concanavalin-a and Liver-Failure--Acute

Acute liver failure can be the consequence of various etiologies, with most cases arising from drug-induced hepatotoxicity in Western countries. Despite advances in this field, the management of acute liver failure continues to be one of the most challenging problems in clinical medicine. The availability of adequate experimental models is of crucial importance to provide a better understanding of this condition and to allow identification of novel drug targets, testing the efficacy of new therapeutic interventions and acting as models for assessing mechanisms of toxicity. Experimental models of hepatotoxicity related to acute liver failure rely on surgical procedures, chemical exposure or viral infection. Each of these models has a number of strengths and weaknesses. This paper specifically reviews commonly used chemical in vivo and in vitro models of hepatotoxicity associated with acute liver failure.

Topics: Acetaminophen; Animals; Apoptosis; Chemical and Drug Induced Liver Injury; Concanavalin A; Disease Models, Animal; Fas Ligand Protein; Galactosamine; Humans; Liver Failure, Acute

Acute liver failure (ALF) can be the consequence of various etiologies, which immune response plays a pivotal role in the pathogenesis. For the diversity of etiologies, more animal models are still needed in this field. Here, we developed a new acute liver injury mouse model induced by a fungal lectin AAGL (Agrocybe aegerita galectin). Intravenous injection of AAGL could induce the infiltration and activation of T, NKT and NK cells in liver and T cell played an important role in the pathogenesis. However, compared with the widely used concanavalin A model, AAGL model showed different immune mechanism. Transcriptome analysis of live tissue suggested that inflammation mediated by chemokine and cytokine signaling pathway was different between AAGL and Con A model. Fluorescent quantitative PCR verification assay showed that IL-1β was expressed much higher in AAGL-treated mice and anti-IL-1β could ameliorate AAGL-induced liver injury by inhibiting NF-κB and p38 signaling pathway. The expression of CXCL9 which was responsible for T cell infiltration in liver was also inhibited in AAGL model. We found a critical role of IL-1β in the pathogenesis of AAGL model through recruiting T cells to liver, which highlighted that IL-1β antibody might be a candidate therapy for ALF.

Topics: Agrocybe; Animals; Antibodies; Cell Movement; Concanavalin A; Galectins; Interleukin-1beta; Liver; Liver Failure, Acute; Mice; T-Lymphocytes

Amphiregulin (Areg), a glycoprotein from the epidermal growth factor receptor (EGFR) ligand family, has a well-documented protective role against tissue injury; however, its effects on immune-mediated liver injury are still unclear. Here, we used a concanavalin A (ConA)-induced acute liver hepatitis model to explore the effects of Areg on immune-mediated acute liver injury.. Some C57BL/6 mice were administered ConA at a dose of 20 mg/kg (model mice), and some received 5 µg of Areg (treated mice). Then, their survival rates over 36 h were analyzed. After 5 h of treatment, liver function, hepatic histology, and apoptosis in liver tissue were investigated, and cytokine expression and neutrophil infiltration and activity in the liver were detected. Moreover, the protective effects of Areg were also evaluated without IL-22. Our results showed that Areg administration increased acute liver failure (ALF) mouse survival, restored liver function, and alleviated liver damage. Interestingly, Areg administration increased IL-22 production in hepatic T cells and upregulated IL-22 concentrations in the serum and liver, whereas IL-22 neutralization completely abolished the therapeutic effect of Areg. Meanwhile, Areg administration was concomitant with increased expression of the anti-apoptotic proteins Bcl-2 and Bcl-xL, which are important in the hepatoprotective mechanism of IL-22.. Areg showed direct protective effects against ConA-induced acute liver injury, which suggests the potential therapeutic application of Areg in immune-mediated ALF.

Topics: Amphiregulin; Animals; Apoptosis; bcl-X Protein; Chemical and Drug Induced Liver Injury; Concanavalin A; Disease Models, Animal; Interleukin-22; Interleukins; Liver; Liver Failure, Acute; Mice, Inbred C57BL; Phosphorylation; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; STAT3 Transcription Factor; T-Lymphocytes

Human liver or hepatocyte transplantation is limited by a severe shortage of donor organs. Direct reprogramming of other adult cells into hepatic cells may offer a solution to this problem. In a previous study, we have generated hepatocyte-like cells from mouse fibroblasts using only one transcription factor (TF) plus a chemical cocktail. Here, we show that human urine-derived epithelial-like cells (hUCs) can also be transdifferentiated into human hepatocyte-like cells (hiHeps) using one TF (Foxa3, Hnf1α, or Hnf4α) plus the same chemical cocktail CRVPTD (C, CHIR99021; R, RepSox; V, VPA; P, Parnate; T, TTNPB; and D, Dznep). These hiHeps express multiple hepatocyte-specific genes and display functions characteristic of mature hepatocytes. With the introduction of the large T antigen, these hiHeps can be expanded in vitro and can restore liver function in mice with concanavalin-A-induced acute liver failure. Our study provides a strategy to generate functional hepatocyte-like cells from hUCs by using a single TF plus a chemical cocktail.

Topics: Animals; Cellular Reprogramming; Cellular Reprogramming Techniques; Concanavalin A; Epithelial Cells; HEK293 Cells; Hepatocyte Nuclear Factor 1-alpha; Hepatocyte Nuclear Factor 3-gamma; Hepatocyte Nuclear Factor 4; Hepatocytes; Humans; Liver Failure, Acute; Male; Mice; Transfection; Urine; Young Adult

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Cell Line, Tumor; Concanavalin A; Cytokines; Hep G2 Cells; Humans; Inflammation; Liver Failure, Acute; Macaca fascicularis; Male; Mice; Mice, Inbred C57BL; Rats; Rats, Sprague-Dawley; Receptors, TNF-Related Apoptosis-Inducing Ligand; TNF-Related Apoptosis-Inducing Ligand

Interleukin 35 (IL-35) is a relatively newly identified cytokine required for the regulatory and suppressive functions of regulatory T cells (Treg), playing an important role in the prevention of autoimmune diseases. This study used mesenchymal stem cells (MSCs) as the gene-delivery vehicles for IL-35 gene therapy and investigated their protective effects in Concanavalin A (Con A)-induced autoimmune hepatitis. Results showed that IL-35 gene modified MSCs (IL-35-MSCs) can specifically migrate to the injured liver tissues and significantly narrow the necrosis areas of injured livers. IL-35-MSCs prevented hepatocyte apoptosis by reducing the FASL expression by mononuclear cells. Although MSC treatment can alleviate liver injury to some extent, IL-35-MSCs showed a stronger protective effect, which means some novel mechanisms exist. The results show that IL-35-MSCs could decrease the level of interferon gamma secreted by liver mononuclear cells through the JAK1-STAT1/STAT4 signal pathway. In summary, this study thus demonstrates a novel and efficient treatment for Con A-induced fulminant hepatitis through negatively regulating the secretion of interferon gamma, thus providing a novel therapeutic approach for this devastating liver disease.

