transforming-growth-factor-beta and Necrosis

Graphene-based materials (GBMs) are widely used in many fields, including biomedicine. To date, much attention had been paid to the potential unexpected toxic effects of GBMs. Here, we review the recent literature regarding the impact of GBMs on programmed cell death (PCD). Apoptosis, autophagy, and programmed necrosis are three major PCDs. Mechanistic studies demonstrated that the mitochondrial pathways and MAPKs (JNK, ERK, and p38)- and TGF-β-related signaling pathways are implicated in GBMs-induced apoptosis. Autophagy, unlike apoptosis and necroptosis which are already clear cell death types, plays a vital pro-survival role in cell homeostasis, so its role in cell death should be carefully considered. However, GBMs always induce unrestrained autophagy accelerating cell death. GBMs trigger autophagy through inducing autophagosome accumulation and lysosome impairment. Mitochondrial dysfunction, ER stress, TLRs signaling pathways, and p38 MAPK and NF-κB pathways participate in GBMs-induced autophagy. Programmed necrosis can be activated by RIP kinases, PARP, and TLR-4 signaling in macrophages after GBMs exposure. Though apoptosis, autophagy, and necroptosis are distinguished by some characteristics, their numerous signaling pathways comprise an interconnected network and correlate with each other, such as the TLRs, p53 signaling pathways, and the Beclin-1 and Bcl-2 interaction. A better understanding of the mechanisms of PCD induced by GBMs may allow for a thorough study of the toxicology of GBMs and a more precise determination of the consequences of human exposure to GBMs. These determinations will also benefit safety assessments of the biomedical and therapeutic applications of GBMs.

Topics: Animals; Apoptosis; Autophagy; Graphite; Humans; Macrophages; Mitochondria; Necrosis; NF-kappa B; Signal Transduction; Transforming Growth Factor beta

The HtrA family proteins are serine proteases that are involved in important physiological processes, including maintenance of mitochondrial homeostasis, apoptosis and cell signaling. They are involved in the development and progression of several pathological processes such as cancer, neurodegenerative disorders and arthritic diseases.. We present characteristics of the human HtrA1, HtrA2 and HtrA3 proteins, with the stress on their function in apoptosis and in the diseases. We describe regulation of the HtrAs' proteolytic activity, focusing on allosteric interactions of ligands/substrates with the PDZ domains, and make suggestions on how the HtrA proteolytic activity could be modified. Literature cited covers years 1996 - 2010.. An overview of the HtrAs' function/regulation and involvement in diseases (cancer, neurodegenerative disorders, arthritis), and ideas how modulation of their proteolytic activity could be used in therapies.. HtrA2 is the best target for cancer drug development. An increase in the HtrAs' proteolytic activity could be beneficial in cancer treatment, by stimulation of apoptosis, anoikis or necrosis of cancer cells, or by modulation of the TGF-beta signaling cascade; modulation of HtrA activity could be helpful in therapy of neurodegenerative diseases and arthritis.

Topics: Allosteric Regulation; Animals; Anoikis; Antineoplastic Agents; Apoptosis; Arthritis; Drug Design; Enzyme Activation; High-Temperature Requirement A Serine Peptidase 1; High-Temperature Requirement A Serine Peptidase 2; Humans; Ligands; Mitochondrial Proteins; Necrosis; Neoplasms; Neurodegenerative Diseases; PDZ Domains; Serine Endopeptidases; Signal Transduction; Transforming Growth Factor beta

Extensive bone marrow necrosis and symptomatic hypercalcemia have been described independently as rare complications of chronic myeloid leukemia. Here we report a 66-year-old man who developed B cell blastic transformation 10 years after diagnosis of CML in the chronic phase. Extensive bone marrow necrosis and symptomatic hypercalcemia concurrently developed after transformation, with development of disseminated intravascular coagulation and multifocal osteolysis. Most necrotic cells were readily identifiable as blasts. Mediators related to hypercalcemia, including prostaglandin E2, transforming growth factor-alpha and transforming growth factor-beta, were significantly elevated in the serum. As far as we know, this is the first case report of chronic myeloid leukemia concurrently developing bone marrow necrosis and hypercalcemia; this association was not reported in other types of leukemia or bone marrow malignancies.

Topics: Blast Crisis; Bone Marrow; Dinoprostone; Disseminated Intravascular Coagulation; Fatal Outcome; Humans; Hypercalcemia; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Male; Middle Aged; Necrosis; Osteolysis; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Currently, platelet concentrates are produced either from pooled buffy which are leukoreduced during processing, by various types of WBC removal filters and several apheresis technologies, which are leukocyte-reduced during collection with or without filtration. It is therefore important to define the impact of various leukocyte-removal processes on the acceleration of platelet storage of lesion and cellular apoptosis/necrosis. This overview briefly highlights the effects of exposure to artificial surfaces, during apheresis leukoreduction processes and platelet storage bags on the development of the platelet storage lesion that may contribute to transfusion reactions. The results obtained from three plateletpheresis technologies are compared with data from a "like with like" study on buffy-coat-derived platelet concentrates, using three types of platelet filter/pack assemblies. Emphasis is placed on the combined preparative methods and storage bags on generation/removal of: kallikrein/kinin; activated complement, leukocytes and platelet-derived cytokines and the development of cellular injuries, measured by the release of Annexin V. There was no systematic evidence of significant cellular fragmentation caused by filtration, but the combined preparative methods and storage bags appears to have some impact on the rate of release of soluble HLA. Moreover, large variability was observed between and within groups, in terms of various laboratory markers of biocompatibility and major biological response modifiers, indicating that much still remains to be done on various aspects of quality improvement to fully abrogate platelet concentrates associated transfusion reactions.

Topics: Annexin A5; Apoptosis; Biocompatible Materials; Blood Component Removal; Blood Platelets; Blood Preservation; Blood Transfusion; Chemokine CCL5; Chemokines; Complement C3a; Cytokines; Cytoplasm; Filtration; Humans; Immunologic Factors; Interleukin-8; Leukapheresis; Leukocyte Count; Leukocytes; Necrosis; Platelet Transfusion; Plateletpheresis; Transforming Growth Factor beta

Topics: Adult; Age Factors; Aged; Alcoholism; alpha-Tocopherol; Antiviral Agents; Biopsy; Child; Cicatrix; Enzyme-Linked Immunosorbent Assay; Fatty Liver; Female; Hemochromatosis; Hepacivirus; Hepatitis C, Chronic; HIV Infections; Humans; Immunohistochemistry; Inflammation; Lipid Peroxidation; Liver; Liver Cirrhosis; Male; Necrosis; Phenotype; Risk Factors; Sex Factors; Transforming Growth Factor beta

The uptake and removal of necrotic or lysed cells involves inflammation and an immune response, due in part to processes that involve members of the collectin family, surface calreticulin and CD91. Clearance of apoptotic cells, by contrast, does not induce either inflammation or immunity. Could the phosphatidylserine receptor be the molecular switch that determines what the outcome will be?

Topics: Animals; Antigen Presentation; Apoptosis; Calcium-Binding Proteins; Calreticulin; Carrier Proteins; Cell Adhesion; Cell Death; Cellular Senescence; Collectins; Dendritic Cells; Endopeptidases; Humans; Inflammation; Inflammation Mediators; Jumonji Domain-Containing Histone Demethylases; Low Density Lipoprotein Receptor-Related Protein-1; Membrane Lipids; Models, Biological; Necrosis; Phagocytosis; Phosphatidylserines; Receptors, Cell Surface; Receptors, Immunologic; Ribonucleoproteins; Transforming Growth Factor beta

Infarct scar, a requisite to the rebuilding of necrotic myocardium following myocardial infarction (MI), has long been considered inert. Earlier morphologic studies suggested healing at the infarct site was complete within 6-8 weeks following MI and resultant scar tissue, albeit necessary, was acellular and simply fibrillar collagen. Utilizing molecular and cellular biologic technologies, recent studies indicate otherwise. Infarct scar is composed of phenotypically transformed fibroblast-like cells, termed myofibroblasts (myoFb) because they express alpha-smooth muscle actin (alpha-SMA) and these microfilaments confer contractile behavior in response to various peptides and amines. These cells are nourished by a neovasculature and are persistent at the MI site, where they are metabolically active expressing components requisite to angiotensin (Ang) peptide generation, including converting enzyme, receptors for AngII and transforming growth factor (TGF)-beta1. They continue to elaborate fibrillar type I collagen. Their generation of these peptides contribute to ongoing scar tissue collagen turnover and to fibrous tissue formation of noninfarcted myocardium. Infarct scar contraction accounts for its thinning and its tonus may contribute to abnormal ventricular chamber stiffness with diastolic dysfunction. Infarct scar is a dynamic tissue: cellular, vascularized, metabolically active and contractile. Pharmacologic interventions with angiotensin converting enzyme inhibitor or AT1 receptor antagonist has proven effective in attenuating scar tissue metabolic activity and minimizing adverse accumulation of fibrous tissue in noninfarcted myocardium.

Topics: Angiotensin II; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Cicatrix; Collagen; Extracellular Matrix; Fibroblasts; Humans; Myocardial Infarction; Myocardium; Necrosis; Neovascularization, Pathologic; Transforming Growth Factor beta; Wound Healing

Cytokines have been associated with the pathogenesis of acute coronary syndromes and chronic heart failure (CHF), which are both associated with cardiomyocyte loss. In CHF, increased serum concentrations of proinflammatory cytokines, including tumour necrosis factor alpha (TNF-alpha) and also soluble TNF receptor have been found. Both TNF and Fas-ligand have been able to induce programmed cell death (apoptosis) of cardiomyocytes in various experimental studies. In ischaemic conditions of the heart, increased serum levels of soluble Fas receptor have been found. The proinflammatory cytokines interleukin 1 (IL-1), IL-2 and interferon-gamma can induce TNF production from target cells, including myocytes. TNF and some other cytokines are able to induce nitric oxide production, which depresses cardiac function and can induce apoptosis. However, anti-inflammatory cytokines such as IL-10, IL-4 and IL-13, secreted by T-helper type 2 lymphocytes and other cells, inhibit the production of proinflammatory cytokines. Preliminary studies suggest that cardiotrophin-1, produced by cardiomyocytes, is able to inhibit cytokine-induced cardiomyocyte apoptosis in vitro. As growth hormone is able to inhibit the production of proinflammatory cytokines in many cell types, it may also play an important role in the regulation of apoptosis induced by these cytokines. When the cytokine-induced pathways leading to altered gene expression of cardiomyocytes are understood, this knowledge may aid in the development of drugs that prevent progressive cardiomyocyte loss, in particular by inhibiting cytokine-induced apoptosis.

Topics: Apoptosis; Cytokines; Heart Failure; Humans; Interleukin-6; Myocardium; Necrosis; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

This study aimed to investigate the effect and mechanism of trauma flap healing promoted by the Zhikang capsule after radical breast cancer surgery. The enrolled breast cancer patients were randomly divided into two groups: treatment and observation. The patients in the treatment group were treated with the Zhikang capsule in addition to the conventional dressing changes, while patients in the observation group underwent only the regular dressing changes. Serum samples of 98 breast cancer patients (with complete clinical data) who underwent modified radical mastectomy were collected and analyzed for expressions of transforming growth factor beta (TGF-β) and basic fibroblast growth factor (bFGF). The drainage fluid amount and tissue necrosis rate were found to be lower in the treatment group than in the observation group. Moreover, bFGF expression in peripheral blood was higher in the treatment group than in the observation group. However, no significant difference was found between the two groups in the expression of TGF-β in peripheral blood. In conclusion, Zhikang capsule is effective in promoting flap healing after radical breast cancer surgery, and the increase of bFGF expression in peripheral blood may be the underlying mechanism.

Topics: Adult; Aged; Breast Neoplasms; Drugs, Chinese Herbal; Female; Fibroblast Growth Factor 2; Gene Expression; Humans; Mastectomy, Modified Radical; Middle Aged; Necrosis; Surgical Flaps; Transforming Growth Factor beta; Wound Healing

Patients with peripheral artery disease (PAD) and diabetes compose a high-risk population for development of critical limb ischemia (CLI) and amputation, although the underlying mechanisms remain poorly understood. Comparison of dysregulated microRNAs from diabetic patients with PAD and diabetic mice with limb ischemia revealed the conserved microRNA, miR-130b-3p. In vitro angiogenic assays demonstrated that miR-130b rapidly promoted proliferation, migration, and sprouting in endothelial cells (ECs), whereas miR-130b inhibition exerted antiangiogenic effects. Local delivery of miR-130b mimics into ischemic muscles of diabetic mice (db/db) following femoral artery ligation (FAL) promoted revascularization by increasing angiogenesis and markedly improved limb necrosis and amputation. RNA-Seq and gene set enrichment analysis from miR-130b-overexpressing ECs revealed the BMP/TGF-β signaling pathway as one of the top dysregulated pathways. Accordingly, overlapping downregulated transcripts from RNA-Seq and miRNA prediction algorithms identified that miR-130b directly targeted and repressed the TGF-β superfamily member inhibin-β-A (INHBA). miR-130b overexpression or siRNA-mediated knockdown of INHBA induced IL-8 expression, a potent angiogenic chemokine. Lastly, ectopic delivery of silencer RNAs (siRNA) targeting Inhba in db/db ischemic muscles following FAL improved revascularization and limb necrosis, recapitulating the phenotype of miR-130b delivery. Taken together, a miR-130b/INHBA signaling axis may provide therapeutic targets for patients with PAD and diabetes at risk of developing CLI.

Topics: Animals; Chronic Limb-Threatening Ischemia; Diabetes Mellitus, Experimental; Endothelial Cells; Humans; Inhibins; Ischemia; Mice; MicroRNAs; Necrosis; RNA, Small Interfering; Signal Transduction; Transforming Growth Factor beta

To identify the best low level laser photobiomodulation application site at the same irradiation time to increase the viability of the skin flap in rats. Eighteen male rats (Rattus norvegicus: var. Albinus, Rodentia Mammalia) were randomly distributed into three groups (n = 6). Group I (GI) was submitted to simulated laser photobiomodulation; group II (GII) was submitted to laser photobiomodulation at three points in the flap cranial base, and group III (GIII) was submitted to laser photobiomodulation at 12 points distributed along the flap. All groups were irradiated with an Indium, Galium, Aluminum, and Phosphorus diode laser (InGaAlP), 660 nm, with 50 mW power, irradiated for a total time of 240 s in continuous emission mode. The treatment started immediately after performing the cranial base random skin flap (10 × 4 cm

Topics: Animals; Cell Nucleus; Collagen; Fibroblast Growth Factors; Fibroblasts; Lasers, Semiconductor; Low-Level Light Therapy; Male; Necrosis; Rats, Wistar; Skin; Surgical Flaps; Tissue Survival; Transforming Growth Factor beta

Acute liver failure is a serious consequence of acetaminophen (APAP)-induced hepatotoxic liver injury with high rates of morbidity and mortality. Transforming growth factor beta 1 (TGFβ1) is elevated during liver injury and influences hepatocyte senescence during APAP-induced hepatotoxicity. This study investigated TGFβ1 signaling in the context of inflammation, necrotic cell death, and oxidative stress during APAP-induced liver injury. Male C57Bl/6 mice were injected with 600 mg/kg APAP to generate liver injury in the presence or absence of the TGFβ receptor 1 inhibitor, GW788388, 1 h prior to APAP administration. Acetaminophen-induced liver injury was characterized using histological and biochemical measures. Transforming growth factor beta 1 expression and signal transduction were assessed using immunohistochemistry, Western blotting and ELISA assays. Hepatic necrosis, liver injury, cell proliferation, hepatic inflammation, and oxidative stress were assessed in all mice. Acetaminophen administration significantly induced necrosis and elevated serum transaminases compared with control mice. Transforming growth factor beta 1 staining was observed in and around areas of necrosis with phosphorylation of SMAD3 observed in hepatocytes neighboring necrotic areas in APAP-treated mice. Pretreatment with GW788388 prior to APAP administration in mice reduced hepatocyte cell death and stimulated regeneration. Phosphorylation of SMAD3 was reduced in APAP mice pretreated with GW788388 and this correlated with reduced hepatic cytokine production and oxidative stress. These results support that TGFβ1 signaling plays a significant role in APAP-induced liver injury by influencing necrotic cell death, inflammation, oxidative stress, and hepatocyte regeneration. In conclusion, targeting TGFβ1 or downstream signaling may be a possible therapeutic target for the management of APAP-induced liver injury.

Topics: Acetaminophen; Animals; Antioxidants; Apoptosis; Benzamides; Cell Death; Chemical and Drug Induced Liver Injury; Glutathione; Hepatocytes; Inflammation; JNK Mitogen-Activated Protein Kinases; Liver; Liver Failure, Acute; Male; Mice; Mice, Inbred C57BL; Necrosis; Oxidative Stress; Phosphorylation; Protective Agents; Pyrazoles; Regeneration; Signal Transduction; Transforming Growth Factor beta

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Topics: Actins; Animals; Aquaporin 2; Aquaporin 4; Calcitriol; Cholecalciferol; Cholesterol; Disease Models, Animal; Down-Regulation; Emulsions; Epithelial Cells; Fibronectins; Fibrosis; Kidney; Male; Mice, Inbred ICR; Necrosis; Plasminogen Activator Inhibitor 1; Receptors, Calcitriol; Signal Transduction; Tissue Culture Techniques; Transforming Growth Factor beta; Ureteral Obstruction

Liver injury results in rapid regeneration through hepatocyte proliferation and hypertrophy. However, after acute severe injury, such as acetaminophen poisoning, effective regeneration may fail. We investigated how senescence may underlie this regenerative failure. In human acute liver disease, and murine models, p21-dependent hepatocellular senescence was proportionate to disease severity and was associated with impaired regeneration. In an acetaminophen injury mouse model, a transcriptional signature associated with the induction of paracrine senescence was observed within 24 hours and was followed by one of impaired proliferation. In mouse genetic models of hepatocyte injury and senescence, we observed transmission of senescence to local uninjured hepatocytes. Spread of senescence depended on macrophage-derived transforming growth factor-β1 (TGFβ1) ligand. In acetaminophen poisoning, inhibition of TGFβ receptor 1 (TGFβR1) improved mouse survival. TGFβR1 inhibition reduced senescence and enhanced liver regeneration even when delivered beyond the therapeutic window for treating acetaminophen poisoning. This mechanism, in which injury-induced senescence impairs liver regeneration, is an attractive therapeutic target for developing treatments for acute liver failure.

