The role of the sphingosine axis in immune regulation: A dichotomy in the anti-inflammatory effects between sphingosine kinase 1 and sphingosine kinase 2-dependent pathways.

Background: Sphingosine kinase has been identified as playing a central role in the immune cascade, being a common mediator in the cellular response to a variety of signals. The different effects of sphingosine kinase 1 and 2 (SphK1 and SphK2, respectively) activity have not been completely characterized. Aim: To determine the different roles played by SphK1 and SphK2 in the regulation of immune-mediated disorders. Methods: Nine groups of mice were studied. Concanavalin A (ConA) injection was used to induce immune-mediated hepatitis. Mice were treated with SphK1 inhibitor (termed SphK-I) and SphK2 inhibitor (termed ABC294640), prior to ConA injection, and effects of treatment on liver enzymes, subsets of T lymphocytes, and serum levels of cytokines were observed. Results: While liver enzyme elevation was ameliorated by administration of SphK1 inhibitor, SphK2 inhibitor-treated mice did not show this tendency. A marked decrease in expression of CD25+ T-cells and Foxp+ T-cells was observed in mice treated with a high dose of SphK1 inhibitor. Alleviation of liver damage was associated with a statistically significant reduction of serum IFNγ levels in mice treated with SphK1 inhibitor and not in those treated with SphK2 inhibitor. Conclusions: Early administration of SphK1 inhibitor in a murine model of immune-mediated hepatitis alleviated liver damage and inflammation with a statistically significant reduction in IFN-γ levels. The data support a dichotomy in the anti-inflammatory effects of SphK1 and SphK2, and suggests that isoenzyme-directed therapies can improve the effect of targeting these pathways.

Resveratrol modulates Toxoplasma gondii infection induced liver injury by intervening in the HMGB1/TLR4/NF-κB signaling pathway.

Toxoplasma gondii (T. gondii) is an obligate intracellular parasite that can cause liver diseases in the host, including hepatitis and hepatomegaly. High mobility group box 1 (HMGB1) is the main inflammatory mediator causing cell injury or necrosis. HMGB1 binds to toll like receptor 4 (TLR4), then activates the nuclear factor-κB (NF-κB) signaling pathway, which promotes the release of inflammatory factors. Our previous studies showed that HMGB1 mediated TLR4/NF-κB signaling pathway plays an important role in liver injury induced by T. gondii infection. Resveratrol (RSV) is a small polyphenol, which has anti-inflammatory, anti-cancer, anti-T. gondii effect. However, the effect of RSV on liver injury caused by T. gondii infection is unclear. This study used the RH strain tachyzoites of T. gondii to infect murine liver line, NCTC-1469 cells to establish an in vitro model and acute infection of mice for the in vivo model to explore the protective effect of RSV on liver injury induced by T. gondii infection. The results showed that RSV inhibited the proliferation of T. gondii in the liver, reduced the alanine aminotransferase/aspartate aminotransferase levels and pathological liver damage. Additionally, RSV inhibited the production of tumor necrosis factor-α, inducible nitric oxide synthase and HMGB1 by interfering with the TLR4/NF-κB signaling pathway. These results indicate that RSV can protect liver injury caused by T. gondii infection by intervening in the HMGB1/TLR4/NF-κB signaling pathway. This study will provide a theoretical basis for RSV treatment of T. gondii infection induced liver injury.

Activation of Autophagy Relieves Linoleic Acid-Induced Inflammation in Large Yellow Croaker (Larimichthys crocea).

