malondialdehyde; Oxygen; Pancreatitis; Rats; Rats, Inbred WKY; Superoxide Dismutase

The response of pancreatic exocrine secretion to cholecystokinin (CCK), has been studied in experimental acute pancreatitis induced in rats by supramaximal doses of caerulein. Several doses of caerulein were used (4, 20 and 40 micrograms/Kg) and each one was administered by four subcutaneous injections over 3 h at hourly intervals. Pancreatic juice was collected 9 h after the first injection. The caerulein-treated animals showed a statistically significant increase in serum amylase levels. Secretory activity of ductular cells remained unchanged in all the caerulein-treated animals, but total protein and amylase secretion decreased significantly at all the caerulein doses used, both in resting conditions and under stimulation with CCK (1.25 micrograms/Kg/h). Despite this the acinar cells of rats treated with the lowest dose of caerulein retained a certain degree of secretory function since amylase activity in pancreatic juice was greater than in other groups of rats treated with higher doses of caerulein. Moreover, the percentage of increase observed in total protein and amylase in response to CCK respect to basal secretion is similar to that of the untreated animals. At higher doses (20 and 40 micrograms/Kg) the secretory capacity in response to CCK was inhibited. Therefore CCK administration in slight acute pancreatitis could be used as a therapy since it favours the secretion of pancreatic enzymes at percentual levels similar to those of the controls.

Topics: Acute Disease; Animals; Ceruletide; Cholecystokinin; Dose-Response Relationship, Drug; Male; Pancreas; Pancreatic Juice; Pancreatitis; Rats; Rats, Inbred Strains

To explore the relationship between exocrine pancreas and parotid gland, we measured the changes of parotid gland in in-vitro system at an acute pancreatitis induced by supramaximal dose of caerulein (5 micrograms/kg/h for 3.5 hours) in rats. Both the serum amylase levels and parotid gland amylase content in rats with acute pancreatitis increased significantly compared with normal rats. The dry/wet weight ratio also decreased significantly and LDH discharge from parotid acini as well as lysosomal enzyme leakage from lysosomes in acini increased significantly compared with normal rats. In addition, redistribution of lysosomal enzyme in parotid acini was seen in acute pancreatitis. These results indicate the edema and congestion of amylase in parotid gland, and furthermore the increased cellular and lysosomal fragility of parotid gland at an acute pancreatitis. Thus, there seems to be the intimate organ relationship between exocrine pancreas and parotid gland as well as the important roles of gut hormones such as caerulein in the pathophysiology of parotid gland.

Topics: Acute Disease; Amylases; Animals; Ceruletide; L-Lactate Dehydrogenase; Lysosomes; Male; Pancreatitis; Parotid Gland; Rats; Rats, Inbred Strains

The role of free radicals in the development of cerulein-induced pancreatitis was evaluated by measuring the activity of the endogenous scavengers, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSHpx), as indicators of the defense system, and the level of lipid peroxide (LPO) in the pancreas, as an indicator of the offense system. Acute pancreatitis was induced by 5 hourly intraperitoneal administrations of cerulein (50 micrograms/kg body weight), in 0.9% NaCl, to mice. The presence of acute pancreatitis was confirmed by changes in serum amylase levels and in typical microscopical features. Regarding the changes in the levels of endogenous scavengers, the SOD level was decreased significantly from a basal level of 52.6 +/- 3.94 to 43.1 +/- 2.79 mU/micrograms DNA at 6 h (p less than 0.01) to 38.8 +/- 5.18 mU/micrograms DNA at 9 h (p less than 0.05) and to 31.7 +/- 3.10 mU/micrograms DNA at 12 h (p less than 0.01) after the first intraperitoneal cerulein injection. The CAT level also decreased significantly from a basal level of 7.80 +/- 0.27 to 5.86 +/- 0.46 mU/micrograms DNA at 9 h (p less than 0.01) and to 4.52 +/- 0.21 mU/microgram DNA at 12 h (p less than 0.01). GSHpx increased from a basal level of 6.80 +/- 0.43 to 7.58 +/- 0.50 mU/micrograms DNA at 9 h and to 10.2 +/- 0.52 mU/micrograms DNA at 12 h after the first intraperitoneal cerulein injection.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Animals; Catalase; Ceruletide; Female; Free Radical Scavengers; Glutathione Peroxidase; Lipid Peroxides; Mice; Mice, Inbred BALB C; Oxygen; Pancreas; Pancreatitis; Superoxide Dismutase

Topics: Acute Disease; Animals; Ceruletide; Lysosomes; Pancreatitis; Parotid Gland; Rats

Because somatostatin is a potent inhibitor of pancreatic secretion, we hypothesized that pretreatment with somatostatin analogue octreotide (SMS 201-995) might prevent cerulein-induced edematous pancreatitis. We studied 18 rats prepared with jugular vein catheters. The following agents were administered intravenously to groups of four rats for 6 hours: 1 mL/h (control) crystalloid solution; 1-microgram/kg bolus then 1 microgram/kg per hour of octreotide; and 5 micrograms/kg per hour of cerulein; also, in a fourth group of six rats, octreotide and cerulein were administered simultaneously. At the end of experiments, blood was drawn for plasma amylase determinations; rats were killed and pancreata were examined. Supramaximal cerulein administration to conscious rats induced hyperamylasemia and edematous pancreatitis, confirming previous observations; in both groups of rats receiving cerulein, there was prominent interstitial edema, acinar vacuolization, and mild-to-moderate acute inflammation. While octreotide pretreatment of rats with cerulein-induced acute pancreatitis was associated with a lesser increase of wet pancreas weight and plasma amylase concentration, there was little overall benefit of octreotide pretreatment in this form of experimental acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Octreotide; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

Caerulein-induced acute pancreatitis is characterized by the occurrence of two membrane-bound vacuolar systems in acinar cells. Beside digestive enzymes containing secretory vacuoles, lysosomal autophagic structures can be identified at the ultrastructural level. In the present study glycoconjugate patterns of the surrounding membranes were characterized by ultrastructural lectin-binding experiments using five colloidal-gold labeled lectins with distinct sugar specificities. Furthermore, the profile of membrane glycoproteins of isolated vacuolar fractions was studied by SDS-PAGE and lectin-blotting. In pancreatitis, membranes of secretory vacuoles showed a significant lower degree of lectin-binding compared to normal zymogen granules. In contrast, newly appearing autophagic vacuoles in pancreatitis revealed a strong membrane labelling for most lectins used. The pattern of membrane glycoproteins of secretory and autophagic vacuoles as determined by SDS-PAGE and lectin-blotting differed from those of normal zymogen granules resembling the protein profile of smooth microsomes. Since this pattern requires a previous passage through Golgi stacks, it is assumed that the two types of vacuoles derive from Golgi elements. For the pathogenesis of caerulein pancreatitis these vacuolar post-Golgi structures seem to play an important role.