Topics: Animals; Cellular Senescence; Cyclin-Dependent Kinase Inhibitor p21; Disease Models, Animal; Hepatocytes; Humans; Liver; Liver Regeneration; Macrophages; Male; Mice, Inbred C57BL; Necrosis; Paracrine Communication; Signal Transduction; Transforming Growth Factor beta

Invasion into surrounding normal brain and resistance to genotoxic therapies are the main devastating aspects of glioblastoma (GBM). These biological features may be associated with the stem cell phenotype, which can be induced through a dedifferentiation process known as epithelial-mesenchymal transition (EMT). We show here that tumor cells around pseudopalisading necrotic areas in human GBM tissues highly express the most important EMT inducer, transforming growth factor (TGF-β), concurrently with the EMT-related transcriptional factor, TWIST. In addition, the stem cell markers CD133 and alkaline phosphatase (ALPL) were also highly expressed around necrotic foci in GBM tissues. The high expression of TGF-β around necrotic regions was significantly correlated with shorter progression-free survival and overall survival in patients with GBM. High expression of stem cell markers, ALPL, CD133, and CD44 was also correlated with poor outcomes. These results collectively support the hypothesis that tissue hypoxia induces the stem cell phenotype through TGF-β-related EMT and contributes to the poor outcome of GBM patients.

Topics: AC133 Antigen; Aged; Alkaline Phosphatase; Biomarkers, Tumor; Brain Neoplasms; Disease-Free Survival; Epithelial-Mesenchymal Transition; Female; Glioblastoma; Humans; Hyaluronan Receptors; Male; Middle Aged; Necrosis; Neoplastic Stem Cells; Nuclear Proteins; Retrospective Studies; Transforming Growth Factor beta; Twist-Related Protein 1

Acute kidney injury (AKI) is exacerbated in C-reactive protein transgenic mice but alleviated in Smad3 knockout mice. Here we used C-reactive protein transgenic/Smad3 wild-type and C-reactive protein transgenic/Smad3 knockout mice to investigate the signaling mechanisms by which C-reactive protein promotes AKI. Serum creatinine was elevated, and the extent of tubular epithelial cell necrosis following ischemia/reperfusion-induced AKI was greater in C-reactive protein transgenics but was blunted when Smad3 was deleted. Exacerbation of AKI in C-reactive protein transgenics was associated with increased TGF-β/Smad3 signaling and expression of the cyclin kinase inhibitor p27, but decreased phosphorylated CDK2 and expression of cyclin E. Concomitantly, tubular epithelial cell proliferation was arrested at the G1 phase in C-reactive protein transgenics with fewer cells entering the S-phase cell cycle as evidenced by fewer bromodeoxyuridine-positive cells. In contrast, the protection from AKI in C-reactive protein transgenic/Smad3 knockout mice was associated with decreased expression of p27 and promotion of CDK2/cyclin E-dependent G1/S transition of tubular epithelial cells. In vitro studies using tubular epithelial cells showed that C-reactive protein activates Smad3 via both TGF-β-dependent and ERK/MAPK cross talk mechanisms, Smad3 bound directly to p27, and blockade of Smad3 or the Fc receptor CD32 prevented C-reactive protein-induced p27-dependent G1 cell cycle arrest. In vivo, treatment of C-reactive protein transgenics with a Smad3 inhibitor largely improved AKI outcomes. Thus, C-reactive protein may promote AKI by impairing tubular epithelial cell regeneration via the CD32-Smad3-p27-driven inhibition of the CDK2/cyclin E complex. Targeting Smad3 may offer a new treatment approach for AKI.

Topics: Acute Kidney Injury; Animals; C-Reactive Protein; Cell Line, Tumor; Cell Proliferation; Creatinine; Cyclin E; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase Inhibitor p27; Disease Models, Animal; Epithelial Cells; G1 Phase Cell Cycle Checkpoints; Humans; Isoquinolines; Kidney Tubules; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mice, Knockout; Necrosis; Phosphorylation; Pyridines; Pyrroles; Rats; Receptors, IgG; Regeneration; Smad3 Protein; Transforming Growth Factor beta

Graphene oxide (GO) has been a focus of research in the fields of electronics, energy, and biomedicine, including drug delivery. Thus, single- and multi-layered GO (SLGO and MLGO) have been produced and investigated. However, little information on their toxicity and biocompatibility is available. In the present study, we performed a comprehensive study of the size- and dose-dependent toxicity of GOs in the presence or absence of Pluronic F-127 on THP-1 cells by examining their viability, membrane integrity, levels of cytokine and ROS production, phagocytosis, and cytometric apoptosis. Moreover, as an extended study, a toxicity evaluation in the acute and chronic phases was performed in mice via intravenous injection of the materials. GOs exhibited dose- and size-dependent toxicity. Interestingly, SLGO induced ROS production to a lesser extent than MLGO. Cytometric analysis indicated that SLGO induced necrosis and apoptosis to a lesser degree than MLGO. In addition, cell damage and IL-1β production were influenced by phagocytosis. A histological animal study revealed that GOs of various sizes induced acute and chronic damage to the lung and kidney in the presence or absence of Pluronic F-127. These results will facilitate studies of GO prior to its biomedical application.

Topics: Animals; Apoptosis; Chemokine CCL2; Cytochalasin D; Dose-Response Relationship, Drug; Graphite; Humans; Interleukin-1beta; Kidney; L-Lactate Dehydrogenase; Lung; Mice; Necrosis; Oxides; Phagocytosis; Poloxamer; Reactive Oxygen Species; Tetradecanoylphorbol Acetate; THP-1 Cells; Transforming Growth Factor beta

The deleterious effects of increased cadmium (Cd) serum levels on the cardiovascular system are proven by epidemiological and basic science studies. Cd exposure of animals and humans is known to impair myocardial function, possibly leading to heart failure. This study aims at investigating the effect of Cd treatment on the cardiac system with emphasis on the combined effect of Cd and high serum cholesterol levels as an important cardiovascular risk factor. Detailed analyses of Cd-induced effects on the heart of ApoE-/- mice fed a high fat diet (HFD), ApoE-/- mice fed a normal diet (ND), and C57BL/6J mice fed a ND revealed proinflammatory and fibrotic changes in the presence of cellular hypertrophy but in the absence of organ hypertrophy. Hypercholesterolemia in ApoE-/- mice alone and in combination with Cd treatment resulted in significant cardiomyocyte cell death. Based on further analyses of heart sections, we conclude that severe hypercholesterolemia in combination with ApoE-/- genotype as well as Cd treatment results in necrotic cardiomyocyte death. These data were supported by in vitro experiments showing a Cd-induced depolarization of the mitochondrial membrane and the permeabilization of the plasma membrane arguing for the occurrence of Cd-induced necrotic cell death. In summary, we were able to show for the first time that the combination of high cholesterol and Cd levels increase the risk for heart failure through cardiac fibrosis. This observation could in part be explained by the dramatically increased deposition of Cd in the hearts of ApoE-/- mice fed a HFD.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Apolipoproteins E; Biomarkers; Body Burden; Cadmium Chloride; Cardiomyopathies; Cell Line; Chemokine CCL2; Cholesterol; Diet, High-Fat; Disease Models, Animal; Female; Fibrosis; Hypercholesterolemia; Membrane Potential, Mitochondrial; Mice, Inbred C57BL; Mice, Knockout; Mitochondria, Heart; Myocardium; Necrosis; Transforming Growth Factor beta

The mouse model of arterialized orthotopic liver transplantation (AOLT) has played an important role in biomedical research. The available methods of sutured anastomosis for reconstruction of the hepatic artery are complicated, resulting in a high incidence of complications and failure. Therefore, we developed and evaluated a new model of AOLT in mice.. Male inbred C57BL/6 mice were used in this study. A continuous suture approach was applied to connect the suprahepatic inferior vena cava (SHVC). The portal vein and infrahepatic inferior vena cava (IHVC) were connected according to the "two-cuff" method. The common bile duct was connected by a biliary stent. We used the stent (G3 group) or aortic trunk (G2 group) to reconstruct the hepatic artery. The patency of the hepatic artery was verified by transecting the artery near the graft after one week. The survival rate of the recipients and serum alanine aminotransferase (ALT) levels, hepatic pathologic alterations, apoptosis and necrosis were observed at one week postoperatively.. The patency of the hepatic artery was verified in eight of ten mice in G3 and in six of ten mice in G2. The 7-day survival rate, extents of necrosis and apoptosis, and TGF-β levels were not significantly different among the three groups (P>0.05). However, the serum ALT levels and operation time were markedly lower in G3 compared with G2 or G1 (both P<0.05).. Reconstruction of the hepatic artery using a stent can be performed quickly with a high rate of patency. This model simplifies hepatic artery anastomosis and should be promoted in the field of biomedical research.

Topics: Alanine Transaminase; Anastomosis, Surgical; Animals; Aorta, Abdominal; Apoptosis; Blood Vessel Prosthesis Implantation; Common Bile Duct; Hepatic Artery; Liver; Liver Transplantation; Male; Mice; Mice, Inbred C57BL; Necrosis; Portal Vein; Random Allocation; Stents; Transforming Growth Factor beta; Vascular Patency; Vena Cava, Inferior

n-3 polyunsaturated fatty acids (PUFA) ameliorate fatty liver in experimental models, but their effects on inflammation and fibrosis during steatohepatitis are either controversial or lacking. We compared the effects of supplementation with olive oil (OO) alone or OO and n-3 PUFA on the development and progression of experimental steatohepatitis.. Balb/C mice (≥5 mice/group) were fed a methionine- and choline-deficient (MCD) diet or a control diet for 4 or 8 weeks. At the same time, mice were supplemented with n-3 PUFA (eicosapentaenoic and docosahexahenoic acid, 25 mg together with 75 mg OO), or OO alone (100 mg), two times a week by intragastric gavage.. After 8 weeks, mice on MCD/n-3 had higher ALT levels compared to MCD/OO and more severe scores of inflammation, including a significant increase in the number of lipogranulomas (26.4 ± 8.4 vs. 5.1 ± 5 per field, P < 0.001). Intrahepatic expression of TNF-α and CCL2 was higher in MCD/n-3 mice at both time points. In addition, increased expression of the profibrogenic genes TIMP-1 and TGF-β, and more severe histological scores of fibrosis were evident in MCD/n-3 mice. After 8 week of MCD diet, portal pressure was higher in mice receiving n-3 than in those on OO alone (5.1 ± 1.4 vs. 7.0 ± 0.9 mmHg, P < 0.05). Analysis of hepatic fatty acid profile showed that supplementation resulted in effective incorporation of n-3 PUFA.. In a murine model of steatohepatitis, supplementation with n-3 PUFA and OO is associated with more severe necro-inflammation and fibrosis than in mice treated with OO only.

Topics: Animals; Biomarkers; Chemical and Drug Induced Liver Injury; Choline Deficiency; Dietary Supplements; Disease Models, Animal; Fatty Acids, Omega-6; Inflammation Mediators; Liver; Liver Cirrhosis; Male; Methionine; Mice, Inbred BALB C; Necrosis; Non-alcoholic Fatty Liver Disease; Olive Oil; Plant Oils; Time Factors; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta

To investigate the effect of lymph circulation blockage on the alteration of renal epithelial cell phenotype and the tubular integrity, as well as the underlying mechanisms.. Wistar rats received left renal lymph ligation and right renal nephrectomy (KL group) or right renal nephrectomy without lymph ligation (KN group) and then were killed on day 14, day 28 and day 56. The urine, blood and kidney tissue were collected for the analysis of protein and gene expressions and morphological changes.. The urine albumin and serum creatinine (Cr) in KL group were significantly increased compared with KN group. Masson and PAS staining indicated the epithelial cell degeneration, necrosis, sublethal loss and atrophy in KL rats, but not in KN group. Interestingly, from the atrophic tubules, some epithelial cells exhibited polarity changes with hypertrophy contrasting to the normal epithelial morphology of KN group throughout the experiment. By EM, ligated kidneys showed irregularly wrinkled basement membranes and epithelial cell swelling. Some intertubular areas of the KL kidney were expanded with fibrotic matrix and fibroblast-like cells. In line with these morphological changes, the fibroblast cell markers of FSP1 and α-SMA were markedly increased in contrast to the remarkable reduction in epithelial cell marker E-cadherin and tight junction protein ZO-1. Moreover, the TGF-β1/Smad2/3 signaling pathway was significantly activated in KL rats in contrast to a robust downregulation of BMP7/Smad5 signaling.. Disturbance of renal lymphatic circulation resulted in the epithelial cell phenotypic alteration and impaired tubular integrity possibly via distinct regulation of TGFβ1/Smads and BMP7/Smad5 signaling pathway.

Topics: Actins; Albuminuria; Animals; Atrophy; Bone Morphogenetic Protein 7; Cadherins; Calcium-Binding Proteins; Creatinine; Epithelial Cells; Epithelial-Mesenchymal Transition; Kidney Tubules; Ligation; Lymphatic Vessels; Necrosis; Nephrectomy; Phenotype; Rats, Wistar; RNA, Messenger; Signal Transduction; Smad2 Protein; Smad3 Protein; Smad5 Protein; Transforming Growth Factor beta; Zonula Occludens-1 Protein

To evaluate the antioxidant, immunomodulatory, antinecrotic and antifibrotic effects of hesperidin on CCl4-induced cirrhosis.. Liver damage was produced by giving CCl4 injections (0.4 g/kg, i.p., 3 times per week for 8 weeks) to rats. Hesperidin (200 mg/kg) was administered using gavage. The expression of nuclear factor-κB (NF-κB), transforming growth factor-β (TGF-β), connective tissue growth factor (CTGF), interleukin (IL)-10 and IL-1β was assessed using Western blotting. Alanine aminotransferase (ALT) and γ-glutamyl transpeptidase (γ-GTP) serum activities, glycogen content, reduced/oxidised glutathione (GSH/GSSG) ratio, lipid peroxidation degree and fibrosis (using hydroxyproline content and a histopathological analysis) were measured.. CCl4 increased the enzymatic activities of ALT and γ-GTP, liver lipid peroxidation, the hydroxyproline content as well as NF-κB, TGF-β, CTGF, IL-1β and IL-10 levels and decreased the glycogen content and GSH/GSSG ratio. Hesperidin significantly decreased the modifications produced by CCl4, except in the case of IL-10, which was further increased by the flavone. The group receiving hesperidin alone showed decreases in lipid peroxidation, NF-κB, TGF-β, CTGF and IL-1β and an increase in IL-10. The results of the histopathological analysis were in agreement with the biochemical and molecular findings.. This study demonstrates that hesperidin prevents experimental necrosis and fibrosis. The action mechanism of hesperidin is associated with its ability to reduce oxidative stress and modulate proinflammatory and profibrotic signals. These results support earlier findings demonstrating the beneficial effect of hesperidin against liver damage.

Topics: Animals; Antioxidants; Blotting, Western; Carbon Tetrachloride; Connective Tissue Growth Factor; Disease Models, Animal; Gene Expression Regulation; Hesperidin; Lipid Peroxidation; Liver Cirrhosis; Male; Necrosis; NF-kappa B; Oxidative Stress; Rats; Rats, Wistar; Transforming Growth Factor beta

Antiangiogenic therapy is a promising strategy for cancer therapy. However, antiangiogenic therapy can induce intratumor hypoxia and hypoxia-inducible factor-1 (HIF-1) expression, which slows cancer progression. In our present study, we found that antiangiogenic therapy with sunitinib plus HIF-1 dimerization inhibitor acriflavine retarded tumor growth in a murine model of breast cancer. The combination of sunitinib with acriflavine significantly decreased vascular endothelial growth factor and TGF-β expression and reduced tumor vasculature followed by increased intratumor necrosis and apoptosis. Moreover, decreased accumulation of myeloid-derived suppressor cells in the spleen was observed after the combinational therapy. In conclusion, the combination of HIF-1 inhibition and antiangiogenic therapy may represent a novel strategy for cancer patients.

Topics: Acriflavine; Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Cell Line, Tumor; Disease Models, Animal; Drug Synergism; Female; Hypoxia-Inducible Factor 1; Indoles; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Myeloid Cells; Necrosis; Neovascularization, Pathologic; Protein Multimerization; Pyrroles; Spleen; Sunitinib; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Bovine pancreatic trypsin inhibitor (BPTI) has been reported to relieve liver ischemia-reperfusion-induced injury in rats.. This study was designed to determine whether the recombinant BPTI (rBPTI) can prevent the chronic liver injury induced by CCl4 in rats.. Fifty male Wistar rats were divided into five groups. Rats were treated with 40% CCl4 at a dose of 2 ml/kg body weight twice a week subcutaneously for 12 weeks. In the 8th week, they were administered intraperitoneally with rBPTI (80 MU/kg), BPTI (80 MU/kg) or hepatocyte growth-promoting factor (pHGF; 100 mg/kg) daily for the next 4 weeks.. rBPTI significantly prevented the disruption of liver function of alanine aminotransferase (ALT; 172.7 ± 18.16 versus 141.2 ± 15.28, p=0.003), aspartate aminotransferase (AST; 225.10 ± 36.54 versus 170.06 ± 27.14, p=0.007) and hydroxyproline (Hyp; 1.14 ± 0.27 versus 0.62 ± 0.17, p=0.001). rBPTI significantly decreased the level of thiobarbituric acid reactive substances (TBARS; 1.15 ± 0.16 versus 0.87 ± 0.15, p=0.003) and increased the activities of superoxide dismutase (SOD; 6.07 ± 0.95 versus 7.75 ± 1.12, p=0.007). rBPTI reduced the production of cytokines of IL-1β and TGF-β. The hepatocyte necrosis, fibrosis, fatty degeneration and inflammatory cell infiltration were ameliorated by rBPTI administration.. This study demonstrated that rBPTI exerted a hepatoprotective effect on chronic liver fibrosis induced by CCl4, which suggests that rBPTI may have the potential application for chronic liver injury induced by drugs metabolism and toxic substances.

