glycodeoxycholic-acid and Pancreatitis

Acute pancreatitis exhibits a rapid clinical progression which makes it difficult to observe in human; hence, an experimental animal model is needed. This preliminary study performed an induction of acute pancreatitis using glycodeoxycholic acid (GDOC) in an experimental macaque model.. GDOC injections (initial dose of 11.20 mg/kg) were administered in an escalating manner at specific time points. The injection was given along the bilio-pancreatic duct, followed by measurement of vital signs, serum amylase-lipase, TNF-α, procalcitonin, oxidative stress parameters, and microscopic and macroscopic findings.. The results indicated that acute pancreatitis occurred following induction with low-dose GDOC. Serum amylase and lipase levels increased with subsequent GDOC injections. Blood pressure and heart rate were elevated, indicating abdominal pain. Changes in TNF-α, procalcitonin, and oxidative stress values showed active inflammation. We observed histologic features of pancreatitis and as the dose increased, vasodilation of the splanchnic vasculatures was observed.. Small dose GDOC injection in the bilio-pancreatic duct may have a role to induce acute pancreatitis in Macaca nemestrina.

Topics: Acute Disease; Amylases; Animals; Glycodeoxycholic Acid; Lipase; Macaca nemestrina; Pancreatitis; Procalcitonin; Tumor Necrosis Factor-alpha

Acute lung injury (ALI) is one of the most common extra-pancreatic complications of acute pancreatitis. In this study, we examined the protective effect of protease inhibitor aprotinin and a matrix metalloproteinase inhibitor (MMPi) on pulmonary inflammation in rats with severe pancreatitis-associated ALI.. A rat model of acute pancreatitis (AP) was established by injecting sodium glycodeoxycholate (GDOC) into the pancreatic duct. Pharmacological interventions included pretreatment with a protease inhibitor aprotinin (10mg/kg) and a matrix metalloproteinase inhibitor (MMPi, 100g/kg). The extent of pancreatic and lung injury and systemic inflammation was assessed by examinations of blood, bronchoalveolar lavage (BAL), and lung tissue. Pancreatic or lung tissue edema was evaluated by tissue water content. Pulmonary arterial pressure and alveolar-capillary membrane permeability were evaluated post-injury via a catheter inserted into the pulmonary artery in an isolated, perfused lung model.. Pre-treatment with aprotinin or MMPi significantly decreased amylase and lactate dehydrogenase (LDH), and the wet/dry weight ratio of the lung and pancreas in AP rats. Compared to the GDOC alone group, administration of aprotinin or MMPi prevented pancreatitis-induced IL-6 increases in the lung. Similarly, treatment with aprotinin or MMPi significantly decreased the accumulation of white blood cells, oxygen radicals, nitrite/nitrates in both blood and BAL, and markedly reduced lung permeability.. Pretreatment with either aprotinin or MMPi attenuated the systemic inflammation and reduced the severity of lung and pancreas injuries. In short, our study demonstrated that inhibition of protease may be therapeutic to pulmonary inflammation in this GDOC-induced AP model.

Topics: Acute Lung Injury; Animals; Aprotinin; Cells, Cultured; Disease Models, Animal; Drug Therapy, Combination; Edema; Glycodeoxycholic Acid; Humans; Inflammation; Lung; Male; Matrix Metalloproteinase Inhibitors; Organ Culture Techniques; Pancreatitis; Pulmonary Artery; Rats; Rats, Sprague-Dawley

Severe acute pancreatitis is an inflammatory disease of the pancreas with a high morbidity and mortality. To date, no causal treatment is known. The aim of the present study was to analyze the impact of pancreatic microcirculatory disturbances in severe acute pancreatitis and to correlate the effects with histopathologic tissue damage and outcome.. Severe acute pancreatitis was induced in 129 pigs by injection of glycodeoxycholic acid into the pancreatic duct. Pancreatic microcirculation, pancreatic tissue oxygenation, histopathologic tissue damage, and survival were measured and analyzed.. Our study demonstrates a strong correlation between pancreatic microcirculatory disturbances and histopathologic tissue damage (r = 0.728; P < 0.001). Furthermore, we showed a strong correlation between tissue oxygenation and the severity of the pancreatitis according to an established porcine pancreatitis score (r = 0.694; P < 0.001). In addition, disturbances of the pancreatic microcirculation were shown to be associated with an increased mortality rate in severe acute pancreatitis.. We found that pancreatic microcirculatory disturbances have significant effects on histopathologic tissue damage and the outcome of severe acute pancreatitis. For a better survival of severe acute pancreatitis, the treatment should focus on an improvement of pancreatic microcirculation.