Topics: Animals; Apoptosis; Concanavalin A; Fas Ligand Protein; Gene Expression Regulation; Gene Transfer Techniques; Hepatitis, Autoimmune; Hepatocytes; Humans; Interferon-gamma; Interleukins; Janus Kinase 1; Liver; Liver Failure, Acute; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; T-Lymphocytes, Regulatory

Fasiglifam (TAK-875), a G protein-coupled receptor 40 (GPR40) agonist, was a drug candidate for type 2 diabetes. However, its development was terminated in phase 3 trials due to liver safety concerns. Although TAK-875 was reported to inhibit hepatobiliary transporters and disturb bile acid disposition, pathogenic mechanisms of TAK-875-induced liver injury are not fully understood. In this study, we sought to identify the mechanisms with a hepatic genome-wide transcriptomic analysis in a murine model. We demonstrated that, among the three GPR40 agonists, TAK-875, AMG-837, and TUG-770, only TAK-875 induced acute liver injury in mice. Transcriptome profiles of TAK-875-exposed liver was compared with those of non-hepatotoxic analogues AMG-837 and TUG-770 as negative controls and those of classical hepatotoxicants concanavalin A and carbon tetrachloride as positive controls. The comparative hepatic transcriptome analyses revealed the enrichment of genes involved in inflammation, endoplasmic reticulum (ER) stress, apoptosis, and hepatic lipid accumulation, suggesting that these events play pathophysiologic roles in the development of TAK-875-induced liver injury. These results were validated by quantitative PCR with significant changes in chemokines, danger signals, ER stress mediators, proapoptotic factors, and hepatic steatosis markers only in TAK-875-exposed liver. Pretreatment of TAK-875-administered mice with an ER stress inhibitor 4-phenylbutyric acid (4-PBA) alleviated the liver injury. Consistent with the in vivo study, pretreatment of HepG2 cells with 4-PBA significantly improved the decrease of cell viability induced by TAK-875. In conclusion, by a comprehensive transcriptomic analysis, we found multiple possible processes that contribute to TAK-875-induced acute liver injury in mice.

Topics: Animals; Benzofurans; Carbon Tetrachloride; Chemical and Drug Induced Liver Injury; Concanavalin A; Female; Liver; Liver Failure, Acute; Mice; Mice, Inbred ICR; Molecular Structure; Sulfones; Transcriptome

Fulminant hepatitis is a severe liver disease resulting in hepatocyte necrosis. Galectin‑9 (Gal‑9) is a tandem‑repeat‑type galectin that has been evaluated as a potential therapeutic agent for various diseases that regulate the host immune system. Concanavalin A (ConA) injection into mice results in serious, immune‑mediated liver injury similar to human viral, autoimmune and fulminant hepatitis. The present study investigated the effects of Gal‑9 treatment on fulminant hepatitis in vivo and the effect on the expression of microRNAs (miRNAs), in order to identify specific miRNAs associated with the immune effects of Gal‑9. A ConA‑induced mouse hepatitis model was used to investigate the effects of Gal‑9 treatment on overall survival rates, liver enzymes, histopathology and miRNA expression levels. Histological analyses, TUNEL assay, immunohistochemistry and miRNA expression characterization, were used to investigate the degree of necrosis, fibrosis, apoptosis and infiltration of neutrophils and macrophages. Overall survival rates following ConA administration were significantly higher in Gal‑9‑treated mice compared with control mice treated with ConA + PBS. Histological examination revealed that Gal‑9 attenuated hepatocellular damage, reduced local neutrophil infiltration and prevented the local accumulation of macrophages and liver cell apoptosis in ConA‑treated mice. In addition, various miRNAs induced by Gal‑9 may contribute to its anti‑apoptotic, anti‑inflammatory and pro‑proliferative effects on hepatocytes. The results of the present study demonstrate that Gal‑9 may be a candidate therapeutic target for the treatment of fulminant hepatitis.

Topics: Animals; Apoptosis; Concanavalin A; Disease Models, Animal; Galectins; Gene Expression Profiling; Gene Expression Regulation; Hepatocytes; Liver; Liver Failure, Acute; Liver Function Tests; Macrophages; Male; Mice; MicroRNAs; Neutrophil Infiltration; Neutrophils; Survival Rate

Fulminant hepatic failure is a life-threatening disease which occurs in patients without preexisting liver disease. Nowadays, there is no ideal therapeutic tool in the treatment of fulminant hepatic failure. Recent studies suggested that a novel technology termed CRISPR/Cas9 may be a promising approach for the treatment of fulminant hepatic failure. In this project, we have designed single chimeric guide RNAs specifically targeting the genomic regions of mouse Fas gene. The in vitro and in vivo effects of sgRNAs on the production of Fas protein were examined in cultured mouse cells and in a hydrodynamic injection-based mouse model, respectively. The in vivo delivery of CRISPR/Cas9 could maintain liver homeostasis and protect hepatocytes from Fas-mediated cell apoptosis in the fulminant hepatic failure model. Our study indicates the clinical potential of developing the CRISPR/Cas9 system as a novel therapeutic strategy to rescue Concanavalin-A-induced fulminant hepatic failure in the mouse model. J. Cell. Biochem. 118: 530-536, 2017. © 2016 Wiley Periodicals, Inc.

Topics: Animals; Apoptosis; Cell Line; Clustered Regularly Interspaced Short Palindromic Repeats; Concanavalin A; Disease Models, Animal; fas Receptor; Gene Targeting; Hepatocytes; Liver Failure, Acute; Mice; Mice, Inbred ICR

Liver diseases are mostly accompanied by inflammation and hepatocyte death. Therapeutic approaches targeting both hepatocyte injury and inflammation are not available. Natural compounds are considered as potential treatment for inflammatory liver diseases. Hesperetin, a flavonoid component of citrus fruits, has been reported to have anti-inflammatory properties. The aim of this study was to evaluate the cytoprotective and anti-inflammatory properties of hesperetin both in vitro and in models of fulminant hepatitis.. Apoptotic cell death and inflammation were induced in primary cultures of rat hepatocytes by bile acids and cytokine mixture respectively. Apoptosis was quantified by caspase-3 activity and necrosis by LDH release. The concanavalin A (ConA) and D-galactosamine/LPS (D-GalN/LPS) were used as models of fulminant hepatitis. Liver injury was assessed by alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, liver histology and TUNEL assay and inflammation by inducible NOS (iNOS) expression.. Hesperetin blocked bile acid-induced apoptosis and cytokine-induced inflammation in rat hepatocytes. Moreover, hesperetin improved liver histology and protected against hepatocyte injury in ConA- and D-GalN/LPS-induced fulminant hepatitis, as assessed by TUNEL assay and serum AST and ALT levels. Hesperetin also reduced expression of the inflammatory marker iNOS and the expression and serum levels of TNFα and IFN-γ, the main mediators of cell toxicity in fulminant hepatitis.. Hesperetin has anti-inflammatory and cytoprotective actions in models of acute liver toxicity. Hesperetin therefore has therapeutic potential for the treatment of inflammatory liver diseases accompanied by extensive hepatocyte injury, such as fulminant hepatitis.