Topics: Alanine Transaminase; Animals; Aprotinin; Aspartate Aminotransferases; Carbon Tetrachloride; Chemical and Drug Induced Liver Injury, Chronic; Cytoprotection; Disease Models, Animal; Fatty Liver; Hydroxyproline; Inflammation Mediators; Interleukin-1beta; Liver; Liver Cirrhosis, Experimental; Male; Necrosis; Protective Agents; Rats; Rats, Wistar; Recombinant Proteins; Superoxide Dismutase; Thiobarbituric Acid Reactive Substances; Transforming Growth Factor beta

Endothelial progenitor cells (EPCs) protect the kidney from acute ischemic injury. The aim of this study was to analyze whether pretreatment of murine "early outgrowth" EPCs (eEPCs) with the hormone melatonin increases the cells' renoprotective effects in the setting of murine acute ischemic renal failure. Male (8-12 wk old) C57Bl/6N mice were subjected to unilateral ischemia-reperfusion injury postuninephrectomy (40 min). Postischemic animals were injected with either 0.5×10(6) untreated syngeneic murine eEPCs or with cells, pretreated with melatonin for 1 h. Injections were performed shortly after reperfusion of the kidney. While animals injected with untreated cells developed acute renal failure, eEPC pretreatment with melatonin dramatically improved renoprotective actions of the cells. These effects were completely reversed after cell pretreatment with melatonin and the MT-1/-2 antagonist luzindole. In vitro analysis revealed that melatonin reduced the amount of tumor growth factor-β-induced eEPC apoptosis/necrosis. Secretion of vascular endothelial growth factor by the cells was markedly stimulated by the hormone. In addition, migratory activity of eEPCs was enhanced by melatonin and supernatant from melatonin-treated eEPCs stimulated migration of cultured mature endothelial cells. In summary, melatonin was identified as a new agonist of eEPCs in acute ischemic kidney injury.

Topics: Acute Kidney Injury; Animals; Antioxidants; Apoptosis; Cell Movement; Cells, Cultured; Endothelial Cells; Male; Melatonin; Mice; Mice, Inbred C57BL; Necrosis; Neovascularization, Physiologic; Recovery of Function; Reperfusion Injury; Stem Cells; Transforming Growth Factor beta

Recombinant human bone morphogenetic protein-2 (rhBMP-2) delivered on an absorbable collagen sponge has been shown to induce the healing of acute, primary, large-scale calvarial defects in rabbits. However, clinical circumstances often require the reconstruction of a previously infected and chronically scarred wound. This study was designed to evaluate the efficacy of rhBMP-2/absorbable collagen sponge to improve healing in the previously infected, unfavorable calvarial defect model.. Subtotal defects were made in the calvariae of 15 adult New Zealand White rabbits. The bone flap was inoculated with Staphylococcus aureus and replaced in situ. After a 2-week infection period, animals underwent bone flap removal and a 10-day course of antibiotic therapy. On postoperative day 42, the defect was exposed and treated with (1) no intervention/control (group 1; n = 5), or (2) absorbable collagen sponge with 0.43 mg/ml of rhBMP-2 (group 2; n = 10). Bone growth was analyzed with serial computed tomographic imaging and postmortem histology. Percentage bone healing was compared between groups using the t test.. The treatment group (group 2) demonstrated statistically more healing (55.6 percent) compared with the control group (group 1) (29 percent; p < 0.01). However, rhBMP-2-induced bone was not histologically or radiographically similar to native bone, lacking both continuity and a well-defined diploic space.. These data suggest that rhBMP-2-treated collagen sponges may be useful for the repair of calvarial defects following infection. However, the osseous healing observed in this study was significantly less than previous reports in acute, noninfected models and was dissimilar to native bone. Further work is needed to optimize treatment of the previously infected calvarial wound with rhBMP-2.

Topics: Animals; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Bone Regeneration; Disease Models, Animal; Necrosis; Osteotomy; Rabbits; Recombinant Proteins; Skull; Tomography, X-Ray Computed; Transforming Growth Factor beta; Wound Healing

Cirrhotic septa harbor vessels and inflammatory, fibrogenic, and ductular epithelial cells, collectively referred to as the ductular reaction (DR). Lack of the DR in the stromal compartment around hepatocellular carcinoma (HCC) has been documented; however, the relationship of epithelial keratin 19 (K19) structures to progression of intralesional carcinogenesis has not been explored. K19 immunoreactivity in the stromal compartment around 176 nodules in cirrhotic explants was examined. Quantitative differences (P < 0.0001) were manifested in three distinct histologically identifiable patterns: "complex" around cirrhotic nodules (CN), "attenuated" around dysplastic nodules (DN), and "absent" around HCC. Markers of necrosis or apoptosis could not explain the perinodular K19 epithelial loss; however, multicolor immunolabeling for K19, vimentin, E-Cadherin, SNAIL, and fibroblast-specific protein 1 (FSP-1) demonstrated discrepancies in immunophenotype and cytomorphologic features. Variability of cellular features was accompanied by an overall decrease in epithelial markers and significantly increased fractions of SNAIL- and FSP-1-positive cells in the DR around DN when compared with CN (P < 0.0001). Immunolabeling of transforming growth factor-β signaling components (TGFβR1, SMAD3, and pSMAD2/3) demonstrated increased percentages of pSMAD2/3 around DN when compared with CN (P < 0.0001). These findings collectively suggest marked alterations in cellular identity as an underlying mechanism for the reproducible extralesional K19 pattern that parallels progressive stages of intranodular hepatocarcinogenesis. Paracrine signaling is proposed as a link that emphasizes the importance of the epithelial-stromal compartment in malignant progression of HCC in cirrhosis.

Topics: Apoptosis; Biopsy; Carcinoma, Hepatocellular; Epithelium; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Humans; Keratin-19; Liver Cirrhosis; Liver Neoplasms; Models, Biological; Necrosis; Predictive Value of Tests; Signal Transduction; Transforming Growth Factor beta

We have previously reported that pancreatic stellate cells (PSCs) have a phagocytic function. The aim of the present study was to investigate whether engulfment of necrotic acinar cells affects pancreatic fibrogenesis.. Rat pancreatic acinar cells were incubated for 48 hours to induce necrosis, and PSCs were allowed to interact with them for 12 to 48 hours. Annexin V and propidium iodide staining or detection of DNA fragmentation was used to identify cell death.. A large number of necrotic acinar cells were engulfed by PSCs. When PSCs were exposed to necrotic acinar cells for 12 hours, the number of living PSCs was significantly lower than among the control PSCs, which were not exposed to necrotic acinar cells. DNA degradation was observed in PSCs that had ingested necrotic acinar cells, and they were Annexin V and propidium iodide positive, suggesting that engulfment of necrotic acinar cells induced PSC death. There was no difference between the concentrations of transforming growth factor-beta in the medium of the PSCs that had engulfed acinar cells and the medium of the control PSCs.. Engulfment of necrotic acinar cells by PSCs induces PSC death, suggesting that engulfment of necrotic acinar cells may inhibit the progression of fibrogenesis.

Topics: Animals; Apoptosis; Cell Proliferation; Cells, Cultured; Culture Media, Conditioned; Fibrosis; Male; Necrosis; Pancreas, Exocrine; Phagocytes; Phagocytosis; Rats; Rats, Wistar; Time Factors; Transforming Growth Factor beta

Liver fibrosis is characterized by an excess of collagen fiber deposition, and it is known that Kupffer cells play an important role by immunomodulation of the toxic response. Methyl palmitate (MP) is an effective Kupffer cell inhibitor. The aim of this work was to evaluate the effect of MP on experimental liver fibrosis. Four groups were formed: the control group, which received the vehicles only; CCl(4) group (0.4 g kg(-1), i.p., three times a week, for eight weeks); CCl(4) plus MP (300 mg kg(-1), i.p., daily); and MP alone. Alanine aminotransferase was increased by CCl(4), and MP did not prevent this increase. Lipid peroxidation was increased markedly by CCl(4); again, MP was not able to prevent this effect. Fibrosis increased nearly 6-fold (measured as liver hydroxyproline content) in the CCl(4) group; MP preserved the normal content of collagen. These results were corroborated by histopathology. To elucidate the antifibrogenic mechanism of MP, we measured the production of TGF-beta; CCl(4) increased this cytokine several-fold, and MP abolished this increase. Collectively the present results indicate that MP possesses a strong antifibrogenic effect at least in the CCl(4) model of fibrosis. The antifibrotic effect of MP is probably associated with its ability to reduce TGF-beta content, maybe by immunomodulation of Kupffer cells functioning.

Topics: Animals; Biotransformation; Blotting, Western; Carbon Tetrachloride; Carbon Tetrachloride Poisoning; Collagen; Down-Regulation; Histocytochemistry; Lipid Peroxidation; Liver Cirrhosis; Male; Necrosis; Palmitates; Rats; Rats, Wistar; Transforming Growth Factor beta

Dendritic cells (DC) are sensitive to their local environment and are affected by proximal cell death. This study investigated the modulatory effect of cell death on DC function. Monocyte-derived DC exposed to apoptotic Jurkat or primary T cells failed to induce phenotypic maturation of the DC and were unable to support CD4+ allogeneic T-cell proliferation compared with DC exposed to lipopolysaccharide (LPS) or necrotic cells. Apoptotic cells coincubated with LPS- or necrotic cell-induced mature DC significantly suppressed CD80, CD86 and CD83 and attenuated LPS-induced CD4+ T-cell proliferation. Reduced levels of interleukin-12 (IL-12), IL-10, IL-6, tumour necrosis factor-alpha and interferon-gamma (IFN-gamma) were found to be concomitant with the suppressive activity of apoptotic cells upon DC. Furthermore, intracellular staining confirmed IFN-gamma expression by DC in association with apoptotic environments. The specific generation of IFN-gamma by DC within apoptotic environments is suggestive of an anti-inflammatory role by the induction of indoleamine 2,3-dioxygenase (IDO). Both neutralization of IFN-gamma and IDO blockade demonstrated a role for IFN-gamma and IDO in the suppression of CD4+ T cells. Moreover, we demonstrate that IDO expression within the DC was found to be IFN-gamma-dependent. Blocking transforming growth factor-beta (TGF-beta) also produced a partial release in T-cell proliferation. Our study strongly suggests that apoptosis-induced DC suppression is not an immunological null event and two prime mediators underpinning these functional effects are IFN-gamma-induced IDO and TGF-beta.

Topics: Apoptosis; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Proliferation; Cells, Cultured; Coculture Techniques; Cytokines; Dendritic Cells; Humans; Immune Tolerance; Immunophenotyping; Indoleamine-Pyrrole 2,3,-Dioxygenase; Interferon-gamma; Lipopolysaccharides; Lymphocyte Activation; Necrosis; Transforming Growth Factor beta; Up-Regulation

Podocyte injury and loss contribute to progressive glomerulosclerosis. Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is a nuclear hormone receptor, which we have found to be increased in podocytes in a variety of kidney diseases. It is not known if PPAR-gamma contributes to renal injury or if it serves as a countermeasure to limit renal injury during disease progression. We tested these possibilities utilizing the puromycin aminonucleoside (PAN) model of renal injury in immortalized mouse podocytes. The cultured podocytes expressed PPAR-gamma mRNA at baseline but this was decreased by PAN. Pioglitazone, a pharmacologic agonist of PPAR-gamma, increased both PPAR-gamma mRNA and activity in injured podocytes, as assessed by a reporter plasmid assay. Further, pioglitazone significantly decreased PAN-induced podocyte apoptosis and necrosis while restoring podocyte differentiation. The PPAR-gamma agonist significantly restored expression of the cyclin-dependent kinase inhibitor p27 and the antiapoptotic molecule Bcl-xL while significantly decreasing proapoptotic caspase-3 activity. Pioglitazone tended to decrease PAN-induced transforming growth factor-beta (TGF-beta) mRNA expression. Our study shows that PPAR-gamma is normally expressed by podocytes and its activation is protective against PAN-induced apoptosis and necrosis. We postulate that this protective effect may be mediated in part by effects on p27 and TGF-beta expression.

Topics: Animals; Apoptosis; bcl-X Protein; Caspase Inhibitors; Cell Proliferation; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p27; Kidney Glomerulus; Mice; Necrosis; Pioglitazone; Plasmids; Podocytes; PPAR gamma; Puromycin Aminonucleoside; RNA, Messenger; Thiazolidinediones; Transforming Growth Factor beta

We have previously demonstrated that clinically utilized volatile anesthetics protect against renal ischemia reperfusion injury in rats in vivo and reduce necrosis in vitro via activation of ERK and Akt and by upregulating HSP70. In this study, we further deciphered the upstream cellular signaling mechanism(s) of volatile anesthetic-mediated antinecrotic effects in vitro. We hypothesized that volatile anesthetics perturb the structure of the plasma membrane lipid bilayer, causing externalization of phosphatidylserine (PS) to the outer surface on renal tubule cells leading to the increased generation of transforming growth factor-beta1 (TGF-beta1), a cytokine with antinecrotic properties.. In human proximal tubule (HK-2) cell culture, 16-hour exposure to volatile anesthetics (isoflurane, halothane, sevoflurane) caused membrane externalization of PS detected by positive annexin-V staining and increased the release of TGF-beta1 into the cell culture media. Exogenous TGF-beta1 induced protection and neutralizing TGF-beta1 antibody prevented the cytoprotection by volatile anesthetics against hydrogen peroxide-induced HK-2 cell necrosis.. Volatile anesthetics induce a cytoprotective signaling cascade in proximal tubule cells via membrane externalization of PS initiating TGF-beta1-mediated cytoprotection.

Topics: Anesthetics, Inhalation; Antibodies; Cell Line; Cell Membrane; Humans; Hydrogen Peroxide; Kidney Tubules, Proximal; Necrosis; Phosphatidylserines; Transcription, Genetic; Transforming Growth Factor beta

Oxidative stress plays an important role in acetaminophen (APAP)-induced hepatotoxicity. In addition to inducing direct cellular damage, oxidants can activate transcription factors including NF-kappaB, which regulate the production of inflammatory mediators implicated in hepatotoxicity. Here, we investigated the role of APAP-induced oxidative stress and NF-kappaB in inflammatory mediator production. Treatment of mice with APAP (300 mg/kg, i.p.) resulted in centrilobular hepatic necrosis and increased serum aminotransferase levels. This was correlated with depletion of hepatic glutathione and CuZn superoxide dismutase (SOD). APAP administration also increased expression of the proinflammatory mediators, interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNFalpha), macrophage chemotactic protein-1 (MCP-1), and KC/gro, and the anti-inflammatory cytokine, interleukin-10 (IL-10). Pretreatment of mice with the antioxidant, N-acetylcysteine (NAC) prevented APAP-induced depletion of glutathione and CuZnSOD, as well as hepatotoxicity. NAC also abrogated APAP-induced increases in TNFalpha, KC/gro, and IL-10, but augmented expression of the anti-inflammatory cytokines interleukin-4 (IL-4) and transforming growth factor-beta (TGFbeta). No effects were observed on IL-1beta or MCP-1 expression. To determine if NF-kappaB plays a role in regulating mediator production, we used transgenic mice with a targeted disruption of the gene for NF-kappaB p50. As observed with NAC pretreatment, the loss of NF-kappaB p50 was associated with decreased ability of APAP to upregulate TNFalpha, KC/gro, and IL-10 expression and increased expression of IL-4 and TGFbeta. However, in contrast to NAC pretreatment, the loss of p50 had no effect on APAP-induced hepatotoxicity. These data demonstrate that APAP-induced cytokine expression in the liver is influenced by oxidative stress and that this is dependent, in part, on NF-kappaB. However, NF-kappaB p50-dependent responses do not appear to play a major role in the pathogenesis of toxicity in this model.

Topics: Acetaminophen; Acetylcysteine; Alanine Transaminase; Analgesics, Non-Narcotic; Animals; Blotting, Western; Chemical and Drug Induced Liver Injury; Chemokine CCL2; Chemokines; Gene Expression; Glutathione; Humans; Inflammation Mediators; Injections, Intraperitoneal; Interleukin-1; Interleukin-10; Interleukin-4; Liver; Macrophage Inflammatory Proteins; Male; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Necrosis; NF-kappa B p50 Subunit; Oxidative Stress; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Superoxide Dismutase; Time Factors; Transcription Factors; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Interleukin-1 (IL-1) and transforming growth factor-beta (TGFbeta) are key inhibitors of hepatocyte proliferation after hepatectomy. IL-1 inhibition by heat shock proteins (HSPs) has been reported in inflammatory processes. A recent study indicated the benefits of ischaemic preconditioning in reduced-size orthotopic liver transplantation (ROLT). The present study examined: (a) the effect of ischaemic preconditioning on IL-1 and TGFbeta in ROLT; (b) whether preconditioning protects small liver grafts through HSP induction; and (c) whether the potential benefits of preconditioning on HSP is related to IL-1 inhibition. Our results, obtained with an IL-1 receptor antagonist, indicated the injurious effects of IL-1 in ischaemia-reperfusion (I/R) injury and established a relationship between IL-1 and growth factors. Thus, IL-1 reduced hepatocyte growth factor (HGF) and promoted TGFbeta release, thus contributing to the impaired liver regeneration associated with ROLT. Preconditioning inhibited IL-1 through nitric oxide (NO), thereby protecting against the injurious effects of IL-1. In addition, by another pathway independent of NO, preconditioning induced HSP70 and haem-oxygenase-1 (HO-1). HO-1 protected against I/R injury and liver regeneration, whereas the benefits resulting from HSP70 were mainly related to hepatocyte proliferation. These results suggest a mechanism that explains the effectiveness of preconditioning in ROLT. They suggest, too, that other strategies, in addition to preconditioning, that modulate IL-1 and/or HSPs could be considered in clinical situations requiring liver regeneration such as small liver grafts.