Topics: Acute Disease; Animals; Ceruletide; Endoplasmic Reticulum; Glycosylation; Intracellular Membranes; Lectins; Male; Membrane Glycoproteins; Membrane Proteins; Microscopy, Electron; Microsomes; Organelles; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Vacuoles

Acute edematous pancreatitis was induced in conscious rats by intravenous infusion of cerulein at a supramaximal dose of 7.5 micrograms/kg/h during 6 h. The most important finding of our study was a marked decrease in the protein and non-protein content of sulfhydryl groups parallel to an evident elevation in the malondialdehyde concentration in pancreatic tissue. The presented data suggest that in cerulein-induced acute pancreatitis in rats, oxygen radicals mediate increased peroxidation reactions which are accompanied by depletion of nonenzymatic sulfhydryl-containing free radical scavengers. The above phenomenon contributes to a disturbance in thiol metabolism resulting in serious diminution of pancreatic protein sulfhydryl compounds.

Topics: Acute Disease; Animals; Ceruletide; Free Radicals; Male; Oxygen; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Sulfhydryl Compounds

We evaluated the effects of a new cholecystokinin (CCK) receptor antagonist, loxiglumide, in a model of mild pancreatitis induced by repeated injections of cerulein and in a severe necrotizing form of pancreatitis induced by retrograde ductal injection of sodium taurocholate (NaTc) in rats. A single subcutaneous injection or oral administration of 50 mg/kg of body weight of loxiglumide almost completely reduced the increases of serum amylase activity and pancreatic wet weight, and caused histologic improvements of the cerulein-induced acute pancreatitis when given 30 min before the first cerulein injection. Loxiglumide was also effective in reducing the elevated serum amylase activity, pancreatic wet weight, and histologic alterations even when administered after the induction of acute pancreatitis. However, loxiglumide offered no apparent beneficial effects when given 30 min before and 3 h after the induction of acute pancreatitis by NaTc as determined by changes in serum amylase activity, pancreatic wet weight, and histology. These results do not necessarily suggest that CCK is not important in the pathogenesis of pancreatitis, but do suggest that the sole blockade of peripheral CCK receptors is ineffective against NaTc-induced severe necrotizing pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Cholecystokinin; Disease Models, Animal; Glutamine; Male; Pancreatitis; Proglumide; Rats; Rats, Inbred Strains; Receptors, Cholecystokinin; Taurocholic Acid

The increased incidence of gallbladder diseases after gastrectomy is discussed with regard to contractile motility of the gallbladder. Ultrasonographic findings and contraction of the gallbladder in response to egg yolk or caerulein were studied before and after gastrectomy at intervals ranging from 2 weeks to 6 months. Enlargement of the gallbladder with accumulation of biliary sludge and hypomotility were frequently observed within a month of operation for gastric cancer, suggesting that biliary stasis is an important contributing factor in postoperative acute cholecystitis. Within 3 months of operation, contraction had recovered to close to preoperative levels and the incidence of biliary sludge formation gradually decreased. Daily administration of an opiate antagonist, naloxone (0.8 mg), significantly improved gallbladder dyskinesia and decreased the incidence of biliary sludge formation within 1 month of gastrectomy.

Topics: Acute Disease; Ceruletide; Cholecystitis; Egg Yolk; Gallbladder; Gastrectomy; Humans; Middle Aged; Muscle Contraction; Naloxone; Postoperative Complications; Stomach Neoplasms

We examined the effect of fasting on the course of experimental acute pancreatitis induced in rats by four subcutaneous injections of 20 micrograms/kg body weight of cerulein at hourly intervals. Rats were either fasted from 24 hr before to 9 hr after the first cerulein injection or fed ad libitum throughout the experiment. Twenty-four hours of fasting reduced cerulein-induced increases in serum levels of amylase and anionic trypsin(ogen) to 50 and 70% of those in fed rats, respectively. Increases in pancreatic wet weight after cerulein injections were also less in fasted rats than in fed rats. Pancreatic content of trypsin was significantly decreased after a 24-hr fast, and no further changes were induced by cerulein injections. The histological signs of acute pancreatitis were greatly alleviated by fasting. However, 24 hr of fasting did not alter the sensitivity and responsiveness of the exocrine pancreas to cerulein in both in vivo and in vitro. Plasma CCK bioactivity and immunoreactive secretin concentration in 24-hr-fasted rats were significantly lower than those in fed rats. Administration of CCK receptor antagonist, loxiglumide, 12 hr prior to the induction of acute pancreatitis reduced the increase in serum amylase activity in fed rats to nearly the same levels as that in fasted rats and alleviated histological signs of pancreatitis to some extent. These present observations suggest that fasting lessens the severity of cerulein-induced acute pancreatitis by reducing endogenous CCK release.

Topics: Acute Disease; Animals; Ceruletide; Cholecystokinin; Fasting; Male; Pancreas; Pancreatitis; Proglumide; Rats; Rats, Inbred Strains; Secretin

Little is known about exocrine pancreatic secretory function in patients with acute pancreatitis, in particular during the early phase of the disease. Therefore, this study evaluates basal and stimulated pancreatic secretion in vivo and in vitro in four different models of acute pancreatitis which reflect its clinical spectrum of severity: (a) edematous pancreatitis induced in the rat by seven IP injections of 50 micrograms/kg cerulein at hourly intervals; (b) edematous pancreatitis with cellular necrosis induced in the mouse by seven IP injections of 50 micrograms/kg cerulein at hourly intervals; (c) hemorrhagic pancreatitis induced in the mouse by feeding an ethionine-supplemented, choline-deficient diet for 66 hours; and (d) hemorrhagic pancreatitis induced in the rat by retrograde infusion of 0.6 mL 5% sodium taurocholate into the pancreatic duct. Secretory studies were performed in vivo and in vitro at various times after onset of pancreatitis. The results show that the exocrine pancreas gradually became resistant to cholecystokinin stimulation after the onset of acute pancreatitis in all four animal models. Cholecystokinin-stimulated secretion was almost abolished in vivo and in vitro at the time of maximal histological damage. In vivo basal secretion was also reduced. In vitro there was an increase in basal release of amylase from isolated acini that was not caused by an increase in luminal secretion but by enzyme release from damaged cells. The time course of improvement of secretory function after acute experimental pancreatitis depended on the severity of the pancreatitis. Recovery of secretory capacity took longer after severe necrotizing pancreatitis than after edematous pancreatitis. However, the ultimate resolution of secretory function was remarkable, in particular after severe hemorrhagic pancreatitis. In all four models, secretory capacity became indistinguishable from normal before the morphological alterations had completely resolved. The present experimental data suggest that pancreatic secretion, and particularly pancreatic secretory response to cholecystokinin, may also be reduced in patients early after the onset of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Choline Deficiency; Diet; Female; Male; Mice; Pancreas; Pancreatic Juice; Pancreatitis; Rats; Rats, Inbred Strains; Sincalide; Taurocholic Acid