Topics: Animals; Biological Products; Concanavalin A; Dose-Response Relationship, Drug; Galactosamine; Hesperidin; In Vitro Techniques; Lipopolysaccharides; Liver Failure, Acute; Male; Protective Agents; Rats; Rats, Wistar

Concanavalin A (ConA), a T-cell mitogen that induces acute autoimmune hepatitis, is widely used to model pathophysiological processes of human acute autoimmune liver disease. Although autophagy has been extensively studied in the past decade, little is known about its molecular mechanism underlying the regulation of ConA-induced acute hepatitis. In this study, we used a Cre-conditional atg7 KO mouse to investigate the effects of Atg7-associated autophagy on ConA-induced murine hepatitis. Our results demonstrated that atg7 deficiency in mice enhanced macrophage activation and increased pro-inflammatory cytokines upon ConA stimulation. Atg7 silencing resulted in accumulation of dysfunctional mitochondria, disruption of reactive oxygen species (ROS) degradation, and increase in pro-inflammatory cytokines in Raw264.7 cells. p38/MAPK and NF-κB levels were increased upon ConA induction due to Atg7 deficiency. Blocking ROS production inhibited ConA-induced p38/IκB phosphorylation and subsequent intracellular inflammatory responses. Hence, this study demonstrated that atg7 knockout in mice or Atg7 knockdown in cell culture augmented ConA-induced acute hepatitis and related cellular malfunction, indicating protective effects of Atg7 on regulating mitochondrial ROS via a p38/MAPK-mediated pathway. Collectively, our findings reveal that autophagy may attenuate macrophage-mediated inflammatory response to ConA and may be the potential therapeutic targets for acute liver injury.

Topics: Animals; Apoptosis; Autophagy; Autophagy-Related Protein 7; Concanavalin A; Hepatitis, Autoimmune; Humans; Liver; Liver Failure, Acute; MAP Kinase Signaling System; Mice; Mice, Knockout; Mitochondria; Molecular Targeted Therapy; NF-kappa B; Reactive Oxygen Species; Ubiquitin-Activating Enzymes

To assess alterations in perfusion and liver function in the concanavalin A (ConA)-induced mouse model of acute liver failure (ALF) using two magnetic resonance imaging (MRI)-based methods: dynamic contrast-enhanced MRI (DCE-MRI) with Gd-EOB-DTPA contrast agent and arterial spin labelling (ASL).. BALB/c mice were studied using a 9.4 T MRI system. The IntraGateFLASH. The average perfusion values for the liver of the control and ConA group were equal to 245 ± 20 and 200 ± 32 ml/min/100 g (p = 0.008, respectively). DCE-MRI showed that the time to the peak of the image enhancement was 6.14 ± 1.07 min and 9.72 ± 1.69 min in the control and ConA group (p < 0.001, respectively), while the rate of the contrast wash-out in the control and ConA group was 0.037 ± 0.008 and 0.021 ± 0.008 min. Both the ASL and DCE-MRI techniques represent a reliable methodology to assess alterations in liver perfusion and hepatocyte integrity in murine hepatitis.

Topics: Acute Disease; Animals; Concanavalin A; Contrast Media; Gadolinium DTPA; Hepatitis; Hepatocytes; Humans; Liver; Liver Failure, Acute; Magnetic Resonance Imaging; Mice; Mice, Inbred BALB C; Perfusion; Retrospective Studies; Spin Labels

Cell-based therapies (CBTs) are considered the effective approaches to treat liver failure. However, which cell type is the most suitable source of CBTs for acute liver failure (ALF) or chronic liver failure (CLF) remains unclear. To investigate this, mature hepatocytes in adult liver (adult HCs), fetal liver cells (FLCs), induced hepatic stem cells (iHepSCs) and bone marrow derived mesenchymal stromal cells (BMSCs) were used to CBTs for ConA-induced ALF and Fah-deficient induced CLF in mice. The results showed that only BMSCs remitted liver damage and rescued ALF in ConA-treated mice. In this process, BMSCs inhibited ConA-induced inflammatory response by decreasing the mRNA expressions of TNF-α, IFN-γ and FasL and increasing IL-10 mRNA expression. However, in the CLF model, not BMSCs but adult HCs transplantation lessened liver injury, recovered liver function and rescued the life of Fah-/- mice after NTBC withdrawal. Further study showed that adult HCs offered more effective liver regeneration compared to other cells in Fah-/- mice without NTBC. These results demonstrated that BMSCs and adult HCs are the optimal sources of CBTs for ConA-induced ALF and Fah-deficient induced CLF in mice, respectively. This finding deepens our understanding about how to select a proper CBT for different liver failure.

Topics: Animals; Cell Differentiation; Cell- and Tissue-Based Therapy; Concanavalin A; End Stage Liver Disease; Hepatocytes; Liver Failure, Acute; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice

Acute liver failure (ALF) is a highly complex syndrome characterized by devastating activation of early activation of Kupffer cells (KCs) has been implicated in the pathogenesis of ALF. However, the factors regulating KC early activation are virtually unexplored. The aim of present study was to determine the role of the intracellular high-mobility group box 1 (HMGB1) in modulating the early activation of KCs during ALF. The intravenous injection of Concanavalin A (Con A) was used to establish a mouse model of ALF. The dynamic pro-inflammatory properties and MHC II expression of KCs were measured by qRT-PCR and flow cytometry. HMGB1 expression in KCs was measured by qRT-PCR and Western blotting. The immunofluorescence was implemented to determine the relocation of HMGB1 in KCs, and the siRNA against HMGB1 was utilized to assess the impact of HMGB1 on KC pro-inflammatory properties. The peak of pro-inflammatory cytokines production and MHC II expression in KCs appeared at the early stage of ALF. The up-regulation of HMGB1 expression and the translocation of HMGB1 in KCs were in parallel with the early activation of KCs. The blockade of intracellular HMGB1 expression caused by siRNA significantly inhibited the production of KC-derived pro-inflammatory cytokines, and led to a down-regulation of MAP kinase activation in KCs. The self-derived HMGB1 is an "early alarmin" of KC activation during Con A-induced ALF. HMGB1 might be a potential target for cell-specific strategy in ALF.