Topics: Animals; Heat-Shock Proteins; Heme Oxygenase-1; Hepatocytes; HSP70 Heat-Shock Proteins; HSP90 Heat-Shock Proteins; Immunohistochemistry; Interleukin-1; Ischemic Preconditioning; Liver; Liver Regeneration; Liver Transplantation; Male; Membrane Proteins; Necrosis; NG-Nitroarginine Methyl Ester; Nitric Oxide; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Transforming Growth Factor beta

Flavivirus infection as dengue and yellow fever persists as a terrible menace to pandemics, due to Aedes prevalence in the Americas. Yellow fever is characterized by hepatocyte damage, with steatosis, apoptosis and necrosis, mainly in the midzonal region of the liver, but the injury mechanism has not been studied at the light of recent knowledge, such as the advances in cell death mechanisms, inflammatory response and cytokine cell expression tools. We studied 53 human liver paraffin embedded blocks from patients who died with yellow fever, all with histological demonstration of higher prevalence of apoptosis over necrosis and mild disproportionate inflammatory response. Viral antigens were found most frequently in hepatocytes from the midzonal area than other lobule areas, as detected by specific immunohistochemistry. Infiltrating cell subpopulations showed mainly CD4+ T lymphocytes, with small numbers of CD8+ cytotoxic lymphocytes, CD20+ B lymphocytes, NKT+ cells and S100+ dendritic cells in the sites of inflammation, as compared to normal and leptospirosis liver blocks. Some cells expressed TNF-alpha and IFN-gamma, but a much more intense proportion of TGF-beta expressing cells were found, suggesting both a Th1 and Th3 patterns of immune response in yellow fever. Most affected hepatocyte presented apoptosis markers that appear at the cell death main pathway in this infection. Viral antigens, which production could interfere in hepatocyte biology, could induce the activation of apoptosis cascade, but TGF-beta was also an apoptosis promoter. Our finding supports the key effect of the yellow fever virus in hepatocyte injury, resulting in prevalence of apoptosis over necrosis, aside from a TGF-beta action induced by the inflammatory response.

Topics: Adolescent; Adult; Aged; Antigens, Viral; Apoptosis; Child; Child, Preschool; Dendritic Cells; Female; Hepatocytes; Histocytochemistry; Humans; Immunohistochemistry; Inflammation; Killer Cells, Natural; Lymphocyte Subsets; Male; Middle Aged; Necrosis; Paraffin Embedding; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Yellow Fever; Yellow fever virus

Myofibroblasts and cytokines such as transforming growth factor-beta1 (TGF-beta1) and platelet-derived growth factor (PDGF)-B have been found to play an important role in pancreatitis-associated fibrogenesis. It is still unclear, however, where in the inflamed pancreas and when these fibrogenic cells and cytokines can be detected. In this study we examined pancreatic tissue from patients with alcoholic chronic pancreatitis to determine the localization and distribution of myofibroblasts and the expression of cytokines in relation to the tissue damage and the activity of the inflammatory process. In tissue from pancreatic specimens from 59 patients with alcoholic chronic pancreatitis the inflammatory process was histologically staged. Myofibroblasts and the cytokines latency-associated peptide, a TGF-beta propeptide, TGF-beta receptor II, PDGF-B and the alpha-isoform of the PDGF receptor were immunohistochemically identified in 10 selected cases representing the four defined stages of alcoholic chronic pancreatitis. In stage I, the stage with overt tissue injury, myofibroblasts were numerous and especially associated with macrophages around areas of necrosis. In stage II, the stage with cellular fibrosis, myofibroblasts were the main component of the interlobular tissue. In stage III, the stage with dense fibrosis, myofibroblasts were rare, and in stage IV, when calculi were present, myofibroblasts were only detected adjacent to duct ulcerations caused by calculi. Latency-associated peptide and TGF-beta receptor II as well as PDGF-B and PDGF receptor-alpha were mainly expressed by macrophages, myofibroblasts and epithelial cells in stages I and II. The results suggest that the fibrogenic process in alcoholic chronic pancreatitis is initiated by a cytokine-based interplay of macrophages and myofibroblasts that follows tissue injury.

Topics: Adult; Aged; Biomarkers; Cytokines; Female; Fibroblasts; Fibrosis; Humans; Immunoenzyme Techniques; Macrophages; Male; Middle Aged; Necrosis; Pancreatic Ducts; Pancreatitis, Alcoholic; Protein Isoforms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-sis; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta; Transforming Growth Factor beta1

TGF-beta1 is a powerful regulator of various T-cell functions. However, it has been unclear how the T-cell responsiveness towards TGF-beta1 is affected by its phenotype or signaling intensity. In the present study, we demonstrate that the phenotype and the TCR-signaling intensity of the responding T-cell as well as the presence of anti-CD28 co-stimulation markedly affects how naïve human cord blood T-cells respond to TGF-beta1. In this report we demonstrate that the strength of the stimulatory signal modifies the T-cell response towards TGF-beta1. Thus, the greatest anti-proliferative effect of TGF-beta1 was observed during weak stimulatory conditions (low dose of anti-CD3 with no co-stimulatory signal). However, such anti-proliferative effect was reduced during strong stimulatory signal (high dose of anti-CD3 with a CD28-directed co-stimulatory signal). In addition, our results indicate that CD8+ T-cells are generally more responsive towards TGF-beta1 than CD4+ T-cells. To our surprise, naïve T-cells had a skewed Th1/Tc1 cytokine secretion pattern with high amounts of IL-2, IFNgamma and TNFalpha, but low amounts of IL-4, IL-5 and IL-10. TGFbeta1 significantly reduced the secretion of IL-2 and IFNgamma, but such suppression was partially prevented by anti-CD28-induced co-stimulation. In contrast, the inhibitory effect on IL-5 secretion was unaffected by anti-CD28 co-stimulation. Interestingly, TGF-beta1 induced IL-10 and TNFalpha secretion. However, the induction of IL-10 secretion was reduced during optimal stimulatory conditions while TGF-beta1 further induced TNFalpha secretion. These data demonstrate that the duration, intensity and type of signaling alters the sensitivity of T-cells to powerful immunological modifying agents like TGF-beta1.

Topics: CD28 Antigens; CD3 Complex; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Survival; Cytokines; Humans; Lymphocyte Activation; Necrosis; Receptors, Antigen, T-Cell; Transforming Growth Factor beta; Transforming Growth Factor beta1

The SAMP1/Sku mouse is a substrain of the SAMP1 (senescence-accelerated-mouse prone 1) which exhibits renal mononuclear cell infiltration from a younger age. We hypothesized that this renal characteristic is related to the incidence of tubulointerstitial nephritis (TIN). The purpose of the present study was to evaluate the applicability of the SAMP1/Sku mouse as a murine model for TIN. TIN was experimentally induced by unilateral ureteral obstruction (UUO). The SAMP1/Sku and control ICR of both sexes received either a sham or UUO operation and were sacrificed 7 days after the operation. The kidneys of the mice were observed histopathologically, immunohistochemically and semiquantitatively. UUO kidneys showed mononuclear cell infiltration, tubular atrophy and interstitial fibrosis. In males, semiquantitative scores of mononuclear cell infiltration, tubular atrophy, and F4/80, alpha-smooth muscle actin (alpha-SMA) and transforming growth factor (TGF)-beta1 reactions were significantly higher in SAMP1/Sku than in ICR. Likewise, in females, tubular atrophy and F4/80 reaction scores were significantly higher in SAMP1/Sku than in ICR. In conclusion, induction of TIN damage by UUO was more serious in SAMP1/Sku mice than in ICR. Therefore, we propose that SAMP1/Sku mice, especially male SAMP1/Sku, have congenital risk factors for the development of TIN.

Topics: Actins; Animals; Antigens, Differentiation; Atrophy; Disease Models, Animal; Female; Fibrosis; Immunohistochemistry; Kidney; Male; Mice; Mice, Inbred Strains; Necrosis; Nephritis, Interstitial; Risk Factors; Transforming Growth Factor beta; Ureteral Obstruction

CD11c+ dendritic cells (DC) and plasmacytoid DC (PDC) are the two major DC subsets in human peripheral blood. For the purpose of immunotherapy with DC, it is important to investigate the phagocytosis of killed tumor cells by different DC subsets. Using immature monocyte-derived DC (iMoDC) as reference, we have compared the ability of CD11c+ DC and PDC to phagocytose apoptotic and necrotic K562 leukemia cells. Freshly isolated CD11c+ DC phagocytosed apoptotic and necrotic K562 cells, whereas PDC did not show any evidence of uptake of dead cells. Blocking studies showed that CD36 is importantly involved in uptake of apoptotic and necrotic material. CD91 and CD11c were also involved. In addition, we found that beta5 integrin was expressed on CD11c+ DC but not in its classical association with alphaV. Uptake of apoptotic K562 cells by CD11c+ DC was increased following incubation with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin (IL)-4, alone or in combination with transforming growth factor-beta1, to levels comparable with those observed for iMoDC. Phagocytosis of dead cellular material by the GM-CSF/IL-4-treated CD11c+ DC was largely restricted to a subset expressing low levels of human leukocyte antigen-DR and CD83. Thus, the relationship between phagocytosis of antigenic material and expression of maturation-related cell-surface molecules is similar for CD11c+ DC and MoDC. We conclude that CD11c+ DC in peripheral blood are precursor cells, which under the influence of cytokines, differentiate to cells with DC phenotype and function.

Topics: Apoptosis; CD11c Antigen; Cell Differentiation; Culture Media, Serum-Free; Dendritic Cells; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Integrin beta Chains; Interleukin-4; K562 Cells; Leukemia, Erythroblastic, Acute; Monocytes; Necrosis; Phagocytosis; Plasma Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1

The peptide hormone relaxin has been demonstrated to exert antifibrotic effects in renal and extrarenal tissues. The aims of this study were to identify potential anti-fibrotic effects of relaxin on human renal fibroblasts in vitro and to analyze their mechanisms.. All experiments were performed in established renal fibroblast cell lines and in primary cortical fibroblasts. Effects of relaxin were analyzed on cell proliferation, apoptosis, activation of renal fibroblasts, synthesis and secretion of collagen type I and fibronectin, as well as on the secretion of matrix metalloproteinases (MMPs). Effects on transforming growth factor-beta1 (TGF-beta1) receptor binding were analyzed by flow cytometry and on TGF-beta1 signal transduction by immunoblot analyses for Smad4 and 7, translocation from cytosol to nucleus for Smad2 and 3 as well as for phosphorylated and unphosphorylated forms of p38, c-Jun NH2 terminal kinase (JNK) and extracellular-regulated protein kinase (ERK). Finally, specific siRNAs for Smad2 and 3 were applied to assess the signal transduction pathway.. After stimulation with relaxin, tyrosine phosphorylation of a 220 kD protein was demonstrated, indicating interaction with the receptor. Relaxin had only modest inhibitory effects on cell proliferation, and no effects on apoptosis. Conversely, relaxin exerted robust effects on TGF-beta1-induced fibroblast to myofibroblast transformation as well as on matrix synthesis and secretion even at the smallest dose tested. The secretion of MMP-2 and MMP-9 was induced noticeably by all investigated relaxin concentrations. TGF-beta1 receptor binding was not influenced by relaxin; however, it prevented Smad2 phosphorylation, translocation to nucleus, and complex formation between Smad2 and 3 indicating a possible interaction with TGF-beta1 signaling. These findings were corroborated by studies using siRNAs to Smad2 and 3 where siRNA to Smad2 but not to Smad3 inhibited the TGF-beta1 induction of fibronectin synthesis. There was no influence of relaxin on intracellular Smad3, Smad4, and Smad7 translocation or phosphorylation of mitogen-activated protein (MAP) kinases.. Relaxin is a potent inhibitor of TGF-beta1-induced extracellular matrix (ECM) synthesis and secretion as well as fibroblast activation. Furthermore, it induces ECM degradation by induction of MMP-2 and MMP-9. These effects are mediated, at least in part, by inhibition of TGF-beta1 signaling.

Topics: Apoptosis; Cell Division; Cell Line; DNA-Binding Proteins; Extracellular Matrix; Fibroblasts; Fibrosis; Humans; Kidney; MAP Kinase Signaling System; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Necrosis; Phosphorylation; Receptors, Transforming Growth Factor beta; Relaxin; Smad2 Protein; Smad3 Protein; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tyrosine

Crescentic nephritis is characterized by formation of cellular crescents that soon become fibrotic and result in irreversible damage, unless an effective immunosuppressive therapy is rapidly commenced. TGF-beta1 is involved in the development of crescents through various pathways. The aim of this study was to identify whether the determination of urinary TGF-beta1 levels in patients with crescentic nephritis could be used as a marker of response to treatment.. Fifteen patients with crescentic nephritis were included in the study. The renal expression of TGF-beta1 was estimated in biopsy sections by immunohistochemistry and urinary TGF-beta1 levels were determined by quantitative sandwich enzyme immunoassay (EIA). TGF-beta1 levels were determined at the time of renal biopsy, before the initiation of immunosuppressive treatment (corticosteroids, cyclophosphamide and plasma exchange). Twelve patients with other types of proliferative glomerulonephritis and ten healthy subjects were used as controls.. Improvement of renal function with immunosuppressive therapy was observed in 6 and stabilization in 4 patients (serum creatinine from 3.2 +/- 1.5 to 1.4 +/- 0.1 mg/dl and from 4.4 +/- 1.2 to 4.1 +/- 0.6 mg/dl, respectively). In 5 patients, with severe impairment of renal function who started on dialysis, no improvement was noted. The main histological feature differentiating these 5 patients from others with improved or stabilized renal function was the percentage patients with poor response to treatment were the percentage of glomeruli with crescents and the presence of ruptured Bowman's capsule and glomerular necrosis. Urinary TGF-beta1 levels were significantly higher in patients who showed no improvement of renal function with immunosuppressive therapy (930 +/- 126 ng/24 h vs. 376 +/- 84 ng/24 h, p < 0.01). TGF-beta1 was identified in crescents and tubular epithelial cells, whereas a significant correlation of TGF-beta1 immunostaining with the presence of fibrocellular cresents was observed (r = 0.531, p < 0,05).. Increased TGF-beta1 renal expression and urinary excretion that is related to the response to immunosuppressive therapy was observed in patients with crescentic nephritis. Evaluation of urinary TGF-beta1 levels may be proved a useful marker of clinical outcome in patients with crescentic nephritis.

Topics: Adult; Aged; Biomarkers; Epithelial Cells; Female; Glomerulonephritis; Humans; Immunohistochemistry; Immunosuppression Therapy; Kidney; Kidney Glomerulus; Kidney Tubules; Male; Middle Aged; Necrosis; Transforming Growth Factor beta; Transforming Growth Factor beta1; Treatment Outcome

Thyroid destruction leading to endemic myxoedematous cretinism is highly prevalent in central Africa, where iodine (I) and selenium (SE) deficiencies as well as thiocyanate (SCN) overload are combined. All three factors have been studied experimentally in the etiology of the disease, but they have never been studied in combination. In a model using rats, we have previously shown that combining I and SE deficiencies increases the sensitivity of the thyroid to necrosis after iodide overload, an event unlikely to occur in the African situation. To develop a model that would more closely fit with the epidemiological findings, we have determined whether an SCN overload would also result in thyroid necrosis as does the I overload. The combination of the three factors increased by 3.5 times the amount of necrotic cells, from 5.5 +/- 0.3% in the I-SE+ thyroids to 18.9 +/- 1.6% in the I-SE-SCN-overloaded ones. Methimazole administration prevented the SCN-induced necrosis. SE- thyroids evolved to fibrosis, whereas SE+ thyroids did not. TGFbeta was prominent in macrophages present in SE- glands. Thyroid destruction in central Africa might therefore originate from the interaction of three factors: I and SE deficiencies by increasing H(2)O(2) accumulation, SE deficiency by decreasing cell defense and promoting fibrosis, and SCN overload by triggering follicular cell necrosis.

Topics: Africa, Central; Animals; Antithyroid Agents; Congenital Hypothyroidism; Disease Models, Animal; Endemic Diseases; Female; Fibrosis; Hydrogen Peroxide; Inflammation; Iodine; Macrophages; Methimazole; Myxedema; Necrosis; Perchlorates; Rats; Rats, Wistar; Selenium; Sodium Compounds; Thiocyanates; Thyroid Gland; Transforming Growth Factor beta

Gene therapy has been recently introduced as a novel approach to treat ischemic tissues by using the angiogenic potential of certain growth factors. We investigated the effect of adenovirus-mediated gene therapy with transforming growth factor-beta (TGF-beta) delivered into the subdermal space to treat ischemically challenged epigastric skin flaps in a rat model.. A pilot study was conducted in a group of 5 animals pretreated with Ad-GFP and expression of green fluorescent protein in the skin flap sections was demonstrated under fluorescence microscopy at 2, 4, and 7 days after the treatment, indicating a successful transfection of the skin flaps following subdermal gene therapy. Next, 30 male Sprague Dawley rats were divided into 3 groups of 10 rats each. An epigastric skin flap model, based solely on the right inferior epigastric vessels, was used as the model in this study. Rats received subdermal injections of adenovirus encoding TGF-beta (Ad-TGF-beta) or green fluorescent protein (Ad-GFP) as treatment control. The third group (n = 10) received saline and served as a control group. A flap measuring 8 x 8 cm was outlined on the abdominal skin extending from the xiphoid process proximally and the pubic region distally, to the anterior axillary lines bilaterally. Just prior to flap elevation, the injections were given subdermally in the left upper corner of the flap. The flap was then sutured back to its bed. Flap viability was evaluated seven days after the initial operation. Digital images of the epigastric flaps were taken and areas of necrotic zones relative to total flap surface area were measured and expressed as percentages by using a software program.. There was a significant increase in mean percent surviving area between the Ad-TGF-beta group and the two other control groups (P < 0.05). (Ad-TGF-beta: 90.3 +/- 4.0% versus Ad-GFP: 82.2 +/- 8.7% and saline group: 82.6 +/- 4.3%.). In this study, the authors were able to demonstrate that adenovirus-mediated gene therapy using TGF-beta ameliorated ischemic necrosis in an epigastric skin flap model, as confirmed by significant reduction in the necrotic zones of the flap. The results of this study raise the possibility of using adenovirus-mediated TGF-beta gene therapy to promote perfusion in random portion of skin flaps, especially in high-risk patients.

Topics: Adenoviridae; Animals; Genetic Therapy; Ischemia; Male; Microscopy, Fluorescence; Models, Animal; Necrosis; Neovascularization, Physiologic; Rats; Rats, Sprague-Dawley; Skin; Surgical Flaps; Transforming Growth Factor beta

The therapeutic efficacy of angiotensin II receptor antagonist, losartan, was studied in patients with nonalcoholic steatohepatitis (NASH). Seven patients with both NASH and hypertension were treated with losartan (50 mg/d) for 48 weeks. Treatment with losartan resulted in a significant decrease in blood markers of hepatic fibrosis, plasma TGF-beta1 and serum ferritin concentration concurrently with an improvement in serum aminotransferase levels. Histological assessment showed improvement of hepatic necroinflammation in five patients, reduction of hepatic fibrosis in four patients, and disappearance of iron deposition in two patients. No side effect of treatment was noted at any time during the study. In conclusion, the present data raise the possibility that an angiotensin II receptor antagonist may be therapeutically efficacious for NASH.