The effects of cholecystokinin receptor antagonist CR 1392 was studied in a model of mild acute pancreatitis induced in rats by four subcutaneous injections of the secretagogue caerulein. A single subcutaneous injection of 50 mg/kg body weight of CR 1392 caused a dramatic reduction in serum amylase concentration and pancreatic wet weight as well as histologic improvement of the caerulein-induced acute pancreatitis when given 30 min before the first caerulein injection. CR 1392 was also effective in reducing the elevated serum amylase activity, pancreatic weight, and histologic alterations even when administered after the pancreatitis had been induced. These present observations suggest that CR 1392 remains active for more than 3 h and blocks the action of caerulein on the pancreas.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Glutamine; Humans; Male; Organ Size; Pancreas; Pancreatitis; Proglumide; Rats; Rats, Inbred Strains; Receptors, Cholecystokinin

Rats infused with a supramaximally stimulating dose of the cholecystokinin (CCK) analog caerulein develop acute edematous pancreatitis. Using CCK-JMV-180, a recently developed CCK analog that acts as an agonist at high-affinity CCK receptors but antagonizes the effect of CCK at low-affinity receptors, we have determined that caerulein induces pancreatitis by interacting with low-affinity CCK receptors. Those low-affinity receptors mediate CCK-induced inhibition of digestive enzyme secretion from the pancreas. Our observations, therefore, suggest that this form of experimental pancreatitis results from the inhibition of pancreatic digestive enzyme secretion.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Kinetics; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Receptors, Cholecystokinin; Sincalide

Cerulein-induced acute pancreatitis in rats is associated with a reversible lung injury that is characterized by alveolar capillary endothelial-cell injury, increased microvascular permeability, interstitial edema formation, and intraalveolar hemorrhage and fibrin deposition. The role of mediators in this injury was analyzed using gravimetric data, microvascular permeability indices, electron microscopy, and a quantitative morphometric analysis. Neutrophil depletion induced by a specific antibody was highly protective against lung injury. Interruption of the complement pathway (using low dose Naja naja cobra venom factor) also protected against lung injury. Catalase and superoxide dismutase were also protective. The iron chelator deferoxamine and the hydroxyl radical scavenger, dimethylsulfoxide, were not protective against acute lung injury. These data suggest that complement, neutrophils, and neutrophil-derived (H2O2-dependent) oxygen products mediate lung injury that occurs secondary to cerulein-induced pancreatitis. In contrast to other models of neutrophil-dependent, oxygen-radical-mediated lung injury, this lung injury does not appear to be an iron-dependent and hydroxyl-radical mediated injury. We postulate that the process of acute pancreatitis leads to complement activation followed by neutrophil recruitment, sequestration, and adherence to alveolar capillary endothelial cells. Ultimately lung injury appears to result from local endothelial-cell injury secondary to neutrophil-generated oxygen products that may be myeloperoxidase dependent.

Topics: Acute Disease; Animals; Capillary Permeability; Ceruletide; Complement System Proteins; Drug Combinations; Endothelium, Vascular; Free Radicals; Hydrogen Peroxide; Lung Diseases; Male; Neutropenia; Neutrophils; Oxygen; Pancreatitis; Polyethylene Glycols; Pulmonary Alveoli; Rats; Superoxide Dismutase

We examined the ability of a highly potent synthetic protease inhibitor, nafamostat mesilate (FUT-175), to protect the rat pancrease against AP induced by a supramaximal dose of caerulein (CR). Rats received a 6-h, continuous intravenous (iv) infusion of either CR alone or CR + a 6-h infusion of either 2.5, 5.0, 10.0, 25.0, or 50.0 mg of FUT-175/kg/h. Pancreas weights and serum chymotrypsinogen concentrations were significantly elevated by approximately 85 and 75%, respectively, over values in saline infused rats. Pancreas weights in rats treated with CR + FUT-175 at doses from 2.5-25.0 mg/kg/h were significantly reduced by approximately 20% compared to rats given CR along, and histology showed a reduction in the extent and size of acinar cell vacuolization and reduced interstitial edema compared to rats treated with CR alone. Serum chymotrypsinogen concentrations in rats treated with CR and either 5.0 or 10.0 mg of FUT-175/kg/h were significantly lower than in rats given CR alone. Significant mortality occurred in rats infused with FUT-175 at doses of either 25.0 or 50.0 mg of FUT-175/kg/h. These data indicate that serine proteases appear to be involved in the pathogenesis of CR induced AP in rats and that FUT-175 administered in low doses (2.5-10.0 mg/kg/h) provides significant protection against this form of pancreatitis.

Topics: Acute Disease; Animals; Benzamidines; Ceruletide; Chymotrypsinogen; Guanidines; Male; Organ Size; Pancreatitis; Protease Inhibitors; Rats; Rats, Inbred Strains; Reference Values

A morphological study of the pancreatic microvasculature in experimental pancreatitis was performed in dogs, using three-dimensional scanning electron microscopy vascular casting and transmission electron microscopy. Animals with pancreatitis were characterized by a marked reduction in the number of capillaries outlined by the cast and by distortion of those that were outlined, with irregularity of the capillary lumen, abruptly terminating capillary buds and leakage of the cast material through the capillary membrane. The study provides convincing morphological evidence that one of the earliest changes in acute pancreatitis is distortion of the capillary circulation.

Topics: Acute Disease; Animals; Capillaries; Ceruletide; Dogs; Microscopy, Electron; Microscopy, Electron, Scanning; Pancreas; Pancreatitis; Polyesters; Resins, Synthetic

Previous studies have demonstrated that intravenous catalase infusion protects against the formation of pancreatic edema in cerulein-induced acute pancreatitis; however, polyethylene glycol (PEG)-conjugated catalase given as a bolus was not protective. Using radiolabeled catalase and PEG-catalase in subtherapeutic tracer doses, the pancreas tissue distributions of each were determined in rats with and without pancreatitis. Rats with cerulein-induced pancreatitis developed tissue concentrations of catalase within the pancreas that were three times those of PEG-catalase. The relatively low levels of PEG-catalase in the pancreas outside of the vascular compartment suggest that the failure to prevent edema formation may result from inability of PEG-catalase to reach extravascular sites of injury because of the large molecular size.