Topics: Acute Disease; Animals; Glycodeoxycholic Acid; Microcirculation; Oxygen; Pancreas; Pancreatitis; Severity of Illness Index; Survival Analysis; Swine

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

Platelet-activating factor (PAF) is an important mediator of inflammation and postulated to be involved in the pathogenesis of acute pancreatitis. In this study, we evaluated the therapeutic effect of PAF antagonist WEB 2086 in acute experimental pancreatitis of graded severity in rats.. According to a block design, 64 animals were randomly allocated to 8 groups. Severe necrotizing pancreatitis was induced by intraductal infusion of taurocholic acid (4%, 0.4 mL), and the combination of glycodeoxycholic acid (10 mmol/L, 1.0 mL/kg, intraductal infusion) and cerulein (5 microg/kg per hour, intravenous) was applied to induce intermediate pancreatitis, or cerulein alone (5 microg/kg per hour, intravenous) to establish edematous pancreatitis. WEB 2086 was given 15 minutes after beginning the induction of pancreatitis. Pancreatic microcirculation was analyzed in vivo with an epiluminescent microscope. Histopathology was evaluated by a validated score. Trypsinogen-activating peptide and serum amylase were analyzed sequentially.. WEB 2086 had no significant influence on the breakdown of microcirculation, leukocyte adherence, histopathological damage, and amylase levels in severe necrotizing pancreatitis, intermediate pancreatitis, and edematous pancreatitis. Only in intermediate pancreatitis was there a significant reduction of trypsinogen-activating peptide levels.. In our study, PAF antagonist WEB 2086 had no beneficial effect on microcirculation in acute experimental pancreatitis.

Topics: Amylases; Animals; Azepines; Capillaries; Cell Adhesion; Ceruletide; Disease Models, Animal; Edema; Female; Glycodeoxycholic Acid; Leukocytes; Microcirculation; Oligopeptides; Pancreas; Pancreatitis; Pancreatitis, Acute Necrotizing; Platelet Activating Factor; Platelet Aggregation Inhibitors; Rats; Rats, Wistar; Regional Blood Flow; Severity of Illness Index; Taurocholic Acid; Time Factors; Triazoles

Factors determining severity of acute pancreatitis (AP) are poorly understood. Oxidative stress causes acinar cell injury and contributes to the severity, whereas prophylactic probiotics ameliorate experimental pancreatitis. Our objective was to study how probiotics affect oxidative stress, inflammation, and acinar cell injury during the early phase of AP. Fifty-three male Sprague-Dawley rats were randomly allocated into groups: 1) control, 2) sham procedure, 3) AP with no treatment, 4) AP with probiotics, and 5) AP with placebo. AP was induced under general anesthesia by intraductal glycodeoxycholate infusion (15 mM) and intravenous cerulein (5 microg.kg(-1).h(-1), for 6 h). Daily probiotics or placebo were administered intragastrically, starting 5 days prior to AP. After cerulein infusion, pancreas samples were collected for analysis including lipid peroxidation, glutathione, glutamate-cysteine-ligase activity, histological grading of pancreatic injury, and NF-kappaB activation. The severity of pancreatic injury correlated to oxidative damage (r = 0.9) and was ameliorated by probiotics (1.5 vs. placebo 5.5; P = 0.014). AP-induced NF-kappaB activation was reduced by probiotics (0.20 vs. placebo 0.53 OD(450nm)/mg nuclear protein; P < 0.001). Probiotics attenuated AP-induced lipid peroxidation (0.25 vs. placebo 0.51 pmol malondialdehyde/mg protein; P < 0.001). Not only was AP-induced glutathione depletion prevented (8.81 vs. placebo 4.1 micromol/mg protein, P < 0.001), probiotic pretreatment even increased glutathione compared with sham rats (8.81 vs. sham 6.18 miccromol/mg protein, P < 0.001). Biosynthesis of glutathione (glutamate-cysteine-ligase activity) was enhanced in probiotic-pretreated animals. Probiotics enhanced the biosynthesis of glutathione, which may have reduced activation of inflammation and acinar cell injury and ameliorated experimental AP, via a reduction in oxidative stress.