Topics: Animals; Cell Nucleus; Concanavalin A; Cytokines; Disease Models, Animal; HMGB1 Protein; Humans; Inflammation Mediators; Kupffer Cells; Liver Failure, Acute; Male; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Molecular Targeted Therapy; Protein Transport; RNA, Small Interfering; Up-Regulation

Activation of hepatic macrophages and systemic immunoparesis are prominent features of acute liver failure (ALF). However, the specific mechanism linking macrophage-associated inflammation and immunoparesis remains to be elucidated. The present study investigated the functional status and fate of hepatic macrophages, as well as their association with immunoparesis in ALF. Intravenous injection of concanavalin A (con A) was administered to develop a mouse model of ALF. Flow cytometry was performed to determine toll-like receptor 4 (TLR4) expression and the apoptotic rates of isolated hepatic macrophages in mice exposed to con A. The levels of TNF-α, IL-6 and IL-12p40 in serum and cell cultures were determined with enzyme-linked immunosorbent assay. TLR4 peak expression in hepatic macrophages was observed at ~0.5 h following exposure to con A and rapidly decreased at 1-3 h. The apoptotic rates of hepatic macrophages increased significantly with the exposure time of con A. The dysfunctional hepatic macrophages associated with apoptosis were observed earlier than the biochemical and histopathological changes of ALF. In addition, the production of macrophage-related inflammatory cytokines following exposure to con A in vivo or in vitro increased significantly. These observations indicated that the deactivation and apoptosis of hepatic macrophages may be a potential link between inflammation and immunoparesis in ALF.

Topics: Animals; Apoptosis; Concanavalin A; Enzyme-Linked Immunosorbent Assay; Interleukin-12 Subunit p40; Interleukin-6; Liver; Liver Failure, Acute; Macrophages; Male; Mice; Mice, Inbred C57BL; Time Factors; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

Necrostatin-1 (Nec-1) inhibits receptor-interacting protein 1 (RIP1) kinase and programmed necrosis. This study was designed to examine the protective effects and mechanisms of Nec-1 in concanavalin A- (ConA-) induced hepatitis in mice.. C57BL/6 mice were exposed to ConA via tail vein injection and injected intraperitoneally with Nec-1 or vehicle. Levels of serum liver enzymes and histopathology were determined. Levels of inflammatory cytokines with ConA-induced hepatitis were determined with real-time polymerase chain reaction (real-time PCR). The expression of TNF- α , RIP1, and LC3 was detected with immunohistochemical staining. The expression of TNF- α , IFN- γ , IL2, IL6, caspase 3, RIP1, beclin-1, and LC3 protein was assessed by immunofluorescence and western blotting. Autophagosomes were observed with transmission electron microscopy (TEM).. Amelioration in liver functions and histopathological changes and the suppression of inflammatory cytokine production were observed in Nec-1-injected mice. Western blotting analysis showed that the expression of TNF- α , IFN- γ , IL2, IL6, and RIP1 was significantly reduced in the Nec-1-injected mice, which was confirmed by immunofluorescence and immunohistochemistry. Autophagosome formation was significantly reduced by Nec-1 treatment, as the expression of beclin-1 and LC3, determined with immunofluorescence and western blotting.. These results demonstrate that Nec-1 prevents ConA-induced liver injury via RIP1-related and autophagy-related pathways.

Topics: Animals; Autophagy; Chemical and Drug Induced Liver Injury; Concanavalin A; Cytokines; Disease Models, Animal; Gene Expression Profiling; Gene Expression Regulation; GTPase-Activating Proteins; Imidazoles; Indoles; Liver; Liver Failure, Acute; Male; Mice; Mice, Inbred C57BL; Microscopy, Electron, Transmission; Microtubule-Associated Proteins; Necrosis; Phagosomes; Real-Time Polymerase Chain Reaction; Transaminases; Tumor Necrosis Factor-alpha

We used Concanavalin A-induced liver injury to study the role of Interleukin 33 and its receptor ST2 in the induction of inflammatory pathology and hepatocellular damage.. We tested susceptibility to Concanavalin A induced hepatitis in ST2 deficient and wild type BALB/c mice and analyzed the effects of single injection of Interleukin 33 as evaluated by liver enzyme test, quantitative histology, mononuclear cell infiltration, cytokine production, intracellular staining of immune cells, and markers of apoptosis in the liver.. ST2 deficient mice developed significantly more severe hepatitis and had significantly higher number of mononuclear cells in the liver, CD4+ and CD8+ T cells, NKp46+ and CD3+NKp46+ cells, and F4/80+ macrophages. The level of pro-inflammatory cytokines in the sera and number of TNF alpha, IFN gamma, and IL-17 producing cells was higher in ST2 deficient mice. In contrast, number of CD4+Foxp3+ cells was statistically higher in wild type mice. Additionally, treatment of wild type mice with single (1 μg) injection of Interleukin 33 led to attenuation of the liver injury and milder infiltration of mononuclear cells, increase in total number of liver CD4+Foxp3+ cells and IL-4 producing CD4+ T cells. Interleukin 33 also suppressed the activation of caspase 3, prevented the expression of BAX, and enhanced the expression of antiapoptotic Bcl-2 in the liver.. We concluded that Interleukin 33/ST2 axis downregulated Concanavalin A-induced liver injury and should be evaluated as potential target in fulminant hepatitis in humans.

Topics: Animals; Chemical and Drug Induced Liver Injury; Concanavalin A; Disease Models, Animal; Humans; Inflammation Mediators; Interleukin-1 Receptor-Like 1 Protein; Interleukin-33; Interleukins; Liver Failure, Acute; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Receptors, Interleukin; Signal Transduction; T-Lymphocyte Subsets

Liver injury is closely associated with immune inflammation. Lacking immunostimulatory functions, viral interleukin-10 (vIL-10), a cellular IL-10 homologue, has been an attractive molecule for immunomodulatory therapy. We aimed to reveal a protective effect of the gene transfer of an adenoviral vector encoding vIL-10 on liver injury induced by concanavalin A.. C57BL/6J mice were intravenously injected with adenoviral vector encoding vIL-10 before concanavalin A challenge. Liver injury was assessed. Interferon-γ and interleukin-4 levels were measured by ELISA. The activation of splenic and hepatic immune cells was analysed using an MTT assay.. Adenoviral vector encoding vIL-10 pretreatment significantly decreased concanavalin A-mediated elevations in serum alanine aminotransaminase and aspartate aminotransaminase activity, and necrotic area in liver tissues. The protective effect of adenoviral vector encoding vIL-10 was attributed to its inhibition of T cell activation, and production of interferon-γ and interleukin-4 by the immune cells. Recombinant mouse IL-10, a high homologous cytokine to vIL-10, effectively downregulated interferon-γ and interleukin-4 release by hepatic mononuclear cells.. Adenovirus vector-mediated vIL-10 gene transfer can prevent concanavalin A-induced hepatic injury, minimise pro-inflammatory cytokine release, and inhibit the activation of T lymphocytes.