Topics: Adult; Angiotensin Receptor Antagonists; Biomarkers; Fatty Liver; Female; Ferritins; Hepatitis; Humans; Hypertension; Iron; Liver; Liver Cirrhosis; Losartan; Male; Middle Aged; Necrosis; Osmolar Concentration; Transaminases; Transforming Growth Factor beta; Transforming Growth Factor beta1

The present study was conducted to clarify the effect of cell therapy with autologous synovial tissue-derived fibroblasts activated by transforming growth factor (TGF)-beta1 on the necrotized anterior cruciate ligament (ACL). Thirty-six mature female Japanese white rabbits were used in this study. In Group I, fibrin glue with autologous synovial tissue-derived fibroblasts after TGF-beta stimulation was wrapped around the necrotized ACL after freeze-thaw treatment. In Group II, the glue without TGF-beta stimulation was wrapped around the frozen-thawed ACL. In Group III, the fibrin glue without fibroblasts was applied in the same manner on the frozen-thawed ACL. Histological observation found that implantation of fibroblasts after TGF-beta stimulation accelerated cellular infiltration into the ACL following fibroblast necrosis. Biomechanically, the transplantation of synovial tissue-derived autologous fibroblasts activated by TGF-beta inhibited mechanical deterioration of the ACL after the freeze-thaw treatment. The present study has shown that cell-based therapy using synovial tissue-derived fibroblasts activated by TGF-beta1 is a possible solution to mechanical deterioration of the graft after ACL reconstruction.

Topics: Animals; Anterior Cruciate Ligament; Biomechanical Phenomena; Cell Culture Techniques; Cell Transplantation; Female; Fibroblasts; Necrosis; Rabbits; Synovial Membrane; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transplantation, Autologous

The authors investigated the feasibility of thalidomide employed to treat liver fibrosis.. A cirrhotic model was established using Sprague-Dawley rats fed thioacetamide. Thalidomide-treated group was given thalidomide (10mg/kg/day) intraperitoneally for 10 consecutive days. Mortality, histopathological changes, TNFalpha, TGFbeta1, TIMP-1 and TIMP-2 were determined. Expression of TNFalpha and TGFbeta1 mRNA of Kupffer's cells derived from the experimental rats were determined.. The mortality rates of thalidomide-treated group and vehicle-treated group were 8 and 32%, respectively. The total Knodell score of thalidomide-treated rats was lower than those of vehicle-treated rats. Micro-nodular cirrhosis resolved grossly in thalidomide-treated rats on day 28; while vehicle-treated rats continued to display uneven liver surface on day 28. Expression of TNFalpha, TGFbeta1, TIMP-1, and TIMP-2 was decreased in thalidomide-treated rats compared to those treated with vehicles. Finally, the expression of TNFalpha and TGFbeta1 mRNA of Kupffer's cells derived from rats treated with thalidomide were much lower than those treated with vehicle.. Thalidomide salvages lethal hepatic necroinflammation, accelerates recovery from cirrhosis in rats, and works by suppressing of TNFalpha and TGFbeta1 production of Kupffer's cells.

Topics: Animals; Collagenases; Hepatitis; Kupffer Cells; Liver; Liver Cirrhosis, Experimental; Male; Matrix Metalloproteinase 13; Necrosis; Rats; Rats, Sprague-Dawley; Salvage Therapy; Thalidomide; Time Factors; Tissue Inhibitor of Metalloproteinases; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha

Human endothelial cells were exposed to 5 mM glucose (control), 25 mM (high) glucose, or osmotic control for 72 h. TGF-beta1 production, cell growth, death, and cell cycle progression, and the effects of TGF-beta1 and TGF-beta neutralization on these parameters were studied. High glucose and hyperosmolarity increased endothelial TGF-beta1 secretion (P < 0.0001) and bioactivity (P < 0.0001). However, high glucose had a greater effect on reducing endothelial cell number (P < 0.001) and increasing cellular protein content (P < 0.001) than the osmotic control. TGF-beta antibody only reversed the antiproliferative and hypertrophic effects of high glucose. High glucose altered cell cycle progression and cyclin-dependent kinase inhibitor expression independently of hyperosmolarity. High glucose increased endothelial cell apoptosis (P < 0.01), whereas hyperosmolarity induced endothelial cell necrosis (P < 0.001). TGF-beta antibody did not reverse the apoptotic effects observed with high glucose. Exogenous TGF-beta1 mimicked the increased S phase delay but not endoreduplication observed with high glucose. High glucose altered endothelial cell growth, apoptosis, and cell cycle progression. These growth effects occurred principally via a TGF-beta1 autocrine pathway. In contrast, apoptosis and endoreduplication occurred independently of this cytokine and hyperosmolarity.

Topics: Antibodies; Apoptosis; Autocrine Communication; bcl-2-Associated X Protein; Cell Cycle; Cell Cycle Proteins; Cell Division; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins; Diabetes Mellitus; Endothelium, Vascular; Enzyme Inhibitors; Glucose; Humans; L-Lactate Dehydrogenase; Necrosis; Osmolar Concentration; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Suppressor Proteins

The etiology of autoimmune liver disease is poorly understood. BALB/c mice deficient in the immunoregulatory cytokine TGF-beta1 spontaneously develop necroinflammatory liver disease, but the immune basis for the development of this pathology has not been demonstrated. Here, we show that BALB/c-TGF-beta1(-/-) mice exhibit abnormal expansion in hepatic mononuclear cells (MNCs) compared with wild-type littermate control mice, particularly in the T cell and macrophage lineages. To test whether lymphocytes of the adaptive immune system are required for the spontaneous development of necroinflammatory liver disease, BALB/c-TGF-beta1(-/-) mice were rendered deficient in B and T cells by crossing them with BALB/c-recombinase-activating gene 1(-/-) mice. BALB/c-TGF-beta1(-/-)/recombinase-activating gene 1(-/-) double-knockout mice showed extended survival and did not develop necroinflammatory liver disease. The cytolytic activity of BALB/c-TGF-beta1(-/-) hepatic lymphocytes was assessed using an in vitro CTL assay. CTL activity was much higher in BALB/c-TGF-beta1(-/-) hepatic MNCs compared with littermate control hepatic MNCs and was particularly pronounced in the CD4(+) T cell subset. Experimental depletion of CD4(+) T cells in young BALB/c-TGF-beta1(-/-) mice prevented the subsequent development of necroinflammatory liver disease, indicating that CD4(+) T cells are essential for disease pathogenesis in vivo. These data definitively establish an immune-mediated etiology for necroinflammatory liver disease in BALB/c-TGF-beta1(-/-) mice and demonstrate the importance of CD4(+) T cells in disease pathogenesis in vivo. Furthermore, TGF-beta1 has a critical role in homeostatic regulation of the hepatic immune system, inhibiting the development or expansion of hepatic cytolytic CD4(+) T cells.

Topics: Animals; Autoimmune Diseases; CD4-Positive T-Lymphocytes; Crosses, Genetic; Cytotoxicity, Immunologic; Female; Genetic Predisposition to Disease; Hepatitis, Animal; Immunity, Cellular; Lymphocyte Depletion; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Necrosis; T-Lymphocyte Subsets; Transforming Growth Factor beta; Transforming Growth Factor beta1

Thrombospondin-1 is a major activator of transforming growth factor-beta1 (TGF-beta1), and a peptide derived from the latency-associated peptide, Leu-Ser-Lys-Leu (LSKL), inhibits the activation of TGF-beta1. In this study, the effects of LSKL on the hepatocyte damage and fibrogenesis in dimethylnitrosamine (DMN)-induced rat liver fibrosis were examined.. Animals were given an intraperitoneal (i.p.) injection of DMN or saline three times per week for 4 weeks, and treated with LSKL, a control peptide, or saline i.p. daily.. Liver atrophy caused by DMN-injection was significantly inhibited in the DMN+LSKL group. The degrees of necrosis/degeneration and fibrosis scores were significantly lower in the DMN+LSKL group than in the control groups. The hydroxyproline content was significantly higher in the control groups than in the DMN+LSKL group. The amount of active TGF-beta1 was less in the DMN+LSKL group than in the control groups, and the active/total TGF-beta1 ratio in the DMN+LSKL group was suppressed in the control groups. Phosphorylation of Smad 2 in the liver was significantly decreased in the DMN+LSKL group.. The LSKL peptide prevented the progression of hepatic damage and fibrosis through the inhibition of TGF-beta1 activation and its signal transduction in vivo.

Topics: Animals; Atrophy; Dimethylnitrosamine; Disease Progression; DNA-Binding Proteins; Hydroxyproline; Liver; Liver Cirrhosis; Male; Necrosis; Peptides; Phosphorylation; Rats; Rats, Sprague-Dawley; Smad2 Protein; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1

We compared the incorporation of bone allografts with or without vancomycin in tibial defects of 18 pigs. High-quality radiographs, histological examination, immunological expression of metalloproteinase-13 (MMP-13) and transforming growth factor-beta 2 (TGFbeta2) indicated that there was no significant difference in bone allograft incorporation between up to 220 times the MIC (minimum inhibitory concentration) in bone allografts with 1 g of vancomycin in each 300 g of allograft or without this supplement.

Topics: Animals; Anti-Bacterial Agents; Bone Transplantation; Collagenases; Fibrosis; Immunohistochemistry; Male; Matrix Metalloproteinase 13; Necrosis; Swine; Tibia; Transforming Growth Factor beta; Transforming Growth Factor beta2; Vancomycin; Wound Healing

The role of transforming growth factor beta (TGF-beta), a potent regulator of cellular growth, was investigated in the rat model of fulminant hepatic failure (FHF).. The rat FHF model was created by a combination of a 68% partial hepatectomy (PH) and 7% of necrosis (each n = 25 in Groups 1, 2 and 3). Adenovirus mediated gene transfer of mature human TGF-beta1 gene was performed by the systemic injection of AxCAhTGFb1 (1 x 10(9) pfu) in Group 1, 3 days before FHF. In control Groups 2 and 3, recombinant lacZ adenovirus (AxCAlacZ, Group 2) and normal saline (1 ml, Group 3) were used, instead of AxCAhTGFb1.. An excessive expression of TGF-beta1 in Group 1 resulted in an inhibition of hepatocyte proliferation (24-48 h after FHF) and gaining of liver weight (24-48 h), increased expression of HGF in liver tissue (24 h), and decreased expression of TGF-alpha (24 h), compared to those in control Groups 2 and 3. Serum IL-6 levels were also elevated by a TGF-beta1 over-expression at 24 hrs after FHF in Group 1.. The forced expression of TGF-beta1 in the FHF liver yields both a secondary increase of HGF production and a suppression of liver regeneration, which might explain the mechanism of increased serum HGF observed in a clinical FHF. TGF-beta1 is thus thought to have an important role in inhibiting liver regeneration after FHF.

Topics: Adenoviridae; Alanine Transaminase; Animals; Apoptosis; Aspartate Aminotransferases; Body Weight; Cytokines; Disease Models, Animal; Galactosides; Gene Expression; Gene Transfer Techniques; Hepatocyte Growth Factor; Hepatocytes; In Situ Nick-End Labeling; In Vitro Techniques; Indoles; Liver Failure; Liver Regeneration; Male; Necrosis; Organ Size; Proliferating Cell Nuclear Antigen; Rats; Rats, Wistar; Staining and Labeling; Transforming Growth Factor beta; Transforming Growth Factor beta1; Up-Regulation

The aim of this study was to determine whether phagocytosis of necrotic or apoptotic cells affects antigen presentation by murine bone marrow-derived macrophages. After uptake of necrotic neutrophils, macrophages were able to stimulate significantly higher T cell proliferation in vitro against both the recall antigen albumin and the mitogen concanavalin A. No such effect was seen following phagocytosis of apoptotic neutrophils. Flow cytometry revealed that, within 4h of ingestion, macrophages that had taken up the necrotic cells expressed higher levels of CD40 than those that had phagocytosed apoptotic cells. Macrophage cultures pulsed with apoptotic, but not necrotic, neutrophils contained higher levels of transforming growth factor beta1, but lower concentrations of tumour necrosis factor alpha, compared to untreated controls. Our interpretation of these results is that macrophages that have taken up necrotic neutrophils co-stimulate T cells with greater efficiency due to rapid CD40 up-regulation, whereas those that have ingested apoptotic cells are not only ineffective in co-stimulation, but also secrete inhibitory cytokine.

Topics: Animals; Antigen Presentation; Apoptosis; Bone Marrow Cells; CD40 Antigens; Cells, Cultured; Flow Cytometry; Humans; Interferon-gamma; Interleukin-10; Interleukin-4; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred BALB C; Necrosis; Neutrophils; Phagocytosis; T-Lymphocytes; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha; Up-Regulation

The present study was designed to evaluate the distribution of epithelioid cells, myofibroblasts, and TGF-beta1 in the formation of granuloma caused by Mycobacterium avium intracellulare complex (MAC) lung infection. A retrospective study was performed for 9 cases of positive MAC culture in which lung resections were performed between January 1989 and August 1999. Resected lung specimens were evaluated histologically and immunohistochemically for CD68 (stain for monocytes and macrophages, and epithelioid cells) and alpha-smooth muscle actin as well as vimentin (stain for myofibroblasts), and TGF-beta1 was performed. When granuloma was initially formed, no myofibroblasts were found, but as caseous necrosis appeared, the thin epithelioid cell layer was detected and the outer myofibroblast layer gradually became thick. In the cavitary wall, the layer of epithelioid cells and multinucleated giant cells surrounded necrosis, and was associated with the outer layer of myofibroblasts. In addition, the anti-TGF-beta1 antibody stained the cytoplasm of epithelioid cells and multinucleated giant cells, preceding the advent of myofibroblasts. In summary, our present study evaluated distributions of epithelioid cells, myofibroblasts, and TGF-beta along with the morphogenesis of granuloma, and clearly demonstrated the immunohistochemical difference between granuloma with caseous necrosis and granulomas without caseous necrosis.

Topics: Actins; Adult; Aged; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Epithelioid Cells; Female; Fibroblasts; Granuloma; Humans; Immunohistochemistry; Lung; Male; Middle Aged; Mycobacterium avium-intracellulare Infection; Necrosis; Occupational Diseases; Peptide Fragments; Pneumonectomy; Retrospective Studies; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tuberculosis, Pulmonary; Vimentin

A new experimental drug, pirfenidone (5-methyl-1-phenyl-1H-pyridine-one; S-7701), has been reported to have beneficial effects for the treatment of certain fibrotic diseases. We investigated the anti-inflammatory properties in murine endotoxic shock to determine the pharmacological characteristics. The present study describes the prophylactic effect, cytokine regulatory profiles and therapeutic effect of pirfenidone in murine endotoxic shock, which was induced in mice using an intraperitoneal (i.p.) injection of lipopolysaccharide and D-galactosamine. First, we examined the prophylactic effect and cytokine regulatory profiles. A single oral administration of pirfenidone prior to lipopolysaccharide/D-galactosamine challenge inhibited the production of circulating tumor necrosis factor-alpha (TNF-alpha), interleukin-12 and interferon-gamma, markedly enhanced that of interleukin-10, and offered protection from subsequent lethal symptoms in a dose-dependent manner. Second, we examined the therapeutic effect. A single oral administration of pirfenidone 1, 2, 3, 4 and 5 h post lipopolysaccharide/D-galactosamine challenge provided protection against lethal shock in a time-dependent manner. At the histopathological level, apoptotic positive cells were found to be suppressed in the liver. The transforming growth factor (TGF)-beta1 level was markedly elevated in the liver of lipopolysaccharide/D-galactosamine-challenged mice, suppressed in pirfenidone-treated mice. These findings may offer an alternative for both protective and therapeutical treatment of several human acute or chronic inflammatory diseases by pirfenidone.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Galactosamine; Interferon-gamma; Interleukin-10; Interleukin-12; Lipopolysaccharides; Liver; Liver Failure; Mice; Mice, Inbred C57BL; Necrosis; Pyridones; Shock, Septic; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha

Autoimmune hepatitis (AIH) in humans arises spontaneously in genetically susceptible individuals and is associated with the presence of Th1 cells in the liver. The understanding of AIH has advanced more slowly than that of other organ-specific autoimmune diseases, however, largely because of the lack of an appropriate animal model. We now describe a new mouse model characterized by spontaneous development of necroinflammatory hepatitis that is restricted by genetic background. Mice deficient in the immunomodulatory cytokine TGF-beta1 were extensively back-bred to the BALB/c background. The BALB/c background dramatically modified the phenotype of TGF-beta1(-/-) mice: specifically, BALB/c-TGF-beta1(-/-) mice developed a lethal necroinflammatory hepatitis that was not observed in TGF-beta1(-/-) mice on a different genetic background. BALB/c background TGF-beta1(-/-) livers contained large numbers of activated CD4(+) T cells that produced large quantities of IFN-gamma, but little IL-4, identifying them as Th1 cells. BALB/c background TGF-beta1(-/-)/IFN-gamma(-/-) double knockout mice, generated by cross-breeding, did not develop necroinflammatory hepatitis, demonstrating that IFN-gamma is mechanistically required for its pathogenesis. This represents the first murine model of hepatitis that develops spontaneously, is restricted by genetic background, and is dependent upon the Th1 cytokine IFN-gamma, and that thus recapitulates these important aspects of AIH.

Topics: Animals; Autoimmune Diseases; Cell Differentiation; Crosses, Genetic; Genetic Predisposition to Disease; Hepatitis, Animal; Interferon-gamma; Liver; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Necrosis; Survival Rate; Th1 Cells; Th2 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1

Osteotropic growth factors enhance bone repair, but their efficacy in an area of necrotic bone is not known. The purpose of this study was to investigate the effects and potential side effects of an intraosseous application of absorbable bone morphogenetic protein-2 (BMP-2) and growth and differentiation factor-5 (GDF-5) composites in a sheep model for partial necrosis of the femoral head.. The direct injection of ethanol under fluoroscopy into the superior centre of the right femoral head produced histologically documentable necrosis of the central region of the head in a previous study of ten sheep. Another 27 sheep constituted the sample to study the effects of BMP-2 and GDF-5. Necrosis was produced in the same fashion in these animals. Four weeks later nine sheep received 300 microg recombinant BMP-2 and nine sheep 300 microg recombinant GDF-5 on an absorbable carrier by surgical implantation. Nine sheep received the carrier alone (control group). The animals were sacrificed at 3, 6, and 12 weeks after implantation and both femora were harvested.. Bone density analysis and microscopic examination indicated that bone formation was noticeably induced as early as 3 weeks postoperatively in the growth factor treated animals. Bone regeneration was enhanced by growth factor composites. This was documented by histological scoring and histomorphometric analysis. No severe local side effects secondary to the growth factors, such as heterotopic ossification or inflammation, were observed in either group.. The application of an absorbable growth factor composite in combination with established surgical techniques is a promising approach, that may enhance the healing of devitalised bone defects. Based on these results, further studies regarding biodegradation, dosage of the protein and surgical technique are required.