High levels of soybean oil (SO) in fish diets enriched with linoleic acid (LA, 18:2n-6) could induce strong inflammation. However, the molecular mechanism underlying LA-induced inflammation in the liver of large yellow croaker (Larimichthys crocea) has not been elucidated. Based on previous research, autophagy has been considered a new pathway to relieve inflammation. Therefore, the present study was performed to investigate the role of autophagy in regulating LA-induced inflammation in the liver of large yellow croaker in vivo and in vitro. The results of the present study showed that activation of autophagy in liver or hepatocytes could significantly reduce the gene expression of proinflammatory factors, such as tumor necrosis factor α (TNFα) and interleukin 1ß (IL1ß). The results of the present study also showed that inhibition of autophagy could upregulate the gene expression of proinflammatory factors and downregulate the gene expression of anti-inflammatory factors in vivo and in vitro. Furthermore, autophagy could alleviate LA-induced inflammatory cytokine gene expression in vivo and in vitro, while inhibition of autophagy obtained the opposite results. In conclusion, our study shows that autophagy could regulate inflammation and alleviate LA-induced inflammation in the liver of large yellow croaker in vivo and in vitro for the first time, which may offer considerable benefits to the aquaculture industry and human health.

Natural Killer T Cells in Various Mouse Models of Hepatitis.

Natural killer T (NKT) cells are a key component of innate immunity. Importantly, a growing body of evidence indicates that NKT cells play an integral role in various acute and chronic liver injuries. NKT cells participate in the progression of an injury through the secretion of cytokines, which promote neutrophil infiltration and enhance Fas ligand (FasL) and granzyme-mediated NKT cytotoxic activity. Therefore, examining the role of NKT cells in hepatic disease is critical for a comprehensive understanding of disease pathogenesis and may provide insight into novel approaches for treatment. For more than a century, mouse models that imitate the physiopathological conditions of human disease have served as a critical tool in biological and medical basic research, including studies of liver disease. Here, we review the role of NKT cells in various mouse models of hepatitis.

Macrophage-derived extracellular vesicles regulate concanavalin A-induced hepatitis by suppressing macrophage cytokine production.

Acute liver failure is a clinical syndrome of severe hepatic dysfunction. Immune cells play an important role in acute liver failure. In recent years, the immunoregulatory function of extracellular vesicles (EVs) has been reported; therefore, it is inferred that EVs play a role in immune-mediated hepatitis. In this study, we investigated the immunoregulatory function of EVs in concanavalin A (Con A)-induced hepatitis. The mouse model was prepared by a single intravenous administration of 15 mg/kg Con A, in which there was a significant increase in the serum EVs number. In an in vitro study, the number of secreted EVs was also significantly increased in Con A-treated RAW264.7 cells, a mouse macrophage cell line, but not in Hepa1-6 cells, a mouse hepatoma cell line. In an in vitro EVs treatment study, EVs from Con A-treated mouse serum and Con A-treated RAW264.7 cells suppressed inflammatory cytokine production in Con A-stimulated RAW264.7 cells. miRNA sequencing analysis showed that the expression of mmu-miR-122-5p and mmu-miR-148a-3p was commonly increased in these EVs and EVs-treated cells. The pathways enriched in the predicted miRNA target genes included inflammatory response pathways. The mRNA levels of the target genes in these pathways (mitogen-activated protein kinase, phosphoinositide 3-kinase/Akt and Rho/Rho-associated coiled-coil containing protein kinase pathways) were decreased in the EVs-treated cells. In an in vivo RNA interference study, the knockdown of liver RAB27A, an EVs secretion regulator, significantly exacerbated Con A-induced hepatitis. These data suggest that macrophage-derived EVs play an important role in Con A-induced hepatitis through immunoregulation.

Ly6cLo non-classical monocytes promote resolution of rhesus rotavirus-mediated perinatal hepatic inflammation.

Perinatal hepatic inflammation can have devastating consequences. Monocytes play an important role in the initiation and resolution of inflammation, and their diverse functions can be attributed to specific cellular subsets: pro-inflammatory or classical monocytes (Ly6cHi) and pro-reparative or non-classical monocytes (Ly6cLo). We hypothesized that inherent differences in Ly6cHi classical monocytes and Ly6cLo non-classical monocytes determine susceptibility to perinatal hepatic inflammation in late gestation fetuses and neonates. We found an anti-inflammatory transcriptional profile expressed by Ly6cLo non-classical monocytes, and a physiologic abundance of these cells in the late gestation fetal liver. Unlike neonatal pups, late gestation fetuses proved to be resistant to rhesus rotavirus (RRV) mediated liver inflammation. Furthermore, neonatal pups were rendered resistant to RRV-mediated liver injury when Ly6cLo non-classical monocytes were expanded. Pharmacologic inhibition of Ly6cLo non-classical monocytes in this setting restored susceptibility to RRV-mediated disease. These data demonstrate that Ly6cLo monocytes promote resolution of perinatal liver inflammation in the late gestation fetus, where there is a physiologic expansion of non-classical monocytes, and in the neonatal liver upon experimental expansion of these cells. Therapeutic strategies directed towards enhancing Ly6cLo non-classical monocyte function may mitigate the detrimental effects of perinatal liver inflammation.