Topics: Adenoviridae; Alanine Transaminase; Animals; Aspartate Aminotransferases; Concanavalin A; Down-Regulation; Gene Transfer Techniques; Genetic Vectors; Interferon-gamma; Interleukin-10; Interleukin-4; Liver; Liver Failure, Acute; Mice; Mice, Inbred C57BL; Mitogens; Monocytes; T-Lymphocytes

Dysregulation of signal transducer and activator of transcription (STAT) signaling is usually associated with intricate immune diseases and rebuilding the balance of STAT1 and STAT3 signaling is being explored as a useful approach for the treatment of these diseases. However, few chemicals have been reported to rebuild the balance of these two signalings for immune hepatitis therapy. In the present study, we found that fusaruside, a new kind of cerebroside isolated from an endophytic fungus Fusarium sp. IFB-121 in Quercus variabilis, significantly ameliorated concanavalin A (Con A)-induced T-cell-mediated fulminant hepatitis in mice, which was closely associated with the improvement of histopathological parameters, inhibition of activation of liver CD4(+) T cells and NKT cells, regulation of balance of Th1/Th2/Th17/Treg cytokines and protection of hepatocyte from apoptosis. Moreover, T-cell proliferation and activation was also notably inhibited by fusaruside in vitro. Furthermore, the protective effect of fusaruside was attributable to a novel regulatory mechanism through down-regulating STAT1 activation and T-bet expression in liver CD4(+) T cells and up-regulating STAT3 activation and Bcl-X(L) expression in hepatocytes. In conclusion, fusaruside exhibited its capability against T-cell-mediated liver injury in vivo, through rebuilding the balance of STAT1 and STAT3 signalings. These results suggest that fusaruside is potentially useful for the treatment of T-cell-mediated human liver disorders.

Topics: Animals; bcl-X Protein; CD4 Antigens; Cell Proliferation; Cerebrosides; Concanavalin A; Cytokines; Female; Fusarium; Hepatitis, Animal; Liver; Liver Failure, Acute; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Natural Killer T-Cells; Signal Transduction; Spleen; STAT1 Transcription Factor; STAT2 Transcription Factor; T-Lymphocytes

Fulminant hepatitis progresses to acute liver failure (ALF) when the extent of hepatocyte death exceeds the liver's regenerative capacity. Although small interfering RNA (siRNA) appears promising in animal models of hepatitis, the approach is limited by drawbacks associated with systemic administration of siRNA. The aim of this study is to develop a hepatocyte-specific delivery system of siRNA for treatment of fulminant hepatitis.. Galactose-conjugated liposome nano-particles (Gal-LipoNP) bearing siRNA was prepared, and the particle size and zeta potential of Gal-LipoNP/siRNA complexes were measured. The distribution, cytotoxicity and gene silence efficiency were studied in vivo in a concanavalin A (ConA)-induced hepatitis model. C57BL/6 mice were treated with Gal-LipoNP Fas siRNA by i.v. injection 72 h before ConA challenge, and hepatocyte injury was evaluated using serum alanine transferase (ALT) and aspartate transaminase (AST) levels, as well as liver histopathology and TUNEL-positive hepatocytes. The galactose-ligated liposomes were capable of encapsulating >96% siRNA and exhibited a higher stability than naked siRNA in plasma. Hepatocyte-specific targeting was confirmed by in vivo delivery experiment, in which the majority of Gal-LipoNP-siRNA evaded nuclease digestion and accumulated in the liver as soon as 6 h after administration. In vivo gene silencing was significant in the liver after treatment of Gal-Lipo-siRNA. In the ConA-induced hepatitis model, serum levels of ALT and AST were significantly reduced in mice treated with Gal-lipoNP-siRNA as compared with control mice. Additionally, tissue histopathology and apoptosis showed an overall reduction of injury in the Gal-LipoNP siRNA-treated mice.. This study is the first to our knowledge to demonstrate reduction of hepatic injury by liver-specific induction of RNA interference using Gal-LipoNP Fas siRNA, highlighting a novel RNAi-based therapeutic potential in many liver diseases.

Topics: Animals; Apoptosis; Concanavalin A; fas Receptor; Galactose; Gene Silencing; Gene Transfer Techniques; Hepatocytes; Inflammation; Liposomes; Liver; Liver Failure, Acute; Male; Mice; Mice, Inbred C57BL; Nanoparticles; Organ Specificity; Particle Size; RNA, Small Interfering; Static Electricity; Tissue Distribution

High-mobility group box-1 (HMGB1) plays important roles in inflammation, immune responses, and tumor progression. Since HMGB1 and its components have been shown to be mediators of a number of diseases but several sources of recombinant HMGB1 showed controversial biological activity, it is important to obtain recombinant HMGB1 with properties that resemble the native protein. For this purpose, we cloned genes coding for human HMGB1 and its active components A box and B box by PCR and inserted the cloned genes into pET28a vectors for transformation of Escherichia coli BL21. The E. coli expressed proteins were then purified with a Ni(2+)-NTA column and the endotoxin content was removed. Recombinant human HMGB1 (rhHMGB1) and its B box thus obtained stimulated, but A box inhibited, the production of the chemokine CXCL8/IL-8 by THP-1 monocytic cell line. We also used purified rhHMGB1 to immunize rabbits and generated potent anti-sera, which was capable of neutralizing the activity of rhHMGB1 in vitro and detecting the increased HMGB1 expression in inflammatory tissues in mice and humans. Thus, we have established essential means to produce biologically active rhHMGB1 that will facilitate us to study its role in diseases and to explore its potential as a therapeutic agent.

Topics: Animals; Cell Line; Cloning, Molecular; Concanavalin A; Escherichia coli; HMGB1 Protein; Humans; Immune Sera; Immunization; Inflammation; Interleukin-8; Liver Failure, Acute; Mice; Monocytes; Psoriasis; Rabbits; Recombinant Proteins; Transformation, Bacterial

Cell therapy has been used to attenuate liver injury. Here we evaluated whether genetic engineering of either bone marrow-derived mononuclear cells (MNC) or endothelial progenitor cells (EPC) many enhance their hepatoprotective properties.. Mice with ConA-induced hepatitis or with lethal fulminant hepatitis resulting from administration of an adenovirus encoding CD40L (AdCD40L) received an intra-splenic injection of saline or 2 × 10(6) unmodified MNC or EPC or the same cells transduced ex vivo with an adenovirus expressing luciferase (MNCLUC and EPCLUC) or encoding the hepatoprotective cytokine cardiotrophin-1 (CT-1) (MNCCT-1 and EPCCT-1). We analyzed the extent of liver damage, the intensity of inflammatory reaction, and animal survival.. Luciferase immunohistochemistry showed that after injection into the spleen, the engineered cells migrated efficiently to the damaged liver. In mice with ConA hepatitis EPCCT-1, but not other forms of cell therapy, significantly decreased serum transaminases and induced more intense histological improvement than other treatments. This superior therapeutic effect was associated with upregulation of cytoprotective molecules including IGF-I and EGF, lower expression of proinflammatory cytokines, IL-1b and TNFα, and decreased granzyme B levels. In AdCD40L-induced lethal fulminant hepatitis, EPCCT-1 also exceeded other cell therapies in attenuating the expression of proinflammatory mediators and hepatic injury enabling 35.7% survival while mortality was 100% in the other treatment groups.. Genetic engineering of EPC to overexpress CT-1 enhances the hepatoprotective properties of EPC and constitutes a therapy that deserves consideration for acute liver failure.