Topics: Animals; Bone and Bones; Bone Density; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Escherichia coli; Ethanol; Growth Differentiation Factor 5; Growth Substances; Humans; Necrosis; Recombinant Proteins; Sheep; Spectrometry, Fluorescence; Time Factors; Transforming Growth Factor beta; Wound Healing

After apoptosis, phagocytes prevent inflammation and tissue damage by the uptake and removal of dead cells. In addition, apoptotic cells evoke an anti-inflammatory response through macrophages. We have previously shown that there is intense lymphocyte apoptosis in an experimental model of Chagas' disease, a debilitating cardiac illness caused by the protozoan Trypanosoma cruzi. Here we show that the interaction of apoptotic, but not necrotic T lymphocytes with macrophages infected with T. cruzi fuels parasite growth in a manner dependent on prostaglandins, transforming growth factor-beta (TGF-beta) and polyamine biosynthesis. We show that the vitronectin receptor is critical, in both apoptotic-cell cytoadherence and the induction of prostaglandin E2/TGF-beta release and ornithine decarboxylase activity in macrophages. A single injection of apoptotic cells in infected mice increases parasitaemia, whereas treatment with cyclooxygenase inhibitors almost completely ablates it in vivo. These results suggest that continual lymphocyte apoptosis and phagocytosis of apoptotic cells by macrophages have a role in parasite persistence in the host, and that cyclooxygenase inhibitors have potential therapeutic application in the control of parasite replication and spread in Chagas' disease.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Cells, Cultured; Chagas Disease; Cysteine Proteinase Inhibitors; Dinoprostone; Macrophages; Male; Mice; Mice, Inbred BALB C; Necrosis; Phagocytosis; Putrescine; Receptors, Vitronectin; T-Lymphocytes; Transforming Growth Factor beta; Trypanosoma cruzi

The dystrophin-deficient mdx mouse presents muscle fiber necrosis but active muscle regeneration, probably due to an extensive recruitment of myogenic regulatory factors (MRF), several growth factors and cytokines, and favorable interaction of satellite cells. In contrast, the laminin alpha 2 (merosin)-deficient dy mouse shows progressive muscle fiber necrosis and ineffective muscle regeneration. Using Western blot and immunohistochemical analyses, we investigated the adaptive changes in MRF, growth factors and cytokines and their receptors in the muscles of dy mice during postnatal growth. The relative volume of MyoD, myogenin and Myf-5 proteins was markedly lower in the gastrocnemius and rectus femoris muscles of dy mice. Transforming growth factor-beta 2, leukemia inhibitory factor (LIF) and basic fibroblast growth factor were not up-regulated in the muscles of dy mice. The levels of the LIF receptor and insulin-like growth factor-I receptor levels were markedly decreased in the muscles of dy mice during the entire postnatal period observed in this study. Therefore, unlike the situation in mdx mice, the milieu of regeneration following repetitive damage seems to be degraded in the muscles of dy mice.

Topics: Aging; Animals; Animals, Newborn; DNA-Binding Proteins; Dystrophin; Growth Inhibitors; Insulin-Like Growth Factor I; Interleukin-6; Leukemia Inhibitory Factor; Leukemia Inhibitory Factor Receptor alpha Subunit; Lymphokines; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Muscle Development; Muscle Fibers, Skeletal; Muscle Proteins; Muscle, Skeletal; MyoD Protein; Myogenic Regulatory Factor 5; Myogenin; Necrosis; Receptor, IGF Type 1; Receptors, Cytokine; Receptors, OSM-LIF; Receptors, Transforming Growth Factor beta; Regeneration; Trans-Activators; Transforming Growth Factor beta

The expression of neuron-type glutamate transporters (EAAC-1), AMPA glutamate receptor subunits (GluR1 and GluR2/3), polyadenosine (5'diphosphate-ribose) polymerase (PARP), and transforming growth factor-beta1 was investigated in 20 cases of neonatal pontosubicular neuron necrosis and 12 gestational-age matched controls. Developmental immunoreactivities of EAAC-1, GluR1, and GluR2/3 appeared in the neurons of the pontine nuclei at 29 to 30 weeks' gestation in controls, and then gradually increased with age. However, these activities were decreased in the pontine nucleus of patients with pontosubicular neuron necrosis. Decreases in these immunoreactivities might indicate early degeneration of neurons. Although PARP and transforming growth factor-beta1 immunoreactivity was insignificant or very weak in the pontine nuclei at any age in controls, PARP was markedly expressed in karyorrhectic neurons of the pontine nucleus in patients with pontosubicular neuron necrosis. Transforming growth factor-beta1 immunoreactivity was observed in nonkaryorrhectic neurons of the pontine nuclei. PARP could contribute to the pathogenesis of pontosubicular neuron necrosis more than EAAC-1 or GluR1 or GluR2/3. Transforming growth factor-beta1 could play a role in the protection and repair of damaged neurons.

Topics: Amino Acid Transport System X-AG; Apoptosis; ATP-Binding Cassette Transporters; Brain Stem Hemorrhage, Traumatic; Case-Control Studies; Enzyme Activation; Female; Gene Expression; Gestational Age; Humans; Immunoblotting; Immunohistochemistry; Infant, Newborn; Infant, Newborn, Diseases; Male; Necrosis; Neurons; Poly(ADP-ribose) Polymerases; Receptors, AMPA; Survival Analysis; Transforming Growth Factor beta

Myostatin, a member of the TGF-beta superfamily, is a key negative regulator of skeletal muscle growth. The role of myostatin during skeletal muscle regeneration has not previously been reported. In the present studies, normal Sprague-Dawley and growth hormone (GH)-deficient (dw/dw) rats were administered the myotoxin, notexin, in the right M. biceps femoris on day 0. The dw/dw rats then received either saline or human-N-methionyl GH (200microg/100g body weight/day) during the ensuing regeneration. Normal and dw/dw M. biceps femoris were dissected on days 1, 2, 3, 5, 9 and 13, formalin-fixed, then immunostained for myostatin protein. Immunostaining for myostatin revealed high levels of protein within necrotic fibres and connective tissue of normal and dw/dw damaged muscles. Regenerating myotubes contained no myostatin at the time of fusion (peak fusion on day 5), and only low levels of myostatin were observed during subsequent myotube enlargement. Fibres which survived assault by notexin (survivor fibres) contained moderate to high myostatin immunostaining initially. The levels in both normal and dw/dw rat survivor fibres decreased on days 2-3, then increased on days 9-13. In dw/dw rats, there was no observed effect of GH administration on the levels of myostatin protein in damaged muscle. The low level of myostatin observed in regenerating myotubes in these studies suggests a negative regulatory role for myostatin in muscle regeneration.

Topics: Animals; Dwarfism; Human Growth Hormone; Humans; Immunohistochemistry; Male; Muscle Fibers, Skeletal; Muscle, Skeletal; Myostatin; Necrosis; Rats; Rats, Mutant Strains; Rats, Sprague-Dawley; Regeneration; Transforming Growth Factor beta

Exposure to oocysts of the protozoan Cryptosporidium parvum causes intestinal epithelial cell dysfunction in vivo and in vitro, but effective means by which mucosal injury might be prevented remain unclear. We examined the ability of transforming growth factor beta1 (TGF-beta1)-a cytokine synthesized and released by cells in the intestine-to preserve the barrier function of human colonic epithelia when challenged with C. parvum oocysts and then studied the mechanisms involved. Epithelial barrier function was monitored electrophysiologically, receptors for TGF-beta1 were localized by confocal microscopy, and TGF-beta1-induced protein kinase C activation was detected intracellularly by translocation of its alpha isozyme. TGF-beta1 alone enhanced intestinal epithelial barrier function, while exposure to C. parvum oocysts (> or =10(5)/monolayer) markedly reduced barrier function to < or =40% of that of the control. When epithelial monolayers were pretreated with TGF-beta1 at 5.0 ng/ml, the barrier-disrupting effect of C. parvum oocysts was almost completely abrogated for 96 h. Further investigation showed that (i) the RI and RII receptors for TGF-beta1 were present on 55 and 65% of human epithelial cell line cells, respectively, over a 1-log-unit range of receptor protein expression, as shown by flow cytometry and confirmed by confocal microscopy; (ii) only basolateral and not apical TGF-beta1 exposure of the polarized epithelial monolayer resulted in a protective effect; and (iii) TGF-beta1 had no direct effect on the organism in reducing its tissue-disruptive effects. In exploring mechanisms to account for the barrier-preserving effects of TGF-beta1 on epithelium, we found that the protein kinase C pathway was activated, as shown by translocation of its 80-kDa alpha isozyme within 30 s of epithelial exposure to TGF-beta1; the permeability of epithelial monolayers to passage of macromolecules was reduced by 42% with TGF-beta1, even in the face of active protozoal infection; and epithelial cell necrosis monitored by lactate dehydrogenase release was decreased by 50% 70 h after oocyst exposure. Changes in epithelial function, initiated through an established set of surface receptors, likely accounts for the remarkable barrier-sparing effect of nanogram-per-milliliter concentrations of TGF-beta1 when human colonic epithelium is exposed to an important human pathogen, C. parvum.

Topics: Animals; Cattle; Cell Line; Cell Membrane; Cell Membrane Permeability; Cell Polarity; Colon; Cryptosporidium parvum; Enzyme Activation; Epithelial Cells; Humans; Intestinal Mucosa; Necrosis; Protein Kinase C; Receptors, Transforming Growth Factor beta; T-Lymphocytes; Transforming Growth Factor beta

C-reactive protein (CRP) is a serum protein that is massively induced as part of the innate immune response to infection and tissue injury. As CRP has been detected in damaged tissues and is known to activate complement, we assessed whether apoptotic lymphocytes bound CRP and determined the effect of binding on innate immunity. CRP bound to apoptotic cells in a Ca(2+)-dependent manner and augmented the classical pathway of complement activation but protected the cells from assembly of the terminal complement components. Furthermore, CRP enhanced opsonization and phagocytosis of apoptotic cells by macrophages associated with the expression of the antiinflammatory cytokine transforming growth factor beta. The antiinflammatory effects of CRP required C1q and factor H and were not effective once cells had become necrotic. These observations demonstrate that CRP and the classical complement components act in concert to promote noninflammatory clearance of apoptotic cells and may help to explain how deficiencies of the classical pathway and certain pentraxins lead to impaired handling of apoptotic cells and increased necrosis with the likelihood of immune response to self.

Topics: Apoptosis; Autoimmunity; C-Reactive Protein; Calcium; Complement C1q; Complement C3b; Complement Factor H; Complement Membrane Attack Complex; Complement Pathway, Classical; Flow Cytometry; Fluorescent Antibody Technique; Humans; Inflammation; Jurkat Cells; Macrophages; Necrosis; Opsonin Proteins; Phagocytosis; Protein Binding; Transforming Growth Factor beta

1. Cytokines are soluble factors whose action has been documented in physiological and pathological conditions. Some may be involved in the pathogenesis of cholestasis, whether of acute or chronic origin. 2. The aim of the present study was to evaluate the influence of epidermal growth factor (EGF), transforming growth factor (TGF)-beta 1, interleukin (IL)-6 and tumour necrosis factor (TNF) on cholestasis. Findings from Sprague-Dawley rats submitted to bile duct ligation for 1-28 days were compared with those from controls, which underwent laparotomy but not bile duct ligation. 3. Biochemical and morphological findings confirmed that the experimental procedure was successful. At the end of each follow-up period, the hepatic levels of the cytokines were determined and compared with liver histology findings. 4. The four cytokines studied showed different patterns of activation: hepatic levels of EGF, higher in the experimental than the control group, were comparable with the proliferative picture. The TGF-beta 1 pattern was correlated with data of periportal, perivenular and perineoductular fibrosis, confirming that this cytokine has a role in mediating the synthesis of matrix proteins. A fluctuating, phasic pattern was found for TNF in the experimental group, with high values on day 0, a decrease on the first and second postoperative days and then two peaks on days 8 and 14. Finally, immediately after surgical manipulation, high levels of IL-6 were found in the experimental group, followed by a decrease in levels until zero values were obtained. 5. This suggests that the obstructive condition produces several cytokine responses, each of which contributes to determine the cholestatic condition.

Topics: Animals; Bile Ducts, Intrahepatic; Cholestasis, Intrahepatic; Cytokines; Disease Progression; Epidermal Growth Factor; Interleukin-6; Ligation; Liver; Male; Necrosis; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Bcl-xL is the predominant anti-apoptotic Bcl-2 family member in the liver. Suppression of cell death promotes carcinogenesis and contributes to resistance to radiation and chemotherapeutic agents.. Direct effects of Bcl-xL protein on apoptosis and necrosis were investigated in rat hepatoma cells. Rat hepatoma cell line McA-RH8994 cells were transfected with expression plasmids containing a whole coding sequence of rat bcl-xL cDNA of sense orientation. Stable transfectant cell lines expressing bcl-xL cDNA (designated as RH8994/Bcl-xL-S), or control plasmid DNA (designated as RH8994/pT) were established.. Cellular amounts of Bcl-xL in RH8994/Bcl-xL-S cells were demonstrated to be more than 20-fold that of RH8994/pT and parental cells. Three independent clones of RH8994/Bcl-xL-S were isolated and their susceptibility to various cell death stimuli was compared with that of the control cells. Transforming growth factor-beta1 and tumour necrosis factor-alpha induced apoptosis dose dependently in these cells, but the 50% cytotoxicity concentrations of these factors in RH8994/Bcl-xL-S cells were more than 10-fold higher than those in RH8994/pT and parental cells. Similarly, RH8994/Bcl-xL-S cells were shown to be significantly less susceptible to necrotic cell death induced by a calcium ionophore, A23187; a mutagen, N-methyl-N'-nitro-N-nitrosoguanidine; and UV-irradiation when compared with the control cells.. Over-expression of Bcl-xL was shown to provide protection against apoptotic and necrotic cell death in rat hepatoma cells.

Topics: Animals; Apoptosis; bcl-X Protein; Calcimycin; Cell Death; Gene Expression; Ionophores; Liver Neoplasms, Experimental; Methylnitronitrosoguanidine; Mutagens; Necrosis; Proto-Oncogene Proteins c-bcl-2; Rats; Transfection; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha; Ultraviolet Rays

A major contributor to the development and progression of ischemia-reperfusion (IR)-induced acute renal failure (ARF) is the loss of functioning tubular epithelial cells by means of various cell deletion or death processes. Although the term "acute tubular necrosis" is still used to describe the pathology of ARF, this is a misnomer because apoptotic cell death, as well as necrosis, occurs [1, 2] along with desquamation and loss of viable epithelial cells [3]. Apoptosis was first described in renal disease in 1987 in an animal model of hydronephrosis [4]. In ARF, with reference to only the death processes, the relative contribution of necrosis or apoptosis possibly depends on the extent of the initiating events. For example, after prolonged total renal ischemia, necrosis or "accidental cell death" occurs from the resultant negation of the cell's energy and protein levels. In apoptosis, the cells use their own energy processes and proteins to die, and often the initiating ischemia is more mild [5]. Finally, despite prolonged ischemia, within the heterogeneous renal cell populations there are those that are more sensitive to ischemia, such as the proximal straight tubule and to some extent the thick ascending limb (TAL) of the loop of Henle. It may be hypothesized that these cells tend to undergo necrosis in comparison with the less sensitive segments that undergo apoptosis. Because apoptosis is gene driven, its identification is important because of the possibility of its modulation via molecular controls. However, despite these new concepts of ARF, patient death remains high, at approximately 30 to 50% of ARF cases. Recovery from ARF depends not only on the replacement or regeneration of cells deleted by death, the theme of many recent studies, but also on protection of cells from death. Both processes are dependent on many of the cellular and molecular controls that have evolved in multicellular organisms to manage normal development, differentiation and growth processes, but that then become involved in the pathogenesis and progression of many renal diseases, including ARF.

Topics: Acute Kidney Injury; Animals; Apoptosis; bcl-X Protein; Body Weight; Cell Division; Cell Survival; Epidermal Growth Factor; Epithelial Cells; Insulin-Like Growth Factor I; Loop of Henle; Male; Necrosis; Proliferating Cell Nuclear Antigen; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Regeneration; Reperfusion Injury; Transforming Growth Factor beta

The outcome of myocardial ischemia-reperfusion has been partially attributed to the degree of apoptosis in cardiomyocytes. Aggregating platelets by release of transforming growth factor-beta(1) (TGF-beta(1)) protect the isolated heart against ischemia-reperfusion injury and preserve myocardial TGF-beta(1) content. To gain more insight into the modulation of hypoxia-reoxygenation-induced injury (apoptosis and necrosis) to myocytes by TGF-beta(1) and aggregating platelets, cultured adult rat myocytes were exposed for 48 or 72 h to hypoxia alone, or to hypoxia followed by 3 h of reoxygenation. Apoptosis in the cells was determined by in situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining and DNA fragmentation on gel electrophoresis. Hypoxia alone caused a time-dependent increase in myocyte apoptosis (number of apoptotic cells: 19+/-3% at 48 h and 39+/-5% at 72 h compared with 5+/-1% in control cells, based on a 500-cell count). Three hours of reoxygenation after 48 h of hypoxia further increased the number of apoptotic cells (34+/-8 versus 19+/-3% in hypoxia for 48 h), but reoxygenation after 72 h of hypoxia did not additionally increase the number of apoptotic cells, perhaps because of extensive cell necrosis on prolonged hypoxia. Forty-eight hours of hypoxia followed by 3 h of reoxygenation also resulted in a decrease in Bcl-2 and an increase in Fas protein level. Incubation of myocytes with either recombinant TGF-beta(1) (0.5-5 ng/ml) or aggregated platelet supernatant (from 2-3 x10(7) platelets/ml, containing approximately 0.5 ng/ml of TGF-beta(1)) markedly (P<.01) decreased the number of apoptotic cells after hypoxia-reoxygenation. Incubation with TGF-beta(1) also reduced myocyte necrosis as evident from lactate dehydrogenase release and trypan blue dye exclusion. These data demonstrate that hypoxia-reoxygenation results in apoptosis and necrosis in cultured adult rat myocytes; this can be attenuated by TGF-beta(1). Similarity of data with TGF-beta(1) and aggregated platelet supernatant suggests that platelet-mediated cardioprotection during hypoxia-reoxygenation may relate in part to the release of TGF-beta(1).