Topics: Acute Disease; Animals; Catalase; Ceruletide; Drug Combinations; Edema; Male; Pancreas; Pancreatic Diseases; Pancreatitis; Polyethylene Glycols; Rats; Rats, Inbred Strains

As a result of pancreas stimulation with secretin-ceruletid we were able to measure the release of phospholipase A in ascites, serum, lymph, and urine in acute experimental pancreatitis in the dog. After induction of acute pancreatitis we found no increase over the normal range in serum, lymph, and urine phospholipase A activities. In addition, the stimulation of the exocrine pancreas did not show a significant change in phospholipase A activity. The excessively high phospholipase A activity in ascites following induction of acute pancreatitis fell significantly after pancreas stimulation with secretin-ceruletid.

Topics: Acute Disease; Animals; Ascites; Ceruletide; Dogs; Lymph; Male; Pancreatic Function Tests; Pancreatitis; Phospholipases; Phospholipases A; Secretin

Oxygen derived free radicals have been implicated in the pathogenesis of acute pancreatitis in numerous animal models of the disease. The xanthine oxidase inhibitor allopurinol has been shown to attenuate pancreatic damage in canine and mouse models of acute pancreatitis presumably by preventing the generation of cytotoxic superoxide anions. We therefore examined whether allopurinol could attenuate pancreatic injury in conscious rats with caerulein induced acute pancreatitis. A continuous intravenous infusion of allopurinol (20 mg/kg/h) for six hours along with an acute pancreatitis producing dose of caerulein (10 micrograms/kg/h) reduced pancreas weights by approximately 45% and serum amylase concentrations by approximately 60% compared with rats intravenously infused with either caerulein alone or caerulein plus a lower dose (10 mg/kg/h) of allopurinol. We conclude that the generation of oxygen derived free radicals via pancreatic xanthine oxidase represents an early and perhaps pivotal mechanism in the pathogenesis of acute pancreatitis.

Topics: Acute Disease; Allopurinol; Amylases; Animals; Ceruletide; Dose-Response Relationship, Drug; Male; Organ Size; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains

We examined the protective effects of the trypsin inhibitor, urinastatin, extracted from human urine in experimental acute pancreatitis in conscious rats. Acute pancreatitis was induced by four subcutaneous injections of 20 micrograms/kg body weight of cerulein at hourly intervals. Urinastatin at a dose of 50,000 U/kg body weight/6.5 h was given by continuous i.v. infusion beginning 0.5 h before the first cerulein injection and continuing until 3 h after the last one, for a total of 6.5 h. Urinastatin significantly reduced serum levels of amylase, lipase, and anionic trypsin(ogen) but did not affect pancreatic wet weight or protein or enzyme content. Urinastatin also significantly reduced the degree of acinar cell vacuolization, interstitial edema, and cellular infiltration. These results suggest that urinastatin does not block the induction of acute pancreatitis by cerulein but does substantially reduce its severity.

Topics: Acute Disease; Amylases; Animals; Ceruletide; DNA; Glycoproteins; Lipase; Male; Organ Size; Pancreatitis; Rats; Rats, Inbred Strains; Trypsin; Trypsin Inhibitors

Topics: Acute Disease; Animals; Ceruletide; Edema; Hemorrhage; Magnetic Resonance Spectroscopy; Pancreatitis; Rats; Rats, Inbred Strains; Taurocholic Acid

The effects of a polyethylene glycol linked oxygen free radical scavenger enzyme, superoxide dismutase (PEG:SOD) on caerulein induced acute pancreatitis (AP) in rats were examined. Pancreas weights and serum amylase concentrations in rats given a three hour continuous intravenous infusion of caerulein (7.5 micrograms/kg/h, n = 18) for induction of AP followed by a three hour infusion of normal saline were significantly raised by approximately 25% (p less than 0.005) and 750% (p less than 0.001), respectively, compared with values obtained in control rats (n = 7) infused for six hours with normal saline alone. A single intraperitoneal injection of either 1 X 10(4) U/kg (n = 6), 2 X 10(4) U/kg (n = 5), or 4 X 10(4) U/kg (n = 5) of PEG:SOD immediately before caerulein infusion did not significantly alter pancreas weights, serum amylase content, or pancreatic histopathology compared with rats given caerulein alone. By contrast, a single intravenous bolus injection of 4 X 10(4) U/kg (n = 9) of PEG:SOD before caerulein treatment significantly reduced serum amylase content by approximately 25% (p less than 0.05) and a continuous six hour intravenous infusion of 4 X 10(4) U/kg/h of PEG:SOD (n = 5) produced significant reductions of approximately 25% (p less than 0.001), 35% (p less than 0.05), and 50% (p less than 0.01) in pancreas weights, serum amylase concentrations, and acinar cell vacuolisation (p less than 0.01), respectively, compared with values in rats given caerulein alone. In studies using bovine serum albumin linked to polyethylene glycol and infused for six hours at protein concentrations identical to high dose PEG:SOD (n = 6), no beneficial effects against caerulein induced AP were observed. These data suggest that (a) oxygen derived free radicals are involved in the early pathogenesis of caerulein induced AP in rats, and (b) the greatly extended circulating half life of polyethylene PEG:SOD ( > 35 hours in rats compared with less than six minutes for native superoxide dismutase) may make this compound more suitable than native superoxide dismutase as a potential therapeutic agent in AP.

Topics: Acute Disease; Animals; Ceruletide; Free Radicals; Male; Oxygen; Pancreas; Pancreatitis; Polyethylene Glycols; Rats; Rats, Inbred Strains; Superoxide Dismutase

Topics: Acute Disease; Animals; Ceruletide; Disease Models, Animal; Dose-Response Relationship, Drug; Glutamine; Glycoproteins; Kinetics; Male; Microscopy, Electron; Pancreatitis; Proglumide; Rats; Rats, Inbred Strains; Receptors, Cholecystokinin; Trypsin Inhibitors

Conscious rats were treated with a supramaximal dose of 5.10(-6)g.kg-1.h-1 of cerulein for periods of 3 and 12 h. In both groups of animals typical features of acute oedematous pancreatitis were proved by biochemical and histologic examinations. The most important finding of our study was the decrease of superoxide dismutase (SOD) activity in pancreatic tissue, accompanied by a slight increase of this scavenger enzyme in serum of rats stimulated with cerulein during 3 h. Parallelly, evident elevation of malondialdehyde (MDA) concentration in pancreatic tissue was noted. After the 12-h infusion of cerulein we were not able to detect any SOD activity in pancreatic tissue, whereas this activity appeared in ascitic fluid of tested animals. Further increase of MDA concentration in pancreatic tissue, in comparison with 3-h pancreatitis, was found. These data suggest that in 3-h and 12-h cerulein-induced pancreatitis the oxygen-derived free radicals mediate the increased lipid peroxidation in pancreatic tissue. We think that the depletion of the scavenger enzyme SOD may be responsible for such a disturbance of lipid metabolism.