Chicken infectious anemia virus helps fowl adenovirus break the protection of maternal antibody and cause inclusion body hepatitis-hydropericardium syndrome in layers after using co-contaminated Newcastle disease virus-attenuated vaccine.

Inclusion body hepatitis-hydropericardium syndrome (IBH-HPS) caused by fowl adenovirus type 4 (FAdV-4) has caused huge economic losses for China in the past five years. At present, this disease is controlled in many flocks with the inactivated FAdV vaccine, but the offspring chicks of a layer breeding flock that were vaccinated with this vaccine still became infected and developed IBH-HPS with a 20% mortality rate. Analysis revealed that the NDV-attenuated vaccine in use from the above-mentioned poultry farm was simultaneously contaminated with FAdV-4 and chicken infectious anemia virus (CIAV). The FAdV and CIAV isolated from the vaccine were purified for the artificial preparation of an NDV-attenuated vaccine singly contaminated with FAdV or CIAV, or simultaneously contaminated with both of them. Seven-day-old layers with maternal FAdV antibody were inoculated with the artificially prepared, contaminated vaccines and assessed for corresponding indices. The experiments showed that no obvious symptoms occurred after using the NDV-attenuated vaccine singly contaminated with FAdV or CIAV; however, common IBH and occasional HPS-related death was found in birds after administering the NDV-attenuated vaccine co-contaminated with FAdV and CIAV. In conclusion, this study illustrated that CIAV could assist FAdV in breaking maternal FAdV antibody protection, which then caused the IBH-HPS after vaccination with the co-contaminated NDV vaccine.

Glt25d2 Knockout Directly Increases CD25+CD69- but Decreases CD25-CD69+ Subset Proliferation and is Involved in Concanavalin-Induced Hepatitis.

BACKGROUND/AIMS: The elaborate structure of the extracellular matrix (ECM) and the appropriate surface glycoforms upon it are indispensable to CD4+ T cell regulation. METHODS: To explore the effects of Glcα1,2Galß1 glycosylation mediated by GLT25D2 (Colgalt2) for CD4+ T cell regulation, we prepared C57BL/6J Glt25d2-/- mice. In the induction of hepatitis, after concanavalin A (Con A) challenge for 6, 12, and 24 h, more extensive parenchymal injury was noted in Glt25d2-/- mice than in wild-type (WT) mice at 12 h. Immunohistochemistry and laser scanning confocal microscopy were used to detect GLT25D2 expression, and subsets of CD4+T cells was analyzed by flow cytometry. A total of 26 cytokines in serum samples were determined using Luminex technology. RESULTS: The trend in liver injury score variation was consistent with serum alanine aminotransferase and aspartate aminotransferase levels. The levels of interleukin 4 (IL-4), IL-1ß, IL-9, and several chemokines such as macrophage inflammatory protein-2, eotaxin, and growth-related oncogene α were significantly increased in Glt25d2-/- mice compared with WT mice after Con A challenge. A further phenotype analysis of primary Glt25d2-/- CD4+ T cells showed that Glt25d2 knockout increased the frequency of the CD25+CD69- subset but decreased the frequency of the CD25-CD69+ subset after Con A challenge for 6, 12, and 24 h compared with those of WT CD4+ T cells. Activation-induced apoptosis was also significantly increased in Glt25d2-/- CD4+ T cells after Con A challenge compared with WT CD4+ T cells. Lectin microarray hybridization showed that Glt25d2 knockout increased the binding activity of Narcissus pseudonarcissus lectin to CD4+ T cells but Amaranthus caudatus lectin-binding activity was lost during Con A challenge. CONCLUSION: The present results suggest that collagen glycosylation mediated by GLT25D2 may regulate a subset of CD4+ T cells and be involved in the pathogenesis of Con A-induced hepatitis.