Topics: Animals; Apoptosis; Ceruletide; Glutathione; Glycodeoxycholic Acid; Male; Oxidative Stress; Pancreatitis; Probiotics; Rats; Rats, Sprague-Dawley; Specific Pathogen-Free Organisms

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

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

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

The role of nitric oxide (NO) in microcirculation during the development of acute pancreatitis was not clear. An in vivo microscopic technique was used for evaluating leukocyte-endothelial adherence in the pancreatic microcirculation after induction (cerulein) of acute pancreatitis. Microdialysis was performed to detect pancreatic nitrate concentration (NO level) by high-performance liquid chromatography. Cerulein caused significantly reduced flow velocity in 1 h (31 %) and increased the number of sticking leukocytes in 2 h; both persisted for at least 3 h. Pancreatic NO level was found to be significantly elevated (2.5-fold) in 1 h and also persisted for 3 h. Both microcirculatory changes and NO elevation were significantly alleviated in cerulein-induced animals pretreated with NO synthase inhibitor (NG-nitro-L-arginine), indicating that elevation of NO could precede and account for a major portion of the observed microcirculatory changes. Furthermore, there was a strong positive correlation between numbers of adherent leukocytes and pancreatic NO level, suggesting that during the development of acute pancreatitis, NO could play an adverse role in microcirculation.

Topics: Animals; Ceruletide; Endothelium, Vascular; Enzyme Inhibitors; Glycodeoxycholic Acid; Leukocytes; Male; Microcirculation; NG-Nitroarginine Methyl Ester; Nitrates; Nitric Oxide; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley

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

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

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

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

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

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

Since some authors have stated a certain role for so-called "free fatty acids" (FFA) in the pathogenesis of AP and the subsequent systemic complications we tried to find possible correlations between FFA, pancreatitis and lung injury using a rat model. AP was induced by intraductal infusion of two different concentrations of glycodeoxycholic acid (GDOC 17 mmol and 34 mmol). An equal number of animals had only cannulation of the pancreatic duct without infusion and served as controls (GDOC-control). In another experimental model iv.-infusion of oleic acid (OA) was used to create severe lung injury comparable to human ARDS. In this model control animals received iv.-infusion of saline solution only (SAL). At 2, 6, 12, 24 and 48 hours the animals were sacrificed and blood was collected for determination of FFA, amylase and pO2. The pancreas and lungs were removed for histologic examination and the lungs were weighed. GDOC-34 animals developed severe pancreatitis with hemorrhage and necrosis. Histology of the lungs showed edema, inflammatory infiltrates, hemorrhage and thickening of the alveolar membrane in GDOC-34 rats as well as in OA-animals. In contrast, there was only pancreatic edema until 24 hours in the GDOC 17 group and less severe histological changes in the lungs. Amylase, FFA, pO2 and lung weight were directly influenced by the different kinds of treatment. Furthermore, FFA correlated positively with the levels of amylase and lung weight and negatively with pO2. Infusion of OA alone also caused an increase in levels of amylase with pancreatic edema and focal necroses in some animals. These results show that it was possible to create different degrees of severity of AP which was in concordance with different degrees of morphologic changes and dysfunction in the lungs. FFA values correlated significantly with the clinical course as well as with increasing amylase, lung weight and decreasing pO2.