Topics: Animals; Apoptosis; Cells, Cultured; Electrophoresis, Agar Gel; fas Receptor; Genes, bcl-2; Heart Ventricles; In Situ Nick-End Labeling; L-Lactate Dehydrogenase; Necrosis; Platelet Aggregation; Precipitin Tests; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Time Factors; Transforming Growth Factor beta; Trypan Blue

The induction of pathogenic immune responses may be dependent on the immune system receiving 'danger' signals resulting from tissue damage, rather than tolerogenic stimuli associated with normal cell turnover. The aim was to test this hypothesis by comparing the effects of the uptake of necrotic and apoptotic cells on the ability of antigen-presenting cells (APC) to stimulate immune responses in vitro. The experiments focused on presentation by the macrophage, which is the main cell type adapted for clearing cellular debris in vivo. Murine bone marrow-derived macrophages were pulsed with neutrophils that had been rendered apoptotic or necrotic, and tested for the ability to induce T cell responses. The macrophages that had taken up necrotic, but not apoptotic, cells were able to stimulate recall proliferation by ovalbumin-specific T cells. Furthermore, the response to the mitogen concanavalin A (Con A) was more than 6 times higher when macrophages had been pulsed with necrotic, in comparison with apoptotic, cells. In control experiments, macrophages that had not been exposed to dying neutrophils stimulated weak responses to ovalbumin and Con A. To determine why the uptake of apoptotic and necrotic cells exert opposing effects on the ability of macrophages to stimulate T cells, the expression of costimulatory molecules by treated macrophages, and their production of potentially immunomodulatory cytokines were measured. Flow cytometry revealed that macrophages that had taken up necrotic, but not apoptotic, neutrophils expressed increased levels of CD40 compared to untreated controls within 4 h. Macrophages pulsed with apoptotic cells secreted higher levels of transforming growth factor-beta1 than those ingesting necrotic cells or untreated controls. Our interpretation of these results is that macrophages that have taken up necrotic cell debris present antigens to T lymphocytes with greater efficiency due to transient CD40 upregulation, whereas those that have ingested apoptotic cells are ineffective APC since they secrete inhibitory cytokine.

Topics: Animals; Antigen Presentation; Antigens, CD; Apoptosis; Cells, Cultured; Coculture Techniques; Cytotoxicity, Immunologic; Female; Histocompatibility Antigens Class II; Humans; Interferon-gamma; Interleukin-4; Lymphocyte Activation; Macrophage Activation; Macrophages; Mice; Mice, Inbred BALB C; Necrosis; Neutrophils; Phagocytosis; T-Lymphocytes; Transforming Growth Factor beta

The cellular and humoral factors involved in the pathogenesis of glomerulosclerosis and renal fibrosis following a crescentic glomerulonephritis have not been fully elucidated. Myofibroblasts and transforming growth factor-beta (TGF-beta) have been implicated in the development of experimental and clinical renal fibrosis. We have attempted to identify these mediators in crescentic glomerulonephritis and determine their role in the progression of the disease.. We studied retrospectively 21 patients with crescentic and necrotizing glomerulonephritis (CNG) with emphasis on the renal expression (detected by immunohistochemistry) of myofibroblasts (alpha-smooth muscle actin+ cells), TGF-beta and collagen (III and IV) as well as their relationship with the clinical outcome of these patients. In situ hybridization histochemistry was applied to determine the site of synthesis of TGF-beta1 and collagen III. All the patients were treated by immunosuppression and followed up for a median period of 14 months.. Myofibroblasts and TGF-beta were detected in the crescents as well as in the periglomerular and tubulointerstitial areas in CNG biopsies. TGF-beta1 was also detected within renal tubular cells. The percentage of glomeruli with fibrotic and fibrocellular crescents was positively correlated with the severity of Bowman's capsule disruption (r = 0.631, P < 0.01) and with the intensity of myofibroblast expression in the interstitium (r = 0.504, P < 0.05). Strong interstitial immunostain for myofibroblasts and TGF-beta was also noted in association with interstitial fibrosis. In situ hybridization revealed the site of synthesis of TGF-beta1 to be the renal tubular cells of patients with CNG. By contrast, the site of synthesis of collagen III appeared to be confined to interstitial cells surrounding vessels, tubules and the glomeruli in a distribution identical to that of myofibroblasts. There was a significant positive correlation between the number of interstitial alpha-SMA+ cells and both interstitial TGF-beta (r = 0.591, P < 0.01) and interstitial collagen IV (r = 0.588, P < 0.01). In addition, the number of interstitial alpha-SMA+ cells and the extent of immunostain for collagen IV were positively correlated with the final serum creatinine (r = 0.517, P < 0.05 and r = 0.612, P < 0.01 respectively) and partially predicted functional outcome (R2 = 26.7% and 37.5% respectively) as well as the response to treatment. An association was observed between periglomerular myofibroblasts and the generation of fibrotic and fibrocellular crescents.. These observations suggest a causal link between myofibroblasts and fibrotic crescent formation. We also believe that interstitial myofibroblasts are actively involved in the pathogenesis of interstitial fibrosis in CNG.

Topics: Actins; Adolescent; Adult; Aged; Case-Control Studies; Collagen; Female; Fibroblasts; Fibrosis; Glomerulonephritis; Humans; Immunohistochemistry; In Situ Hybridization; Male; Middle Aged; Muscle, Smooth; Necrosis; Retrospective Studies; RNA, Messenger; Transforming Growth Factor beta

Among our cases of IgA glomerulonephritis (IgAGN), 10% show necrotizing/extracapillary lesions involving a small percentage of glomeruli and associated with a certain degree of inflammation in absence of glomerular and interstitial scarring. In our experience, also in repeat biopsies, these cases of IgAGN have a worse prognosis probably because necrotizing/extracapillary lesions can repeat and accumulate, leading to the progression of damage. As it is well known that transforming growth factor-beta (TGF-beta) and endothelin-1 (ET-1) are key-factors in the progression of glomerulonephritis, aim of the study was to examine their expression in renal biopsies of primary IgAGN with necrotizing/crescentic lesions in complete absence of interstitial fibrosis. To obtain information about the mitogenic effect of ET-1, the expression of c-fos, whose upregulation by ET-1 has been established in culture, was also studied.. Eighteen renal biopsies of patients with necrotizing/crescentic IgAGN were examined by immunohistochemistry with antibodies against TGF-beta, ET-1 and c-fos. The results were compared with those obtained on 22 cases of IgAGN characterized only by pure mesangial proliferation and 25 IgAGN biopsies with advanced, not active, glomerulointerstitial lesions.. In necrotizing/crescentic IgAGN glomerular TGF-beta appeared more positive than in cases characterized only by pure mesangial proliferation and was especially expressed on cellular crescents. In the interstitium, TGF-beta, ET-1 and c-fos were expressed by infiltrating leukocytes, tubules, and small vessels. This positivity, although similar as localization, was less diffuse than in biopsies with advanced interstitial damage, but significantly greater than in cases with pure mesangial proliferation.. Positivity of TGF-beta on cellular crescents is similar to that observed from other authors in different types of necrotizing/crescentic human glomerulonephritis and supports our hypothesis that this is a peculiar type of IgAGN. Moreover, interstitial expression of TGF-beta, ET-1 and c-fos in biopsies with glomerular active lesions but complete absence of interstitial fibrosis may potentially represent a signal of activation of mechanisms that induce and amplify the damage leading to further progression of the disease.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Case-Control Studies; Endothelin-1; Female; Glomerular Mesangium; Glomerulonephritis, IGA; Humans; Immunohistochemistry; Kidney Glomerulus; Male; Middle Aged; Necrosis; Proto-Oncogene Proteins c-fos; Transforming Growth Factor beta

The role of the anti-inflammatory cytokine interleukin-10 (IL-10) was investigated in the mouse model of liver injury induced by carbon tetrachloride (CCl4). To address the role of endogenous IL-10 production, acute hepatitis was induced by CCl4 in C57Bl/6 IL-10 gene knock out (KO) and wild-type (WT) mice. After CCl4 challenge, serum and liver levels of tumor necrosis factor-alpha (TNF-) and serum levels of transforming growth factor-beta 1 (TGF-beta1) increased and were significantly higher in IL-10 KO mice, whereas IL-6 serum levels were only slightly increased compared with WT mice. At histological examination, the livers disclosed a significantly more prominent neutrophilic infiltration in IL-10 KO mice 12 and 24 hours after CCl4 injection. In contrast, hepatocyte necrosis, evaluated by histological examination and serum alanine aminotransferase levels, was only marginally affected. The proliferative response of hepatocytes, assessed by the proliferating cell nuclear-antigen labeling index, was significantly increased in IL-10 KO mice, compared with WT mice 48 hours after CCl4 injection. Finally, repeated CCl4 injections led to more liver fibrosis in IL-10 KO mice after 7 weeks. In conclusion, endogenous IL-10 marginally affects the hepatocyte necrosis although it controls the acute inflammatory burst induced by CCl4. During liver repair, it limits the proliferative response of hepatocytes and the development of fibrosis.

Topics: Animals; Carbon Tetrachloride; Cell Division; Cell Movement; Chemical and Drug Induced Liver Injury; Injections; Interleukin-10; Interleukin-6; Liver; Liver Cirrhosis, Experimental; Mice; Mice, Inbred C57BL; Mice, Knockout; Necrosis; Neutrophils; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Using an intracranial rat C6 glioma model, we tested the hypothesis that gene modification of glioma cells to block the expression of the immunosuppressive cytokine TGF-beta (transforming growth factor beta) may enhance anti-tumor immune responses and thereby prolong survival of tumor-bearing animals. The cDNA for simian TGF-beta 2 was ligated in antisense orientation into the episomal plasmid mammalian expression vector pCEP-4. This TGF-beta-antisense vector was transfected into C6 glioma cells by standard electroporation techniques. PCR was used to determine that the rat C6 clones were successfully transfected with the antisense-TGF beta construct. Twenty-nine adult female Wistar rats harboring 7-day-old intracranial C6 tumors were then subcutaneously injected with either saline (n = 9), unmodified C6 glioma cells (n = 10), or TGF-beta-antisense-modified C6 cells (n = 10). Animals were followed for survival, and Fisher's exact method was used to interpret the significance of difference between experimental groups. The survival of tumor-bearing rats injected with TGF-beta-antisense-modified C6 cells was significantly prolonged, relative to the survival of rats receiving injections of saline or unmodified C6 cells alone. Six of the ten (60%) TGF-beta-antisense treated animals survived for 12 weeks, whereas none of the nine (0%) animals treated with saline and none of ten (0%) of those treated with C6 cells alone survived past 5 weeks. These results indicate that the genetic inhibition of immunosuppressive cytokines (such as TGF-beta) may reverse the phenotypic immunosuppression caused by such factors, and thereby prolong the survival of C6 tumor-bearing animals. Future investigations using cytokine gene modifications in other brain tumor models are warranted.

Topics: Animals; Antisense Elements (Genetics); Brain; Brain Neoplasms; Female; Genetic Therapy; Glioma; Necrosis; Phenotype; Polymerase Chain Reaction; Rats; Rats, Wistar; Survival Analysis; Transfection; Transforming Growth Factor beta; Treatment Outcome

Developmentally programmed cell death occurs in several regions of the chick wing bud. We have studied the nature and control of this cell death in vitro in tissues from two of these regions, the posterior necrotic zone (PNZ) and the opaque patch (OP). When tissue from these regions is excised prior to normal cell death and placed into organ culture, cell death ensues. Under these conditions, cell death in tissue from both of these regions is inhibited by fibroblast growth factor-2 (FGF-2). The only other growth factor we have found to have this function is insulin-like growth factor-II. Cell death in tissue from the OP and PNZ occurs by apoptosis, as indicated by the internucleosomal degradation of DNA and the inhibition of cell death by cycloheximide, an inhibitor of protein synthesis. If cell death is inhibited by FGF-2 and then the growth factor is washed away, a compensatory burst of cell death occurs in the PNZ tissue but not the OP tissue. This finding may indicate that in the PNZ, a death program progresses in the face of FGF-2 inhibition, resulting in more cells on the brink of death when the growth factor is removed.

Topics: Animals; Apoptosis; Chick Embryo; Cycloheximide; Epidermal Growth Factor; Fibroblast Growth Factor 2; Growth Substances; In Vitro Techniques; Insulin-Like Growth Factor II; Mesoderm; Microscopy, Electron; Necrosis; Platelet-Derived Growth Factor; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Wings, Animal

It is known that transforming growth factor beta (TGF-beta) is involved in the modulation of cell growth, differentiation, and repair following injury. We performed an immunohistochemical study of human brain autopsy and biopsy material for the expression of TGF-beta isoforms beta 1, beta 2 and beta 3, and TGF-beta receptor (T beta R) type I in different cells of necrotizing lesions such as infarction and abscess, and compared them with controls. Various cell types, both inside and in the proximity of lesions, showed immunoreactivity indicating the presence of all three isoforms. Significant values of immunoreaction for various TGF-beta s and T beta R-I were observed in cells such as astrocytes, macrophages, neurons, microvascular endothelial cells, and granulocytes. In the control cases, comprising biopsy material without necrotizing lesions, a prominent TGF-beta 2 immunoreactivity was observed in glial cells and neurons. TGF-beta 1 and TGF-beta 3 reactivity in controls, when compared with TGF-beta 2, was less. T beta R-I antiserum showed clear and distinct signals in the same type of cells as for TGF-beta s in the necrotizing lesions with varying values of significance. Our findings suggest that TGF-beta s and their receptor type I are involved in reactive processes around necrotizing human brain lesions like glial and macrophage responses, angiogenesis, and deposition of extracellular matrix.

Topics: Adult; Aged; Aged, 80 and over; Brain; Brain Abscess; Brain Chemistry; Cerebral Infarction; Female; Humans; Immunohistochemistry; Male; Middle Aged; Necrosis; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Based on studies that show a role for the low-density lipoprotein (LDL)-receptor in arachidonic acid delivery and eicosanoid synthesis in macrophages, the present study investigated the effect of cholesterol supplementation on pathological changes and thromboxane (TX) synthesis in alcoholic liver injury. Male Wistar rats were intragastrically fed ethanol with either corn oil or fish oil for 1 month. Control rats received isocaloric amounts of dextrose instead of ethanol. An additional group of rats fed either ethanol or dextrose with fish oil or corn oil were supplemented with 1% cholesterol. At the time of killing, all rats had the following evaluated: liver histopathology, lipid peroxidation, liver and plasma thromboxane levels, plasma endotoxin and messenger RNA (mRNA) levels of LDL-receptor, tumor necrosis factor alpha (TNF-alpha), cyclooxygenase (Cox)-1 and -2, and transforming growth factor beta (TGF-beta). Rats fed ethanol with either fish oil or corn oil developed fatty liver, necrosis, inflammation, and central vein collagen deposition. Cholesterol supplementation enhanced the degree of fibrosis but prevented necrosis and inflammation. These alterations in pathological changes by cholesterol were accompanied by absent TNF-alpha and Cox-2 mRNAs, decreased thromboxane levels, decreased lipid peroxidation, and increased TGF-beta mRNA. Cholesterol enrichment of the diet thus decreases proinflammatory components, but enhances fibrosis in ethanol-fed rats.

Topics: Animals; Cholesterol; Inflammation; Lipids; Liver; Liver Cirrhosis; Liver Diseases, Alcoholic; Male; Necrosis; Rats; Rats, Wistar; RNA, Messenger; Thromboxane B2; Transforming Growth Factor beta

We explored the protective activity of glutathione (GSH) in a rat model of carbon tetrachloride-induced acute liver damage. Histological examination of livers from GSH-pretreated rats revealed minor damage, confirmed by biochemical parameters of liver cell necrosis evaluated both 24 and 48 hr after hepatotoxin delivery. In addition we quantified changes in hepatic steady-state levels of albumin, type I procollagen, transforming growth factor-beta 1 and tumour necrosis factor-alpha mRNAs. Even at the molecular level and above all for the albumin gene, it appears that GSH lessens the effect of the hepatotoxin, however the protection of the thiol is restricted to the first 24 hrs and is almost totally exhausted after 48 hr. Since, only 24 hr after CC14 delivery, GSH abundance determined in erythrocytes and liver is almost equal in the controls and in the GSH injected rats, but significantly higher than in the only intoxicated animals (P < 0.05, intraerythrocyte content), we conclude that the thiol pretreatment exerts an effective but transient protection.

Topics: Animals; Biomarkers; Blotting, Northern; Carbon Tetrachloride; Erythrocytes; Glutathione; Liver; Male; Necrosis; Procollagen; Rats; Rats, Sprague-Dawley; RNA, Messenger; Serum Albumin; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

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

Recently, cases of liver damage and liver tumors have been reported after treatment of prostate cancer patients with the antiandrogen cyproterone acetate (CPA). In rat liver, CPA initiates a wave of DNA synthesis that is accompanied by apoptosis. In apoptotic hepatocytes, a latent form of transforming growth factor beta 1 (TGF-beta 1) is detectable by immunohistochemistry. Injection of a single dose of TGF-beta 1 induces apoptosis in the liver of animals pretreated with CPA but has an insignificant effect in untreated animals. In this study, we show by Northern analysis that there is increased expression of TGF-beta 1 in the liver after CPA treatment. Detection of TGF-beta 1 with in situ hybridization showed that TGF-beta 1 was synthesized in the parenchymal cells. Time course and dose-response experiments performed 48 hours after the last application of CPA showed that apoptotic nuclei with chromatin condensed at the nuclear periphery (AN) were already visible 2 hours after injection (0.13%), and apoptotic bodies (ABs) increased 2 to 9 hours after the injection (from 1.28% to 6.67%) after 25 micrograms TGF-beta 1/kg. At 4.5 hours after injection, an induction of apoptosis could be detected with 0.25 microgram TGF-beta 1/kg and after the maximum dose (250 micrograms TGF-beta 1/kg) ANs (0.24%) and ABs (16.74%) were homogeneously distributed throughout the liver lobe. Irrespective of the dose or time after injection of TGF-beta 1, 82% of the ABs were localized within hepatocytes. Liver enzymes were detected in high amounts in the serum (eightfold elevation of glutamate dehydrogenase, fivefold elevation of alanine transaminase [ALT]) 7 hours after the first visible sign of apoptosis. After an additional 20 hours, the liver contained many necrotic figures. These results suggest that the combination of TGF-beta 1 expression coupled with a strikingly enhanced sensitivity to the induction of apoptosis could be responsible both for the liver damage and the development of liver tumors observed after treatment with CPA.