Topics: Acute Disease; Animals; Ceruletide; Cytosol; Free Radicals; Male; Malondialdehyde; Pancreatitis; Rats; Rats, Inbred Strains; Superoxide Dismutase

The appearance of vacuoles inside acinar cells characterizes an early stage of development in different models of acute pancreatitis and, possibly, also in human disease. The vacuoles have been shown to contain both digestive and lysosomal enzymes. This abnormal admixture may have important implications for the pathogenesis of pancreatitis because the lysosomal enzyme cathepsin B can activate trypsinogen and may, by this way, trigger pancreatic autodigestion. For the activation process of trypsinogen by cathepsin B, however, an acidic pH is required. This study, therefore, looked for evidence of vacuole acidification in two different models of acute pancreatitis. Edematous pancreatitis was induced in rats by hyperstimulation with cerulein and hemorrhagic pancreatitis was induced in mice by feeding a choline-deficient, ethionine-supplemented diet. Pancreatic acinar cells were isolated at different times after induction of pancreatitis and incubated with 50 microM of acridine orange to identify acidic intracellular compartments. As shown in previous work, zymogen granules are the main acidic compartment of normal acinar cells; they remained acidic throughout the course of pancreatitis in both models. Vacuoles became increasingly more frequent in both models as pancreatitis progressed. Throughout development of pancreatitis, vacuoles accumulated acridine orange indicating an acidic interior. Addition of a protonophore (10 microM monensin or 5 microM carbonyl cyanide m-chlorophenylhydrazone [CCCP] or a weak base (5 mM NH4Cl) completely and rapidly abolished acridine orange fluorescence inside both zymogen granules and vacuoles providing further evidence for an acidic interior. The acidification of vacuoles seen in two different models of pancreatitis may be an important requirement for activation of trypsinogen by cathepsin B and thus for the development of acute pancreatitis.

Topics: Acute Disease; Animals; Cell Compartmentation; Ceruletide; Diet; Female; Hemorrhage; Hydrogen-Ion Concentration; Male; Mice; Organoids; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Regression Analysis; Vacuoles

This study examines the effects of cerulein-induced acute pancreatitis on the secretory response of rat pancreatic acini to carbamylcholine and concentration of acinar muscarinic receptors. Rats were injected subcutaneously every 8 hr with cerulein, 12 micrograms/kg, for two days. They were sacrificed 2 and 4 hr after the first injection, 4 hr after the second and third, and 8 hr after the sixth. By 2 hr after the first injection, carbamylcholine showed decreased potency for stimulating amylase release; decreased potency becomes maximal after the second injection. Four hours after the first injection, carbamylcholine also showed decreased efficacy for causing maximal amylase release. In the course of development of pancreatitis, progressive reductions in muscarinic receptor concentrations were evident from 4 hr after the second injection. Following the complete treatment (8 hr after the sixth injection), no alteration could be observed in the affinity or proportions of each agonist class of muscarinic receptors. These studies indicate that the pancreatic acinar cells still remain functional after acute cerulein-induced pancreatitis, although significant reductions in potency and efficacy of carbamylcholine to cause amylase release and reduced muscarinic receptor concentration occur.

Topics: Acute Disease; Amylases; Animals; Carbachol; Ceruletide; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Receptors, Muscarinic

Acute pancreatitis (AP) is believed to result from intraparenchymal activation of trypsin and other digestive enzymes within the pancreas followed by autodigestion of the gland. Gabexate mesilate (FOY), a synthetic guanidino acid ester exhibiting potent and versatile inhibitory actions on a number of proteinases (e.g., trypsin, kallikrein, C1-r, C1 esterase, plasmin, thrombin, phospholipase A2), was examined for its ability to protect the rat pancreas against development of AP induced by pharmacological doses of ceruletide (CRT). Rats were i.v. infused for 6 h with either CRT (5 micrograms/kg/h) or CRT + FOY (50 mg/kg/h). In FOY-treated rats the serum amylase and trypsinogen concentrations were reduced by 60 and 80%, respectively, compared to rats infused with CRT alone. Histologically, the extent of acinar cell vacuolization in the pancreas was significantly reduced and interstitial edema, although not assessed by quantitative morphometric techniques, appeared to be qualitatively lessened in the FOY-treated rats. The ability of FOY to inhibit significantly AP produced by supramaximal doses of CRT, coupled with its inhibitory properties on components of the coagulation and complement cascades, stress the importance of continued research on this compound as a potential therapeutic agent for treatment of AP and its systemic sequelae.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Gabexate; Guanidines; Infusions, Intravenous; Male; Pancreatitis; Protease Inhibitors; Rats; Rats, Inbred Strains; Trypsinogen

The development of acute pancreatitis involves a number of pathophysiological changes which result in pancreatic tissue damage. Data from several models of acute pancreatitis suggest that the in vivo conversion of the enzyme xanthine dehydrogenase to xanthine oxidase may cause tissue damage by the subsequent generation of oxygen-derived free radical products. In the present studies, acute pancreatitis was induced in mice by the administration of supramaximal secretory doses of caerulein, a cholecystokinin analogue. Pancreatic xanthine oxidase activity was observed to occur in the dehydrogenase form in both control and treated mice. Artifactual conversion to the oxidase form could be induced by exclusion of 2-mercaptoethanol and phenylmethylsulfonyl fluoride from the buffer during tissue preparation. These data indicate that no significant conversion of xanthine dehydrogenase to oxidase is associated with this model of acute pancreatitis in mice.

Topics: Acute Disease; Animals; Ceruletide; Female; Mice; Mice, Inbred ICR; Pancreas; Pancreatitis; Xanthine Dehydrogenase; Xanthine Oxidase

Topics: Acute Disease; Allopurinol; Animals; Ceruletide; Edema; Esters; Gabexate; Guanidines; Male; Pancreatitis; Rats; Rats, Inbred Strains; Trypsin Inhibitors

Alterations in the pulmonary surfactant system are partly responsible for the respiratory insufficiency seen with acute pancreatitis. In this model of cerulein-induced pancreatitis in rats, we utilized a new stable isotope metabolic tracer technique to examine one aspect of the pulmonary surfactant system and its relationship to associated lung injury. We have demonstrated primary, early depression of lung phospholipid synthesis reflected in both lung tissue and alveolar washings. We suggest that this quantitative change in pulmonary surfactant synthetic rate may partly explain the occurrence of respiratory failure with acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Lung Diseases; Male; Pancreatitis; Phosphatidylcholines; Phospholipids; Pulmonary Surfactants; Rats; Rats, Inbred Strains

The mechanism of cerulein-induced acute pancreatitis may involve the production of free radicals in excess of the capacity of endogenous intracellular scavengers. These radicals destroy the cellular membranes, releasing digestive enzymes and cellular proteins into the interstitium. Thereafter, a cascade of events, including polymorphonuclear infiltration and complement activation, leads to pancreatic destruction. The present study demonstrates that superoxide dismutase and catalase reduce the ultrastructural and biochemical injury associated with cerulein-induced acute pancreatitis in rats. Pretreatment with superoxide dismutase and catalase 30 minutes before injury did not appear to be protective, presumably because the half-life of intravenous superoxide dismutase is only 6 minutes. This and similar studies suggest a potential clinical role for free radical scavengers in acute established pancreatitis.