Topics: Adenoviridae; Animals; Cell Movement; Chemical and Drug Induced Liver Injury; Concanavalin A; Disease Models, Animal; Endothelial Cells; Genetic Engineering; Genetic Therapy; Liver Failure, Acute; Male; Mice; Mice, Inbred C57BL; Mitogens; Organic Cation Transport Proteins; Phenotype; Solute Carrier Family 22 Member 5; Stem Cell Transplantation; Stem Cells; Survival Rate; Transgenes

Con A-induced fulminant hepatitis is a well-known animal model for acute liver failure. However, the role of γδ T cells in this model is undefined. In this report, using TCR δ(-/-) mice, we demonstrated a protective role of γδ T cells in Con A-induced hepatitis model. TCR δ(-/-) mice showed significantly decreased levels of IL-17A and IL-17F in the Con A-treated liver tissue, and reconstitution of TCR δ(-/-) mice with wild-type (Wt), but not IL-17A(-/-), γδ T cells significantly reduced hepatitis, strongly suggesting a critical role of IL-17A in mediating the protective effect of γδ T cells. Interestingly, only Vγ4, but not Vγ1, γδ T cells exerted such a protective effect. Furthermore, depletion of NKT cells in TCR δ(-/-) mice completely abolished hepatitis, and NKT cells from Con A-challenged liver tissues of TCR δ(-/-) mice expressed significantly higher amounts of proinflammatory cytokine IFN-γ than those from Wt mice, indicating that γδ T cells protected hepatitis through targeting NKT cells. Finally, abnormal capacity of IFN-γ production by NKT cells of TCR δ(-/-) mice could only be downregulated by transferring Wt, but not IL-17(-/-), Vγ4 γδ T cells, confirming an essential role of Vγ4-derived IL-17A in regulating the function of NKT cells. In summary, our report thus demonstrated a novel function of Vγ4 γδ T cells in mediating a protective effect against Con A-induced fulminant hepatitis through negatively regulating function of NKT cells in an IL-17A-dependent manner, and transferring Vγ4 γδ T cells may provide a novel therapeutic approach for this devastating liver disease.

Topics: Animals; Concanavalin A; Disease Models, Animal; Down-Regulation; Female; Interferon-gamma; Interleukin-17; Liver Failure, Acute; Male; Mice; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Natural Killer T-Cells; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocyte Subsets

A large amount of evidence suggests that monocytes/macrophages infiltration is implicated in a variety of inflammatory diseases including acute liver injury. Monocyte chemoattractant protein 1 (MCP-1) plays a crucial role in the process of macrophages recruitment. We herein presented a small-molecule library and a feasible quick screening method of evaluating potency of inhibition of chemotaxis of RAW264.7 cells stimulated by MCP-1. Fifty-three small molecules were synthesized and screened, and four compounds (2g, 2h, 4f, and 6h) showed inhibitory effects with IC(50) values range from 0.72 to 20.47 microM, with compound 4f being the most efficient. Further in vivo studies demonstrated that oral administration of 2g, 2h, 4f, or 6h decreases, most significantly for 4f, the serum levels of alanine aminotransaminase (ALT) and asparate aminotransaminase (AST) in ConA-induced acute livery injury BALB/c mice. Histopathological evaluation liver sections confirmed 4f as a potent, orally active compound for hepatoprotective effects against ConA-induced acute liver injury in BALB/c mice.

Topics: Animals; Cell Movement; Cells, Cultured; Chemical and Drug Induced Liver Injury; Concanavalin A; Disease Models, Animal; Drug Discovery; Drug Evaluation, Preclinical; Female; Liver Failure, Acute; Liver Function Tests; Mice; Mice, Inbred BALB C; Small Molecule Libraries; Stereoisomerism; Structure-Activity Relationship; Thiazolidinediones

Due to the high morbidity and mortality of fulminant hepatitis, early diagnosis followed by early effective treatment is the key for prognosis improvement. So far, little is known about the gene expression changes in the early stage of this serious illness. Identification of the genes related to the very early stage of fulminant hepatitis development may provide precise clues for early diagnosis.. Balb/C mice were used for ConA injection to induce fulminant hepatitis that was confirmed by pathological and biochemical examination. After a gene chip-based screening, the data of gene expression in the liver, was further dissected by ANOVA analysis, gene expression profiles, gene network construction and real-time RT-PCR. At the very early stage of ConA-triggered fulminant hepatitis, totally 1,473 genes with different expression variations were identified. Among these, 26 genes were finally selected for further investigation. The data from gene network analysis demonstrate that two genes, MPDZ and Acsl1, localized in the core of the network.. At the early stages of fulminant hepatitis, expression of twenty-six genes involved in protein transport, transcription regulation and cell metabolism altered significantly. These genes form a network and have shown strong correlation with fulminant hepatitis development. Our study provides several potential targets for the early diagnosis of fulminant hepatitis.

Topics: Algorithms; Analysis of Variance; Animals; Concanavalin A; Gene Expression Profiling; Hepatitis, Animal; Liver Failure, Acute; Mice; Mice, Inbred BALB C; Oligonucleotide Array Sequence Analysis

T-cell-mediated fulminant hepatitis is a potentially life-threatening event for which the underlying pathogenic mechanisms are not fully understood. Here, we demonstrate a key regulatory role of IL-28A in T-cell-mediated hepatitis.. We cloned the murine IL-28A gene by reverse-transcription polymerase chain reaction, assessed the effects of recombinant IL-28A, and generated IL-28A-transgenic mice.. IL-28A induced TH1 cytokine production by CD4+ T lymphocytes in a T-bet-dependent manner and was up-regulated in a murine model of T-cell-mediated hepatitis upon Con A administration. In vivo, CD4+ T cells from newly created IL-28A-transgenic animals revealed an increased proliferation and proinflammatory TH1 cytokine production, as compared with wild-type mice. In addition, IL-28A-transgenic mice showed markedly augmented Con A-induced hepatitis with up-regulated interferon-gamma cytokine production, as compared with wild-type mice. Transgenic mice exhibited an up-regulation of the interferon-gamma-T-bet signaling pathway in Con A hepatitis, and augmented hepatitis in these mice was suppressed by crossing them with T-bet-deficient mice. In addition, in vivo blockade of interferon-gamma but not IL-4 suppressed augmented liver inflammation in transgenic mice, suggesting that IL-28A induces the T-bet signaling pathway in T-cell-induced hepatitis. Finally, IL-28A-specific antisense phosphorothioate oligonucleotides suppressed liver pathology in Con A-treated wild-type mice, as compared with the case of control oligonucleotides.. IL-28A emerges as a key regulatory cytokine with pathogenic function in T-cell-mediated liver injury. Thus, targeting of IL-28A represents a potential novel approach for therapy of Th1-mediated inflammatory diseases such as T-cell-mediated hepatitis.