Topics: Androgen Antagonists; Animals; Apoptosis; Body Weight; Cyproterone Acetate; Dose-Response Relationship, Drug; Enzymes; Female; Hyperplasia; Liver; Necrosis; Phagocytosis; Rats; Rats, Wistar; Time Factors; Transforming Growth Factor beta

Incubation of primary cultures of fetal hepatocytes with lipopolysaccharide (LPS) elicited the expression of nitric oxide (NO) synthetase as well as antagonized the apoptotic cell death evoked by treating the cells with transforming growth factor beta 1 (TGF-beta 1). In addition to LPS, exposure of the cells to chemical NO donors also protected against apoptotic cell death when assayed at concentrations in the low micromolar range. Treatment of hepatocytes with large concentrations of NO donors promoted both apoptotic and necrotic cell death. These results suggest that NO synthesis by hepatocytes might be involved in the protection against apoptotic death.

Topics: Animals; Apoptosis; Cells, Cultured; Fetus; Lipopolysaccharides; Liver; Necrosis; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Wistar; Transforming Growth Factor beta

We found that TGF-beta1, a cytokine that previously has been reported to have neuroprotective effects, was able to prevent the toxicity induced by the HIV-1 coat protein gp120 in hippocampal pyramidal neuron cultures. In the presence of glia, gp120 induced time- and dose-dependent cell death, which was more pronounced in mature (7-19 d in culture) than in young neurons (2-7 d in culture). Staining with nuclear dyes (propidium iodide and Hoechst 33342), in situ detection of DNA fragments, and DNA analysis on agarose gels indicated that apoptosis was mainly responsible for the death caused by the viral protein. However, after several days of treatment, death-displaying necrotic features also occurred. Neurotoxicity induced by gp120 was dependent on the activation of NMDA receptors and required the presence of glia as well as new protein synthesis. Thus, the effect of gp120 was abolished by the NMDA receptor antagonist APV and partially reduced by cycloheximide. Only modest neurotoxicity was observed in pure neuronal cultures deprived of the glia feeder layer. Fura-2-based videoimaging showed that treatment with gp120 enhanced the ability of NMDA to increase neuronal [Ca2+]i. The impairment of neuronal Ca2+ homeostasis was prevented completely by TGF-beta1. Therefore, it is likely that the neuroprotective action of the cytokine is attributable to its ability to stabilize neuronal [Ca2+]i.

Topics: Animals; Apoptosis; Calcium; Cell Survival; Cells, Cultured; Hippocampus; HIV Envelope Protein gp120; Intracellular Membranes; Necrosis; Nerve Tissue Proteins; Neuroglia; Neurons; Neuroprotective Agents; Neurotoxins; Rats; Receptors, N-Methyl-D-Aspartate; Transforming Growth Factor beta

Injured skeletal muscle degeneration comprises early microvascular changes and inflammatory cell infiltration, possibly under the control of several growth factors. We have studied the role of basic fibroblast growth factor (bFGF), insulin-like growth factor-1 (IGF1), and transforming growth factor beta-1 (TGF beta 1), by injecting specific anti-growth factor neutralizing antibodies into mouse extensor digitorum longus muscle at the time of injury (denervation and devascularization). Four days later, at the height of damaged myofiber phagocytosis, we assessed quantitatively revascularization, phagocytic activity, and inflammation. The immune neutralization of bFGF reduced the number of capillaries, macrophages and mast cells, and delayed necrotic myofiber phagocytosis. The immune neutralization of IGF1 or TFG beta 1 promoted muscle revascularization, macrophage infiltration and necrotic myofiber phagocytosis. While IGF1 neutralization reduced the number of mast cells and did not modify that of T-cells or neutrophils, TGF beta 1 neutralization increased the number of all of these cells. This study strongly suggests differing roles for bFGF, IGF1 and TFG beta 1 in angiogenic and inflammatory responses during muscle degeneration, apart from their known effects on the behaviour of myogenic cells.

Topics: Animals; Antibodies; Antibody Specificity; Fibroblast Growth Factor 2; Insulin-Like Growth Factor I; Macrophages; Male; Mast Cells; Mice; Muscle Denervation; Muscle, Skeletal; Myositis; Necrosis; Neovascularization, Physiologic; Neutrophils; Phagocytosis; Regeneration; T-Lymphocytes; Transforming Growth Factor beta; Wound Healing

During limb development, the mesenchymal cells in restricted areas of limb bud, anterior necrotic zone, posterior necrotic zone, opaque zone and interdigital necrotic zones, are eliminated by programmed cell death. The transcripts of bone morphogenetic protein (Bmp)-2 and -4 were first detected in the areas where cell death was observed, then showed overlapping expression with the programmed cell death zones except the opaque zone. To investigate the function of BMP-2 and BMP-4 during limb pattern formation, the dominant negative form of BMP receptor was overexpressed in chick leg bud via a replication-competent retrovirus to block the endogenous BMP-2/-4 signaling pathway. This resulted in excess web formation at the anterior and posterior regions of limb buds in addition to marked suppression of the regression of webbing at the interdigital regions. Significant reductions in the number of apoptotic cells in these three necrotic zones were found in the limb buds which received the virus carrying dominant negative BMP receptor. This indicates that extra tissue formation is due to suppression of programmed cell death in the three necrotic zones. Moreover, BMP-2/-4 protein induced apoptosis of mesenchymal cells isolated from the interdigital region in vitro. Other TGFbeta family proteins as TGFbeta1 and Activin did not show this effect. These results suggest that BMP-2 and BMP-4 are the apoptotic signal molecules of the programmed cell death process in the chick limb buds.

Topics: Animals; Apoptosis; Bone and Bones; Bone Morphogenetic Protein 2; Bone Morphogenetic Protein 4; Bone Morphogenetic Protein Receptors; Bone Morphogenetic Proteins; Chick Embryo; Ducks; Gene Transfer Techniques; Limb Buds; Mesoderm; Necrosis; Receptors, Cell Surface; Receptors, Growth Factor; Signal Transduction; Species Specificity; Transforming Growth Factor beta

Xenografted tumours were produced in nude mice by injection of HCT-8 human colon tumour cells. At average volumes of about 750 mm3, animals were injected with fast green vital dye, and 20 min later, tumours were excised and dissected into viable (stained) and necrotic portions (unstained). Viable and necrotic regions were then examined for cell yields, colony forming efficiencies, and levels of basic fibroblast growth factor (FGF-2), transforming growth factors-beta 1 and -alpha (TGF-beta 1, TGF-alpha), platelet derived growth factor (PDGF), and vascular endothelial growth factor (VEGF) using enzyme-linked immunoassay (ELISA) procedures. Levels in the viable and necrotic regions were compared to levels in unseparated tumours. The average extent of necrosis in HCT-8 tumours of this size was 64%. The data for cell yields, colony forming efficiencies FGF-2, VEGF, TGF-beta 1 and TGF-alpha indicated that values determined in the unseparated tumours could be understood on the basis of the weighted average between viable and necrotic tissue, with the higher values occurring in the viable tissue. Low levels of FGF-2 and VEGF were found in the necrotic portions of the tumour while no measurable levels of TGF-beta 1 and TGF-alpha could be determined. PDGF levels were, however, equivalent in both the viable and necrotic regions indicating that necrotic tissue could be an important reservoir for this growth factor.

Topics: Colonic Neoplasms; Endothelial Growth Factors; Enzyme-Linked Immunosorbent Assay; Fibroblast Growth Factor 2; Growth Substances; Humans; Lymphokines; Necrosis; Neoplasms, Experimental; Platelet-Derived Growth Factor; Tissue Distribution; Transforming Growth Factor alpha; Transforming Growth Factor beta; Transplantation, Heterologous; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Hepatocyte growth factor (HGF), a potent hepatocyte mitogen in vitro, triggers hepatocyte regeneration after partial hepatectomy and acute liver cell necrosis induced by chemicals. In contrast, transforming growth factor beta 1 inhibits hepatocyte proliferation in vitro and suppresses liver regeneration in vivo. We assessed the expression of HGF and TGF beta 1 mRNA in an endotoxin-related hepatic cell necrosis model. Intravenous injection of Gram-negative lipopolysaccharide (LPS) into rats previously given heat-killed Propionibacterium acnes induced endotoxin-related hepatic cell necrosis. In this model, serum ALT began to rise to more than 100IU as early as 3 h after LPS injection, reaching 300IU 12h after injection. HGF mRNA levels in the liver did not increase significantly until 5h after LPS injection; at 12h, they had increased about threefold compared with controls. TGF beta 1 mRNA expression increased threefold after P. acnes treatment alone and increased further after LPS injection. In the spleen, HGF mRNA levels increased within 3h, but in the lung no increase in HGF mRNA was observed. Early elevation of liver TGF beta 1 mRNA levels and delayed elevation of HGF mRNA levels, with low expression of HGF in the lung, may play a role in the pathogenesis of endotoxin-related hepatic necrosis.

Topics: Alanine Transaminase; Animals; Blotting, Northern; Disease Models, Animal; Endotoxins; Hepatocyte Growth Factor; Lipopolysaccharides; Liver; Lung; Male; Necrosis; Propionibacterium acnes; Rats; Rats, Wistar; RNA, Messenger; Spleen; Transforming Growth Factor beta

Peptide growth factors are potent regulators of osteoblast differentiation, proliferation, and maturation. Two of these growth factors, transforming growth factor beta (TGF-beta) and basic fibroblast growth factor (bFGF), were used in an attempt to stimulate osteoneogenesis and angiogenesis in a molded vascularized bone graft in the rat. Custom chambers containing cancellous autograft bone and agarose beads and incubated with TGF-beta, bFGF, or a control solution, were closed around the femoral artery/vein pedicle for 2-4 weeks. Control grafts were completely necrotic and without mechanical integrity. TGF-beta grafts demonstrated active osteogenesis around necrotic bone, osteoclastic activity, and limited angiogenesis. Basic FGF grafts demonstrated substantial angiogenesis with limited osteoblastic activity. This study suggests that TGF-beta and bFGF stimulate populations of cells in the formation of a molded vascularized bone graft. TGF-beta induces the proliferation and/or activity of osteoblastic cells, while bFGF stimulates cells involved in angiogenesis. Despite these findings, an insoluble demineralized matrix component may be required for complete transformation and graft consolidation.

Topics: Animals; Blood Vessels; Bone Transplantation; Fibroblast Growth Factor 2; Necrosis; Osseointegration; Rats; Rats, Inbred Lew; Transforming Growth Factor beta; Transplantation, Autologous

The cytotoxicity of transforming growth factor beta 1 (TGF beta 1) was assessed in rat hepatocytes cultured under periportal (PP)-or pericentral (PC)-equivalent conditions. TGF beta 1 induced a 5-fold greater DNA fragmentation and LDH release in PC cultures than in PP cultures. At low exposure level (1 ng/ml TGF beta 1), albumin secretion and mitochondrial activity (rhodamine-123 uptake) were selectively reduced in PP cultures, whereas the incidence of apoptotic cells in PC cultures was about 10-fold higher than that in PP cultures. The time profiles of TGF beta 1-induced apoptotic and necrotic events and the concentration-response relationship differed in PC and PP cultures. In PC cultures the early appearance of cells with apoptotic nuclei was not associated with DNA fragmentation nor with an increase in LDH release or impaired mitochondrial function. At a high exposure level (5 ng/ml TGF beta 1), again cells with apoptotic nuclei were much more strongly induced in PC cultures but DNA fragmentation, LDH release, and impairment of mitochondrial activity all increased in an exposure-time dependent manner in both PP and PC cultures. At this exposure level 48 and 72% of the apoptotic cells detected in PC cultures after continuous exposure for 24 hr were induced within an exposure of 1 and 4 hr, respectively. Aurintricarboxylic acid (50 microM), an inhibitor of endonucleases, significantly inhibited the appearance of apoptotic cells and the progression in apoptosis. Clearly, TGF beta 1 preferentially induced apoptotic cell death in hepatocytes with PC-equivalent metabolism at low exposure levels. High exposure levels or prolonged exposure periods produced both apoptosis and necrosis.

Topics: Animals; Apoptosis; Cell Survival; Cells, Cultured; Culture Techniques; Dose-Response Relationship, Drug; L-Lactate Dehydrogenase; Liver; Male; Necrosis; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

The effect of an 80 to 90% hepatectomy in stimulating proliferation immediately after irradiation of the liver was studied. A dose of 15 Gy was not lethal for animals with intact livers, but all animals with subtotal hepatectomies exposed to this dose died from apparent liver failure 28 to 60 days after exposure. To elucidate the mechanism for this mortality, plasma aspartate aminotransferase, retention of intravenous injected rose bengal, liver weight and liver hydroxyproline content were measured 0 to 90 days after 15 Gy irradiation of the liver to determine temporal changes in necrosis, function, mass and fibrosis, respectively, in animals with either intact livers or livers with subtotal resection. Irradiation of the liver had no significant effect on these parameters in animals with intact livers. In subtotally hepatectomized animals the same radiation dose that suppressed liver mass restoration significantly increased hepatocyte necrosis within 7 days, which was followed by increased liver hydroxyproline concentration and hepatic dysfunction. This radiation-induced temporal change in hepatic dysfunction correlated with increased concentration of hydroxyproline but not with liver mass, indicating that liver fibrosis was the cause of hepatic dysfunction. Since similar sequelae are produced in intact livers after higher doses and longer intervals after irradiation, the proliferation stimulus induced by partial hepatectomy must accelerate the expression of damage and lower the radiation tolerance of the liver. However, in subtotally hepatectomized animals radiation-induced hepatocyte necrosis precedes fibrosis, whereas the reverse is normally true for animals with intact livers.

Topics: Animals; Aspartate Aminotransferases; Cell Division; Dose-Response Relationship, Radiation; Hepatectomy; Hydroxyproline; Liver; Liver Regeneration; Male; Necrosis; Rats; Transforming Growth Factor beta

We studied the effects of transforming growth factor beta 1 (TGF-beta 1) in a feline model of myocardial ischemia (1.5 hr) and reperfusion (4.5 hr). Myocardial ischemia followed by reperfusion resulted in severe myocardial injury, endothelial dysfunction, high cardiac myeloperoxidase activity indicative of neutrophil accumulation in the ischemic myocardium, and significant neutrophil adherence to the ischemic coronary endothelium. In contrast, intravenous administration of TGF-beta 1 (20 micrograms/kg) 30 min prior to reperfusion significantly attenuated myocardial necrosis (13.8% +/- 3.5% vs. 32.2% +/- 2.9% of area-at-risk, P < 0.01) and attenuated endothelial dysfunction (P < 0.01) associated with ischemia-reperfusion. Moreover, myeloperoxidase activity in the ischemic myocardium was significantly lower than vehicle controls (0.2 +/- 0.1 vs. 1.7 +/- 0.3 units/100 mg of tissue, P < 0.01) and neutrophil adherence to ischemic coronary endothelium was significantly (P < 0.01) attenuated in TGF-beta 1-treated cats. These results demonstrate that TGF-beta 1 exerts a significant cardioprotective effect in a feline model of myocardial ischemia and reperfusion. The mechanism of this protective effect appears to relate to endothelial preservation by TGF-beta 1 inhibiting circulating neutrophils from adhering to the endothelium, a critical step in neutrophil-induced reperfusion injury.

Topics: Animals; Cats; Cell Adhesion; Electrophysiology; Endothelium, Vascular; Hemodynamics; Injections, Intravenous; Male; Myocardial Reperfusion Injury; Myocardium; Necrosis; Neutrophils; Peroxidase; Transforming Growth Factor beta

Cell death by apoptosis is a major determinant of growth of normal tissues and tumours. The present study aimed to elucidate signal factors involved in its regulation. Epithelial cells in control liver, during regression of cyproterone acetate induced liver hyperplasia, in liver (pre)neoplasia and in uterus undergoing apoptosis in vivo show immunostaining for transforming growth factor beta 1 (TGF-beta 1) as detected by anti-pre(266-278) TGF-beta 1 antibodies. Positive immunostaining is also seen in a few intact cells of hyperplastic, regressing liver apparently preparing for apoptosis, but is virtually not found in hepatocytes of normal or growing liver nor in cells undergoing death by necrosis. Recombinant latency associated protein (rLAP, dimer of the pro-region non-covalently associated with the mature region) complex and mature TGF-beta 1 induce apoptosis in isolated hepatocytes cultured in vitro. These findings suggest an involvement of TGF-beta 1 in the induction of apoptosis in certain epithelia in vivo.

Topics: Animals; Apoptosis; Biomarkers; Cell Death; Cyproterone Acetate; Female; Hyperplasia; Liver; Necrosis; Rats; Transforming Growth Factor beta

Transforming growth factor-beta 1 (TGF-beta 1) is a multifunctional growth factor that has profound regulatory effects on many developmental and physiological processes. Disruption of the TGF-beta 1 gene by homologous recombination in murine embryonic stem cells enables mice to be generated that carry the disrupted allele. Animals homozygous for the mutated TGF-beta 1 allele show no gross developmental abnormalities, but about 20 days after birth they succumb to a wasting syndrome accompanied by a multifocal, mixed inflammatory cell response and tissue necrosis, leading to organ failure and death. TGF-beta 1-deficient mice may be valuable models for human immune and inflammatory disorders, including autoimmune diseases, transplant rejection and graft versus host reactions.

Topics: Animals; Base Sequence; Cytokines; Gene Expression; Genes; Homozygote; Inflammation; Leukocyte Count; Mice; Mice, Transgenic; Molecular Sequence Data; Mutagenesis, Insertional; Necrosis; Oligodeoxyribonucleotides; Polymerase Chain Reaction; Restriction Mapping; RNA, Messenger; Transforming Growth Factor beta