Topics: Acute Disease; Animals; Catalase; Ceruletide; Disease Models, Animal; DNA; Free Radicals; Humans; In Vitro Techniques; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; RNA; Superoxide Dismutase

Pharmacological doses of ceruletide administered intravenously to unconscious rats uniformly induces acute pancreatitis (AP) as well as a striking reduction in pure pancreatic juice (PPJ) and protein output. High-dose intravenous secretin administered to rats with ceruletide-induced AP effects a reestablishment of PPJ flow and a significant increase in PPJ protein output. Light microscopy of the pancreas in ceruletide-induced AP rats revealed marked acinar cell vacuolization and intense interstitial edema. By contrast, pancreatic histology in AP rats treated with high-dose secretin revealed a distinct lessening of acinar cell vacuolization and interstitial edema. We have established that high-dose intravenous secretin given to rats with ceruletide-induced AP is (1) not harmful, (2) reestablishes PPJ flow and evokes a partial restoration of protein output, and (3) appears to reduce pancreatic histopathology when compared to non-secretin-treated rats with AP.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Disease Models, Animal; Infusions, Parenteral; Male; Pancreas; Pancreatic Juice; Pancreatitis; Proteins; Rats; Rats, Inbred Strains; Secretin; Time Factors

The regenerative capacity of the different cell types in the rat exocrine pancreas has been studied in a model of hormone-induced acute pancreatitis in which pancreatic edema, inflammation, and acinar cell destruction were induced within 12 h of infusion of supramaximal concentrations of cerulein (5 micrograms/kg/h). A sequential biochemical and structural analysis of the pancreas in daily intervals was combined with the autoradiographic quantitation of labeling indices of five cell populations following 3H-thymidine injection at days 1-7 after induction of pancreatitis. Desquamation of acinar cell apical cytoplasm and release of cytoplasmic segments into the acinar lumen on the first day following induction of pancreatitis led to formation of duct-like tubular complexes. Enzyme content in the pancreas decreased progressively following the formation of the edema to levels 15-20% of controls and remained reduced during the initial 5 days. Thymidine incorporation into total DNA showed a biphasic pattern with a distinct peak at day 1 and a second broader peak between days 4 and 7. Autoradiographic quantitation of labeling indices demonstrated the exclusive incorporation into intercalated duct cells and interstitial cells during the initial 24 h, while the second peak was predominantly due to labeling of acinar cells. Larger interlobular ducts and islets did not show changes in labeling index. In vivo labeling with 3H-thymidine during the first day and analysis of labeling indices 14 days later showed the persistence of label in intercalated duct cells and interstitial cells and argued against the stem cell hypothesis and against transformation of duct cells into acinar cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acute Disease; Amylases; Animals; Autoradiography; Ceruletide; Histocytochemistry; Male; Mitosis; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Regeneration; Thymidine; Tritium

Topics: Acute Disease; Animals; Ceruletide; Choline Deficiency; Disease Models, Animal; Duodenum; Enzymes; Ethionine; Humans; Lysosomes; Pancreas; Pancreatic Ducts; Pancreatitis; Trypsin; Trypsinogen

Large pharmacological doses of ceruletide administered to conscious dogs by intravenous (i.v.) infusion uniformly induce a severe acute necrotizing pancreatitis within 4 h. High-dose i.v. secretin administered for a period of 24 h after cessation of ceruletide infusion resulted in a significant amelioration of the acute pancreatitis compared to non-secretin-treated dogs with acute pancreatitis. Light microscopy of the pancreas in secretin-treated dogs revealed a significant decrease in edema, polymorphonuclear leukocyte infiltration, cell necrosis and acinar cell vacuolization. Serum amylase levels in secretin-treated dogs were significantly decreased compared to non-secretin-treated dogs. The results of this study suggest that high-dose i.v. secretin exerts a beneficial effect on pre-established, ceruletide-induced acute pancreatitis in dogs.

Topics: Acute Disease; Animals; Ceruletide; Dogs; Female; Male; Pancreas; Pancreatitis; Secretin

Topics: Acute Disease; Animals; Ceruletide; Injections, Subcutaneous; Male; Pancreatitis; Rats; Rats, Inbred Strains

The onset, course, and regression of the biochemical and structural alterations associated with pancreatitis induced by various doses of caerulein were studied in the mouse. In addition, the protective effect of secretin was compared with that of the cholecystokinin-receptor antagonists proglumide and benzotript. Subcutaneous or intraperitoneal injections of caerulein induced increases in serum amylase concentration and pancreatic weight and histologic evidence of acute pancreatitis, all effects being dose-related. Cytoplasmic vacuoles were the earliest histologic alterations. As the pancreatitis progressed these vacuoles increased to an enormous size. Interstitial inflammation and acinar cell necrosis were prominent after 6 h, reached a maximum after 12 h, and mostly disappeared after 4 days. During the course of pancreatitis approximately 40% of the acinar cells showed signs of severe degeneration or necrosis at the most effective doses of caerulein. Electron microscopy showed both intact and degenerating granules inside the vacuoles. Signs of basolateral exocytosis of zymogen granules were not observed. During the regression of pancreatitis, focal atrophy was a remarkable histologic finding. Repetitive initiation of pancreatitis (six courses of caerulein injections over 5 wk) produced marked focal atrophy and early fibrosis. High doses of proglumide or benzotript markedly ameliorated both the biochemical and structural alterations induced by caerulein. Secretin, even at very high doses, had only minor protective effects. This study presents a model of acute necrotizing pancreatitis in which the severity of the induced pancreatitis ranges dose-dependently from mild interstitial inflammation to severe necrosis. The ultrastructural alterations described herein support the hypothesis that the trigger mechanism of acute pancreatitis appears to be a primary intracellular event rather than an interstitial event that secondarily damages the acinar cells.