Topics: Animals; Antibodies; CD4-Positive T-Lymphocytes; Cell Division; Cloning, Molecular; Codon, Initiator; Concanavalin A; Interferon-gamma; Interleukin-2; Interleukin-4; Interleukins; Liver Failure, Acute; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mitogens; Oligonucleotides, Antisense; Receptors, Cytokine; Signal Transduction; STAT1 Transcription Factor; T-Box Domain Proteins; T-Lymphocyte Subsets

Osteopontin (OPN) has been implicated in various helper T cell type 1 immunity-mediated diseases including rheumatoid arthritis (RA), multiple sclerosis (MS), Crohn's disease, and fulminant hepatitis. Increased expression of OPN has been detected in pathological foci of these diseases. RA and fulminant hepatitis have been successfully treated by administration of neutralizing anti-OPN antibody in mice. Antibody treatment may elicit side effects including allergic reactions against heterologous antibody proteins, thus necessitating humanization of antibody. To provide alternative means to neutralize OPN function, in this study we explored the possibility of using OPN small interfering RNA (siRNA) to silence OPN gene expression. In vitro, OPN siRNA efficiently silenced the expression of both exogenous and endogenous OPN gene. After hydrodynamic intravenous injection of OPN siRNA, OPN siRNA was efficiently delivered to the liver, which resulted in the efficient silencing of OPN gene expression in liver. In a murine model of concanavalin A (ConA)-induced fulminant hepatitis, OPN expression was elevated in liver and severe hepatic necrosis was induced. Importantly, after OPN siRNA treatment, the OPN expression level in liver was significantly reduced and liver tissue injury was ameliorated, as reflected by the significant reduction of serum alanine aminotransferase levels and almost normal liver histology. Thus, this study indicates that OPN siRNA delivery has therapeutic potential in various inflammatory diseases in which OPN play a critical role by silencing OPN gene expression in vivo.

Topics: Animals; Chemical and Drug Induced Liver Injury; Concanavalin A; Genetic Therapy; Liver; Liver Failure, Acute; Mice; Mice, Inbred C57BL; Osteopontin; RNA Interference; RNA, Small Interfering; Transfection

Recent reports suggest that the adipose tissue and adipokines are potent modulators of inflammation. However, there is only scarce knowledge on the functional role and regulation of endogenous adiponectin in non-fat tissues such as the liver under conditions of acute inflammation.. In the present study, we investigated adiponectin expression in healthy murine liver tissue and under inflammatory conditions in vivo.. Adiponectin mRNA was readily detectable in healthy liver tissue and further increased in ConA-mediated acute liver failure. Adiponectin protein expression was mainly found in hepatic endothelial cells. In vitro adiponectin mRNA expression was detectable in several cell types, including primary hepatic sinusoidal endothelial cells, stellate cells, and macrophages. Mice pretreated with adiponectin before ConA administration developed reduced hepatic injury as shown by decreased release of transaminases and reduced hepatocellular apoptotis. Of note, TNF-alpha levels were not affected by adiponectin, whereas IL-10 production was increased. Neutralisation of IL-10 diminished the protective effect of adiponectin.. Adiponectin expression is up-regulated in ConA-mediated acute liver failure. Therefore, adiponectin might play a role in the control and limitation of inflammation in the liver. Moreover, our data suggest a role for IL-10 in adiponectin-mediated hepatoprotection.

Topics: Adiponectin; Animals; Concanavalin A; Disease Models, Animal; Endothelial Cells; Female; Hepatocytes; Interleukin-10; Liver Failure, Acute; Macrophages; Mice; Mice, Inbred BALB C; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Up-Regulation

Concanavalin A (Con A) activates T lymphocytes and induces CD4+ T cell-mediated hepatic injury in mice. Pro-inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), and interleukin-6 (IL-6), are critical mediators in this experimental model. Activation of adenosine A2A receptors reduces the production of various pro-inflammatory cytokines and suppresses T cell activation. A selective adenosine A2A receptor agonist (ATL-146e) has been shown to be a potent inhibitor of inflammation by increasing intracellular cyclic AMP (cAMP) in leukocytes. The aim of the present study was to determine whether ATL-146e could ameliorate Con A-induced hepatic injury, reduction of pro-inflammatory cytokine production.. Balb/c mice were injected with 25mg/kg Con A with or without a single injection of ATL-146e (0.5-50 microg/kg), 5 min prior to Con A administration. Liver enzymes, histology, and serum levels of tumor necrosis factor-alpha, interferon-gamma, and interleukin-6 were examined. We also assessed the effects of ATL-146e on pro-inflammatory cytokine production with CD4+ T cell.. Pretreatment with ATL-146e significantly reduced serum levels of liver enzymes (P<0.001). The serum pro-inflammatory cytokines were all increased after Con A administration and reduced to near normal levels by ATL-146e. ATL-146e also inhibited CD4+ T cell pro-inflammatory cytokine production.. A selective adenosine A2A receptor agonist, ATL-146e, can prevent concanavalin A-induced hepatic injury that is presumably mediated by its anti-inflammatory properties.

Topics: Adenosine A2 Receptor Agonists; Animals; Concanavalin A; Cyclohexanecarboxylic Acids; Interferon-gamma; Interleukin-6; Liver Failure, Acute; Male; Mice; Mice, Inbred BALB C; Purines; Tumor Necrosis Factor-alpha

Fulminant liver failure (FLF) consists of a cascade of events beginning with a presumed uncontrolled systemic activation of the immune system. The etiology of FLF remains undefined. In this study, we demonstrate that CCR5 deficiency promotes the development of acute FLF in mice following Con A administration by preventing activated hepatic CD1d-restricted NKT cells (but not conventional T cells) from dying from activation-induced apoptosis. The resistance of CCR5-deficient NKT cells from activation-induced apoptosis following Con A administration is not due to a defective Fas-driven death pathway. Moreover, FLF in CCR5-deficient mice also correlated with hepatic CCR5-deficient NKT cells, producing more IL-4, but not IFN-gamma, relative to wild-type NKT cells. Furthermore, FLF in these mice was abolished by IL-4 mAb or NK1.1 mAb treatment. We propose that CCR5 deficiency may predispose individuals to the development of FLF by preventing hepatic NKT cell apoptosis and by regulating NKT cell function, establishing a novel role for CCR5 in the development of this catastrophic liver disease that is independent of leukocyte recruitment.