Topics: Acute Disease; Amylases; Analysis of Variance; Animals; Fatty Acids, Nonesterified; Glycodeoxycholic Acid; Lung; Male; Oleic Acid; Oleic Acids; Organ Size; Oxygen; Pancreas; Pancreatitis; Rats; Rats, Sprague-Dawley; Respiratory Distress Syndrome

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

Intrapancreatic activation of trypsinogens is believed to occur either as a cause or a consequence of acute pancreatitis and to be associated with the more severe forms of the disease. Trypsinogen-activation peptides (TAPs) were measured in plasma, urine, and ascites of rats (n = 54) assigned to different pancreatitis-inducing regimens reproducing the entire spectrum of severity. Compared with survivors, nonsurvivors at 9 hours had significantly higher TAP levels in plasma at 3 hours (P = 0.0001), urine (peak, 1-4 hours) (P = 0.004), and ascites (P = 0.0001) after death. Stepwise discriminant analysis showed that TAP in urine and plasma were the most accurate predictors of outcome (88.2% of animals) compared with other routine laboratory parameters. Morphometric analysis showed that the best histopathologic correlates of TAP elevation were acinar necrosis and intrapancreatic hemorrhage. In a second series of experiments using a homogeneous technique of induction producing pancreatitis with a mortality of 55% at 48 hours, plasma TAP level at 3 hours (cutoff, 0.5 nmol/L) and/or urinary TAP level (peak, 1-6 hours; cutoff, 25 nmol/L) accurately predicted outcome in 85% of animals. It is concluded that the TAP assay gives an accurate early prediction of outcome in different pancreatitis models and correlates best with acinar necrosis and hemorrhage.

Topics: Acute Disease; Animals; Ascites; Glycodeoxycholic Acid; Male; Pancreatitis; Peptides; Prognosis; Rats; Rats, Inbred Strains; Severity of Illness Index; Trypsinogen

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

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

The relationship between pancreatic duct pressure, duct permeability to macromolecules and the development of acute pancreatitis was studied in a cat model. Perfusion of the pancreatic duct with 15 mM glycodeoxycholic acid, ethanol administration, or secretagogue-stimulated pancreatic secretion against greater than 50 per cent duct obstruction resulted in an increase in peak pancreatic duct pressure in all animals. Duct permeability to 20,000 molecular weight dextran molecules was increased in 22 of 29 experimental animals compared with two of 22 control animals (P less than 0.01). Perfusion of the pancreatic duct with activated pancreatic enzymes resulted in acute pancreatitis in 24 of 29 experimental animals compared with three of 22 control animals (P less than 0.01). These results suggest that pancreatic ductal hypertension, resulting in increased ductal permeability to large molecules, may be a common early event in gallstone and alcoholic pancreatitis.

Topics: Acute Disease; Animals; Cats; Cell Membrane Permeability; Constriction; Dextrans; Ethanol; Glycodeoxycholic Acid; Molecular Weight; Pancreatic Ducts; Pancreatitis; Pressure

Controlled intraduct infusion and peri-acinar dispersal of 100 microliter buffer containing sodium glycodeoxycholate (GDOC) at concentrations of 8.5, 17 and 34 mmol/l in rats caused a progressively severe acute pancreatitis from which none of the animals died over the experimental period. Infusion of affinity-purified active human enterokinase in buffer did not cause pancreatitis, presumably because of the inability of the macromolecule to gain access to its specific intracellular substrate trypsinogens. The addition of enterokinase 200 ng to GDOC 34 mmol/l in the infusate resulted in a severe systemic disturbance and a form of acute necrotizing pancreatitis which was uniformly and rapidly lethal. This effect was not seen when equimolar trypsin was substituted for enterokinase. These findings show that enterokinase specifically increases the lethality of experimental bile salt pancreatitis and suggest that this bile-borne enzyme may in some cases pose a significant clinical threat.