Topics: Acute Disease; Animals; Benzamides; Ceruletide; Dose-Response Relationship, Drug; Glutamine; Male; Mice; Necrosis; Pancreas; Pancreatitis; Proglumide; Receptors, Cell Surface; Receptors, Cholecystokinin; Secretin; Time Factors

The functional activity of the sphincter of Oddi complex has been examined by ceruletide manometry in patients undergoing cholecystectomy with a normal peroperative cholangiogram. In Group I (n = 14), which included patients with previous acute cholecystitis/biliary colic, the sphincter activity appeared to be normal and responded to intravenous ceruletide by a marked relaxation with a significant fall in both the infusion and postinfusion pressures. In patients undergoing cholecystectomy for gallstone-associated pancreatitis (n = 8), the sphincter exhibited manometric features of hypotonia with low infusion and postinfusion pressures which were not significantly altered by intravenous ceruletide.

Topics: Acute Disease; Adult; Aged; Ampulla of Vater; Ceruletide; Cholecystectomy; Cholecystitis; Cholelithiasis; Colic; Common Bile Duct; Humans; Manometry; Middle Aged; Pancreatitis; Sphincter of Oddi

This long-term follow-up of 27 patients treated with conservative surgery for necrohemorrhagic pancreatitis (NHP) showed that an almost complete recovery of the exocrine function is achieved within 4 years after discharge, while about half of the patients presented still abnormal endocrine function. The morphological sequelae, pointed out by endoscopic retrograde pancreatography in almost 50% of the cases, remained unchanged during the follow-up period. Therefore, these data seem to exclude an evolution of NHP towards chronic pancreatitis.

Topics: Acute Disease; Alcoholism; Ceruletide; Female; Follow-Up Studies; Gallstones; Glucose Tolerance Test; Hemorrhage; Humans; Male; Necrosis; Pancreas; Pancreatic Function Tests; Pancreatitis; Postoperative Period; Radiography; Secretin

Rats infused with a supramaximally stimulating dose of the cholecystokinin-pancreozymin analogue caerulein develop acute interstitial pancreatitis (M. Lampel and H.F. Kern. Virchows Arch. A 373: 97-117, 1977). We have studied the early (30-180 min) morphological changes in pancreatic acinar cells induced by infusing caerulein (2.5 micrograms X kg-1 X h-1). The techniques of thin-section electron microscopy, freeze fracture, and enzyme and immunocytochemistry were employed. Shortly (30 min) after the onset of caerulein infusion, large vacuoles appeared in the Golgi area. After longer periods of infusion, these vacuoles further enlarged (probably by fusion with other such vacuoles as well as autophagic vacuoles) and became more widely distributed in the cytoplasm. These large vacuoles were found to be acid phosphatase positive and to be labeled by antibodies directed against digestive zymogens as well as the lysosomal enzyme cathepsin D. These observations indicate that the large vacuoles contain both digestive zymogens and lysosomal hydrolases. During caerulein infusion, morphological evidence of exocytosis at the luminal plasmalemma was reduced or absent, and evidence of basolateral exocytosis was not noted. These studies suggest that secretagogue hyperstimulation with caerulein interferes with the processes involved in condensing vacuole maturation, which normally lead to the separation of digestive zymogens and lysosomal hydrolases. As a result, both types of enzymes remain within the same compartment. This may lead to the intracellular activation of digestive enzymes by lysosomal hydrolases and be an important step in the development of acute pancreatitis.

Topics: Acute Disease; Animals; Ceruletide; Cytoplasmic Granules; Disease Models, Animal; Freeze Fracturing; Golgi Apparatus; Lysosomes; Male; Microscopy, Electron; Pancreatitis; Rats

Unconscious rats given intravenous ceruletide (diethylamine salt of the decapeptide caerulein) in large pharmacologic doses consistently developed moderate acute pancreatitis by 3 h and florid pancreatitis by 6 h. Biochemical serum markers of acute pancreatitis tended to parallel the severity of the pancreatic damage. In 50% of the rats, mesenteric fat necrosis was present, free peritoneal fluid containing massive elevations of trypsinogen and amylase were noted in most animals. Intravenous secretion at a low dose given simultaneously with ceruletide exerted a variable protective effect on the pathological process. A high dose of secretin produced a striking macroscopic, microscopic, and biochemical protective effect on ceruletide-induced pancreatitis. High resolution light microscopy and electron microscopy showed a marked cellular disorganization in the acini of animals treated with ceruletide alone. By contrast, there was a striking apical redirection of zymogen granules in acini of the animals treated with secretin. The results of this study suggest that high dose intravenous secretin may exert a beneficial effect on acute pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Edema; Fat Necrosis; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Secretin; Trypsinogen

The effects of hormonal or cholinergic stimulation on survival and on activities of lysosomal enzymes and amylase in pancreatic tissue and ascites were studied in rats with induced pancreatitis. Pancreatitis per se caused an increase of the activities of cathepsin D, N-acetyl-beta-D-glucosaminidase and amylase, and a decrease of acid phosphatase in pancreatic tissue. Pancreatic protein concentration was not influenced. In pancreatitic rats administration of cerulein or carbachol markedly decreased survival rate. Cerulein increased the activities of cathepsin D and amylase in ascites and cathepsin D and acid phosphatase in pancreatic tissue. Carbachol increased the activities of cathepsin D and amylase in ascites and acid phosphatase in pancreatic tissue. Both cerulein and carbachol decreased the activity of amylase in pancreatic tissue. Administration of secretin or the anticholinergic drug Pro-Banthine did not influence survival rate or the activities of lysosomal enzymes and amylase in ascites. In pancreatic tissue the activity of acid phosphatase was slightly increased by secretin or Pro-Banthine. In conclusion, the results show a nonparallel alteration of lysosomal enzyme activities in pancreatic tissue in rats with pancreatitis. Cerulein and cholinergic stimulation decreased survival rate and brought about a marked increase of cathepsin D activity in ascites and, in the case of cerulein, also in pancreatic tissue. The implication of lysosomes and especially the catheptic proteases in the pathogenesis and outcome of acute pancreatitis deserves further attention.