The LPS Responsiveness in BN and LEW Rats and Its Severity Are Modulated by the Liver.

Differences in LPS responsiveness influence the outcome of patients with sepsis. The intensity of the response is highly variable in patients and strain dependent in rodents. However, the role of the liver for initiating the LPS response remains ill defined. We hypothesize that hepatic LPS uptake is a key event for initiating the LPS response. In the present study, the severity of the LPS-induced inflammatory response and the hepatic LPS uptake was compared in two rat strains (Lewis (LEW) rats and Brown Norway (BN) rats). Using a transplantation model, we demonstrated the decisive role of the liver. The expression of hepatic TNF-α, IL-6, and IL-1ß mRNA levels in BN rats was significantly lower than that in LEW rats. LEW rats were sensitized to LPS via G-CSF pretreatment. Sensitization caused by G-CSF pretreatment induced severe liver injury and mortality in LEW rats, but not in BN rats (survival rate: 0% (LEW) versus 100% (BN), p < 0.01). LEW rats presented with higher liver enzymes, more alterations in histology, and higher expression of caspase 3 and higher cytokines levels. One of the reasons could be the increased hepatic LPS uptake, which was only observed in LEW but not in BN livers. Using the transplantation model revealed the decisive role of the LPS responsiveness of the liver. Injection of LPS to the high-responding LEW recipient before transplantation of a low-responder BN liver resulted in a 50% survival rate. In contrast, injecting the same dose of LPS into the high-responding LEW recipient after transplanting the low-responding BN liver resulted in a 100% survival rate. The severity of inflammatory response in different strains might be related to the differences in hepatic LPS uptake. This observation suggests that the liver plays a genetically defined decisive role in modulating the inflammatory severity.

Critical Contribution of NK Group 2 Member D Expressed on Invariant Natural Killer T Cells in Concanavalin A-Induced Liver Hepatitis in Mice.

Natural killer group 2D (NKG2D) is a well-characterized activating receptor expressed on many immune cells, including invariant natural killer T (iNKT) cells. These cells were shown to be responsible of liver injury in the model of concanavalin A (Con A)-induced hepatitis, considered to be an experimental model of human autoimmune hepatitis. In this study, we investigated whether NKG2D plays a role in the hepatitis induced by iNKT cell-mediated immune response to Con A. By using killer cell lectin-like receptor subfamily K, member 1 deficient (Klrk1-/-) mice, we found that the absence of NKG2D reduced the hepatic injury upon Con A administration. This was not due to an intrinsic functional defect of NKG2D-deficient iNKT cells as mice missing NKG2D have normal distribution and function of iNKT cells. Furthermore, increased resistance to Con A-induced hepatitis was confirmed using neutralizing anti-NKG2D antibodies. The reduced pathogenic effect of Con A in the absence of NKG2D correlates with a reduction in pathogenic cytokine production and FAS-Ligand (FAS-L) expression by iNKT cells. We also found that Con A administration led to an increase in the retinoic acid early inducible (RAE-1) surface expression on wild-type hepatocytes. Finally, we found that Con A has no direct action on FAS-L expression or cytokine production by iNKT cells and thus propose that NKG2D-L expression on stressed hepatocytes promote cytotoxic activity of iNKT cells via its interaction with NKG2D contributing to hepatic injury. In conclusion, our results highlight NKG2D as an essential receptor required for the activation of iNKT cells in Con A-induced hepatitis and indicate that it represents a potential drug target for prevention of autoimmune hepatitis.

Antihistone Autoantibodies in Dobermans With Hepatitis.