Topics: Acute Disease; Animals; Endopeptidases; Enteropeptidase; Glycodeoxycholic Acid; Male; Pancreatic Ducts; Pancreatitis; Rats

Acute edematous pancreatitis was induced in cats by perfusing activated pancreatic enzymes through their pancreatic ducts. The ducts had been made permeable to large molecules by one of two techniques. The cats either received ethanol (2 ml/kg every 8 h) and aspirin (25 mg/kg every 8 h) orally for 48 h or had their pancreatic ducts perfused for 1 h with 7.5 mM glycodeoxycholate. When the same procedure was followed, but using 16,16-dimethyl prostaglandin E2 (dmPGE2) (2 micrograms/kg X h infused intravenously for 1 h before and during ductal perfusion with activated enzymes), hemorrhagic pancreatitis developed instead. To investigate whether an increase in pancreatic blood flow or microvascular permeability (both caused by dmPGE2) was important in this phenomenon, we tested the effects of isoproterenol (which increased blood flow) and histamine (which increased microvascular permeability) in the model. Thus in similar experiments, either isoproterenol (0.3 micrograms/kg . min) or histamine phosphate (2 micrograms/kg . min) was infused instead of dmPGE2. The animals that received histamine also developed hemorrhagic pancreatitis. Those that received isoproterenol did not. These observations suggested that an increase in microvascular permeability in the pancreas converted edematous pancreatitis to hemorrhagic pancreatitis. These findings suggest also that clinical studies using prostaglandins to treat patients with pancreatitis should be approached with caution.

Topics: 16,16-Dimethylprostaglandin E2; Animals; Aspirin; Capillary Permeability; Cats; Ethanol; Glycodeoxycholic Acid; Hemorrhage; Histamine; Isoproterenol; Pancreas; Pancreatic Juice; Pancreatitis; Prostaglandins E, Synthetic; Regional Blood Flow

In a model of acute pancreatitis which requires that pancreatic enzymes leak from a permeable duct, we studied the role of intravenous enterokinase (195,000 daltons) in pancreatic enzyme activation. Anesthetized cats were given intravenous 16,16-dimethyl prostaglandin E2 to increase pancreatic blood flow and microvascular permeability. In some animals the permeability of the pancreatic duct was increased by perfusion of the duct with glycodeoxycholic acid (7.5 mM). Endogenous enzyme secretion was stimulated by IV CCK and secretin. Some cats also received enterokinase intravenously. Those animals that received PGE2, glycodeoxycholate, and enterokinase all developed pancreatitis. When any of these agents were not given the pancreases appeared normal. These findings were consistent with the hypothesis that intravenous enterokinase leaked from small pancreatic blood vessels into the pancreatic parenchyma and/or ducts where activation of pancreatic enzymes occurred. The development of pancreatitis appeared to require an increase in both microvascular and ductal permeability.

Topics: 16,16-Dimethylprostaglandin E2; Acute Disease; Animals; Capillary Permeability; Cats; Disease Models, Animal; Enteropeptidase; Glycodeoxycholic Acid; Pancreas; Pancreatic Ducts; Pancreatitis; Particle Size; Permeability

The cascade enterokinase-trypsinogen-prophospholipase A2 lecithin, generating trypsin, phospholipase A2 and lysolecithin, respectively, was studied in vitro using a novel phospholipase A2 assay. The rate of enterokinase catalysed activation of trypsinogen was maximal at 4 mmol/1 glycodeoxycholic acid; higher concentrations of bile salt progressively inhibited enterokinase activity. Net phospholipase A2 activity in reaction mixtures was critically dependent on the trypsin/prophospholipase A2 molar ratio. Lecithin hydrolysis by phospholipase A2 was dependent on the bile salt/lecithin molar ratio and was optimal at 1.25 to 1. The addition of enterokinase to lecithin and bile salt mixtures, containing trypsinogen and prophospholipase A2 at presumed pathophysiological concentrations, resulted in the generation of concentrations of lysolecithin lytic for pancreatic acinar cells within 5 min. These findings would support the concept that the entry of bile containing active enterokinase into the pancreatic duct system in vivo may in some cases be involved in the initiation of necrotising acute pancreatitis in man.

Topics: Acute Disease; Endopeptidases; Enteropeptidase; Enzyme Activation; Enzyme Precursors; Glycodeoxycholic Acid; Humans; Hydrolysis; Kinetics; Lysophosphatidylcholines; Necrosis; Pancreatitis; Phospholipases; Phospholipases A; Phospholipases A2; Trypsin; Trypsinogen