Topics: Acetylglucosaminidase; Acid Phosphatase; Acute Disease; Amylases; Animals; Ascitic Fluid; Carbachol; Cathepsin D; Cathepsins; Ceruletide; Hexosaminidases; Lysosomes; Male; Pancreas; Pancreatitis; Propantheline; Rats; Rats, Inbred Strains; Secretin

The influence of exogenous administration and endogenous release of certain g.i. hormones on the course of acute experimental pancreatitis was studied. Administration of 2 g of a pellet diet every eight hours decreased survival, as did repeated s.c. administration of the cholecystokinin-analogue caerulein. Also oral administration of a trypsin inhibitor--releasing intestinal factors or hormones stimulating pancreatic enzyme synthesis and secretion--decreased survival. On the other hand repeated s.c. administration of secretin or an anticholinergic drug (Pro-Banthine), or oral administration of 0.1 N HCl every eight hours did not influence survival. At blind macroscopic evaluation, caerulein was found to cause signs of more severe disease. All pancreatic rats had increased S-amylase levels, but there was no difference between any of the groups. In peritoneal fluid, however, caerulein caused an increase in the amylase activity. The results suggest that elevated S-levels of g.i. hormones, which primarily stimulate pancreatic enzyme synthesis and secretion, are harmful in acute experimental pancreatitis.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Cholecystokinin; Gastrointestinal Hormones; Male; Pancreatitis; Propantheline; Rats; Rats, Inbred Strains; Secretin; Trypsin Inhibitors

Infusion of supramaximal doses of cerulein induces acute edematous pancreatitis in the rat. Cannulation of the main pancreatic duct does not prevent the formation of the edema but reveals an almost complete reduction of pancreatic flow. Using freeze-fracture techniques and thin-section electron microscopy, earliest structural alterations were observed at membranes of zymogen granules and the plasma membrane. Fusion of zymogen granules among each other leads to formation of large membrane-bound vacuoles within the cytoplasm. These and individual zymogen granules fuse with the basolateral plasma membrane, discharging their content into the interstitial space. The findings indicate severe changes in the specificity of the intracellular membrane fusion process induced by supramaximal doses of a pancreatic secretagogue, which finally result in autodigestion of the pancreas.

Topics: Acute Disease; Animals; Cell Membrane; Ceruletide; Cytoplasm; Freeze Fracturing; Male; Pancreatitis; Rats; Rats, Inbred Strains

The influence of hormonal stimulation by caerulein administration on acute experimental pancreatitis was investigated in the rat. An experimental model of moderate acute pancreatitis was selected after injecting buffer solution containing sodium taurodeoxycholate and various concentrations of trypsin into the bile-pancreatic duct. During acute experimental pancreatitis caerulein administration increased the mortality rate, the incidence of ascites and the activity of amylase in ascites. Caerulein rendered the pancreatitis more severe also as judged from blind macroscopic evaluation. Amylase and insulin levels in serum and plasma were elevated 6 and 25 h after induction of pancreatitis irrespective of caerulein administration. In pancreatitic rats caerulein decreased the activities of digestive enzymes in pancreatic tissue. The results show that hormonal stimulation by the cholecystokinin-pancreozymin analogue caerulein during acute experimental pancreatitis is harmful.

Topics: Acute Disease; Amylases; Animals; Ceruletide; Insulin; Lipase; Male; Pancreas; Pancreatitis; Rats; Rats, Inbred Strains; Stimulation, Chemical; Taurodeoxycholic Acid; Trypsin

Rat exocrine pancreatic function was studied structurally and biochemically after the in vivo production of actue interstitial pancreatitis by supramaximal stimulation with caerulein. Two major phases in the reaction of the gland were observed: During the first two days after cessation of the supramaximal stimulation a progressive infiltration of the interstitium and the pancreatic tissue with polymorphonuclear leucocytes, lymphocytes and macrophages occurred which led to further destruction of the gland and to decreased functional response. From two days after the cessation of the treatment, hypertrophy of centro-acinar cells and an increased rate of mitotic activity indicated regeneration of the pancreas. This was combined with an accelerated in vitro discharge of newly synthesized proteins over a period of four days. Between days three and six after the initial treatment mitotic activity was also observed in fully differentiated exocrine cells. Total structural and functional recovery of the pancreas was achieved nine to tweleve days after the cessation of the supramaximal stimulation.

Topics: Acute Disease; Animals; Ceruletide; Hypertrophy; Leukocytes; Lymphocytes; Macrophages; Male; Mitosis; Pancreas; Pancreatitis; Rats; Regeneration; Remission, Spontaneous; Time Factors

Topics: Acute Disease; Animals; Ceruletide; Choline Deficiency; Disease Models, Animal; Female; Mice; Pancreas; Pancreatitis

Conscious rats were infused via a jugular vein catheter with 5 x 10-6 g/kg/h caerulein for periods up to 24 h. On macroscopic inspection a progressive interstitial oedema is seen to develop in the pancreas, from one hour of infusion on and is most marked at twelve hours. This oedema is largely reabsorbed after 24 h treatment, but the pancreas is considerably indurated by this time. Serum amylase levels increase consistently to reach a tenfold elevation above controls after three, six or twelve hours infusion. Premature fusion of condensing vacuoles and secretory granules leads to formation of large vacuoles in the cytoplasm of exocrine pancreatic cells. These vacuoles fuse with the lateral and basal plasma membrane and realease their content into the extracellular space. Regular discharge of zymogen granules at the cell apex into the duct system does not occur. Vacuole formation is associated with cytoplasmic destruction of the pancreatic cells. The rate of protein synthesis decreases consistently as a result of these structural alterations and this change corresponds largely to a reduction of cellular respiration. Release of amylase from isolated pancreatic lobules of caerulein infused animals shows a progressive increase of unstimulated discharge, while in vitro stimulation with 5 x 10-6M carbamylcholine gives secretion patterns of wash-out kinetics. Stimulated discharge of labeled secretory proteins indicates a progressive reduction in the in vitro sensitivity of the pancreatic cells to secretagogues. After 24 h infusion of 5 x 10-6 g/kg/h caerulein the pancreatic lobules are totally insensitive to the in vitro effect of carbamylcholine or caerulein.

Topics: Acute Disease; Amylases; Animals; Carbachol; Ceruletide; Edema; Male; Microscopy, Electron; Pancreas; Pancreatitis; Proteins; Rats; Secretory Rate

Serum lipase activity, which is a more specific indicator of pancreatic damage than serum amylase activity, was determined in 51 patients before and immediately after endoscopic retrograde cholangio-pancreatography (ERCP) and 24 hours subsequently. An increase in serum lipase activity without clinical signs of acute pancreatitis was observed immediately after pancreatography in 20 cases. Increased serum lipase activity after 24 hours is largely observed in patients with parenchymography or those displaying increased values already in the fasting state before ERCP. This observation emphasizes the need for caution in the assessment of the indication for ERCP in patients with preexisting increased serum lipase activity in view of the very real risk of acute pancreatitis and the need to avoid parenchymography of the pancreas. According to our experience the addition of a broadspectrum antibiotic such as gentamycin to the contrast medium may, to a large extent, avoid the development of acute pancreatitis. Lipase evocation tests carried out 10 days after the ERCP showed negative results in patients with increased enzyme activity and parenchymography after pancreatography. Hence, it is concluded that ERCP causes symptomless reversible biochemical pancreatic changes, as opposed to irreversible pancreatic damage.

Topics: Acute Disease; Ceruletide; Cholangiography; Chronic Disease; Clinical Enzyme Tests; Endoscopy; Humans; Lipase; Pancreatitis; Stimulation, Chemical