Topics: Animals; Antigens, CD1; Antigens, CD1d; Apoptosis; Concanavalin A; fas Receptor; Immunity, Innate; Interleukin-4; Killer Cells, Natural; Liver; Liver Failure, Acute; Lymphocyte Activation; Lymphocyte Depletion; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Receptors, CCR5; Spleen; T-Lymphocyte Subsets

Prostacyclin (PGI(2)) is a potent mediator in the inflammatory and coagulation processes. The aim of this study was to test whether beraprost sodium, a PGI(2) analogue, could prevent experimental hepatic injury induced by concanavalin A (Con A), which is a model of fulminant hepatic failure.. Beraprost (100 microg/kg) was administered intraperitoneally simultaneously with Con A (40 mg/kg) in C57B6J mice. Blood circulation in the liver was determined by laser-Doppler flowmetry. Plasma levels of alanine aminotransferase (ALT), tumor necrosis factor (TNF)-alpha, interferon (IFN)-gamma, and interleukin (IL)-6 were determined. Levels of TNF-alpha and IFN-gamma in culture supernatant of splenocytes were also determined.. Beraprost administration reduced the incidence of death following hepatic failure (76.5% vs. 29.4%, P<0.05). Plasma levels of ALT were significantly lower in the beraprost-treated group than in the control group, and in the former, there was concomitant suppression of the histological features of injury. Beraprost significantly increased hepatic blood flow volume in Con A-treated mice. Plasma levels of TNF-alpha and IFN-gamma were significantly reduced at 6 and 12 h after Con A injection, respectively, but the levels of IL-6 were increased at 6 h. In vitro, beraprost also suppressed Con A-induced TNF-alpha production in splenocytes, while it stimulated IFN-gamma production.. These findings imply that beraprost suppresses Con A-induced liver injury. These data also suggest that beraprost, which is clinically effective in treating pulmonary hypertension, may have therapeutic potential for preventing hepatic injury.

Topics: Animals; Concanavalin A; Epoprostenol; Interferon-gamma; Interleukin-6; Liver Circulation; Liver Failure, Acute; Male; Mice; Mice, Inbred C57BL; Nitric Oxide; Spleen; Tumor Necrosis Factor-alpha

TNF-alpha has been clearly identified as central mediator of T cell activation-induced acute hepatic injury in mice, e.g., Con A hepatitis. In this model, liver injury depends on both TNFRs, i.e., the 55-kDa TNFR1 as well as the 75-kDa TNFR2. We show in this report that the hepatic TNFRs are not transcriptionally regulated, but are regulated by receptor shedding. TNF directly mediates hepatocellular death by activation of TNFR1 but also induces the expression of inflammatory proteins, such as cytokines and adhesion molecules. Here we provide evidence that resistance of TNFR1(-/-) and TNFR2(-/-) mice against Con A hepatitis is not due to an impaired production of the central mediators TNF and IFN-gamma. Con A injection results in a massive induction of ICAM-1, VCAM-1, and E-selectin in the liver. Lack of either one of both TNFRs did not change adhesion molecule expression in the livers of Con A-treated mice, presumably reflecting the fact that other endothelial cell-activating cytokines up-regulated adhesion molecule expression. However, treatment of TNFR1(-/-) and TNFR2(-/-) mice with murine rTNF revealed a predominant role for TNFR1 for the induction of hepatic adhesion molecule expression. Pretreatment with blocking Abs against E- and P-selectin or of ICAM(-/-) mice with anti-VCAM-1 Abs failed to prevent Con A hepatitis, although accumulation of the critical cell population, i.e., CD4(+) T cells was significantly inhibited. Hence, up-regulation of adhesion molecules during acute hepatitis unlikely contributes to organ injury but rather represents a defense mechanism.

Topics: Animals; Antibodies, Monoclonal; Antigens, CD; CD4-Positive T-Lymphocytes; Cell Adhesion Molecules; Cell Movement; Chemical and Drug Induced Liver Injury; Concanavalin A; Cytokines; E-Selectin; Injections, Intraperitoneal; Injections, Intravenous; Intercellular Adhesion Molecule-1; Liver; Liver Failure, Acute; Lymphocyte Function-Associated Antigen-1; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Receptors, Tumor Necrosis Factor, Type II; Recombinant Proteins; RNA, Messenger; Solubility; Tumor Necrosis Factor-alpha; Up-Regulation; Vascular Cell Adhesion Molecule-1

Fulminant hepatic failure (FHF) is a clinical syndrome resulting from massive death of liver cells or sudden and severe impairment of liver function. The causes of FHF are diverse and the overall mortality is very high. Recently, it became clear that apoptosis of hepatocytes is the critical cause of acute hepatic failure in FHF. It is well known that a family of cysteine proteases called caspase is one of the key mediators of the apoptotic pathway. Thus, caspases are attractive potential targets for the treatment of disorders resulting from excessive apoptosis. In this report, we examined the activity of a new caspase inhibitor, Xyz 033 mp. This nonpeptide inhibitor showed broad-spectrum caspase-inhibiting activity and protected primary rat hepatocytes from apoptotic death. In a mouse model of FHF induced by concavalin A (Con A), Xyz 033 mp suppressed elevated AST and ALT and specifically reduced IL-1 beta concentration. Also, Xyz 033 mp rescued mice from lethal experimental hepatitis induced by Con A. In addition, histological examinations indicated that Xyz 033 mp protected hepatocytes from the fatal apoptogenic effect of Con A. These results suggest that Xyz 033 mp may be a candidate therapeutic agent for FHF caused by massive apoptotic death of hepatocytes.

Topics: Animals; Apoptosis; Benzene Derivatives; Caspase Inhibitors; Cell Death; Concanavalin A; Cysteine Proteinase Inhibitors; Cytosol; Dactinomycin; Female; Hepatocytes; Liver Failure, Acute; Male; Mice; Mice, Inbred BALB C; Rats; Rats, Sprague-Dawley; Tumor Necrosis Factor-alpha

The significance of tumor necrosis factor receptor 1 (TNFR1) for TNF function in vivo is well documented, whereas the role of TNFR2 so far remains obscure. In a model of concanavalin A (Con A)-induced, CD4+ T cell-dependent experimental hepatitis in mice, in which TNF is a central mediator of apoptotic and necrotic liver damage, we now provide evidence for an essential in vivo function of TNFR2 in this pathophysiological process. We demonstrate that a cooperation of TNFR1 and TNFR2 is required for hepatotoxicity as mice deficient of either receptor were resistant against Con A. A significant role of TNFR2 for Con A-induced hepatitis is also shown by the enhanced sensitivity of transgenic mice overexpressing the human TNFR2. The ligand for cytotoxic signaling via both TNF receptors is the precursor of soluble TNF, i.e. transmembrane TNF. Indeed, transmembrane TNF is sufficient to mediate hepatic damage, as transgenic mice deficient in wild-type soluble TNF but expressing a mutated nonsecretable form of TNF developed inflammatory liver disease.

Topics: Animals; Antigens, CD; Concanavalin A; Hepatitis, Animal; Immunity, Innate; Interferon-gamma; Liver Failure, Acute; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Knockout; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Receptors, Tumor Necrosis Factor, Type II; Tumor Necrosis Factor-alpha