BACKGROUND: Immune system involvement is suggested as an underlying cause for Doberman hepatitis (DH) based on female predisposition, lymphocyte infiltration, abnormal hepatocyte expression of major histocompatibility complex class II antigens, and homozygosity for dog leukocyte antigen DRB1*00601. OBJECTIVE: To measure serum antinuclear antibodies (ANA) and serum antihistone antibodies (AHA) in Dobermans with hepatitis. To determine whether increased serum ANA or serum AHA could be used to support the diagnosis of Doberman hepatitis (DH). ANIMALS: Privately owned 25 subclinically and 13 clinically affected DH Dobermans and 17 healthy control Dobermans. METHODS: Case-control study. Indirect immunofluorescence (IIF) microscopy and line blot tests were employed for the ANA pilot studies and an enzyme-linked immunosorbent assay (ELISA) assay for detection of IgG AHA. RESULTS: Indirect immunofluorescence revealed ANA-positive cases, and line blot showed AHA reactivity. In ELISA, importantly increased concentrations of AHA were found in 92% (23/25) of dogs in the subclinical stage and 84.6% (11 of 13) of dogs in the clinical stage of DH compared with no control dogs (0/17) (P < 0.0005). The mean AHA absorbance values of the blood samples obtained from the 25 subclinical DH dogs (1.36 ± 0.60, mean ± SD) and the 13 clinically affected dogs (1.46 ± 0.49) were significantly higher than in 17 control dogs (0.51 ± 0.18; P < 0.0001). CONCLUSIONS AND CLINICAL IMPORTANCE: As the presence of AHA indicates autoimmune activity, our results favor an autoimmune background as one cause for DH. Antihistone antibody could represent a novel means for screening Dobermans with increased serum alanine transaminase concentrations and suspicion of DH.

Identification of Glyceraldehyde-3-Phosphate and Alcohol Dehydrogenases as Autoantigens in Doberman Hepatitis.

An autoimmune background is suspected for Doberman hepatitis (DH). It is based on the finding of mononuclear cell infiltrates in the liver, strong female bias, association to the homozygous risk factor dog leucocyte antigen (DLA) allele DRB1*00601 and aberrant major histocompatibility complex (MHC) class II expression on hepatocytes that correlates with the degree of inflammation in the liver. The aim of this study was to search for autoantibodies against liver-related antigens associated with DH. Twenty-five Dobermans with subclinical DH (SDH), 13 that clinically manifest DH (CDH) and 17 healthy controls were studied. Immunoblotting analysis detected specific antibodies in the DH sera. By mass spectrometry the targets were identified as liver-related enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and alcohol dehydrogenase (ADH). Using ELISA, anti-GAPDH IgG was detected in 36% (9/25) of SDH dogs and 69.2% (9/13) of the CDH dogs compared to healthy controls (0/17) (P < 0.0005). Anti-ADH IgG was detected in 72% (18/25) of SDH dogs and 76.9% (10/13) of CDH dogs and only in one (1/17) control (P < 0.0005). The finding of novel autoantigens, GAPDH and ADH strengthen the hypothesis that DH is an autoimmune disease of the liver. These findings suggest that DH could be diagnosed by screening for autoantibodies against the defined antigens.

Sibiriline, a new small chemical inhibitor of receptor-interacting protein kinase 1, prevents immune-dependent hepatitis.

Necroptosis is a regulated form of cell death involved in several disease models including in particular liver diseases. Receptor-interacting protein kinases, RIPK1 and RIPK3, are the main serine/threonine kinases driving this cell death pathway. We screened a noncommercial, kinase-focused chemical library which allowed us to identify Sibiriline as a new inhibitor of necroptosis induced by tumor necrosis factor (TNF) in Fas-associated protein with death domain (FADD)-deficient Jurkat cells. Moreover, Sib inhibits necroptotic cell death induced by various death ligands in human or mouse cells while not protecting from caspase-dependent apoptosis. By using competition binding assay and recombinant kinase assays, we demonstrated that Sib is a rather specific competitive RIPK1 inhibitor. Molecular docking analysis shows that Sib is trapped closed to human RIPK1 adenosine triphosphate-binding site in a relatively hydrophobic pocket locking RIPK1 in an inactive conformation. In agreement with its RIPK1 inhibitory property, Sib inhibits both TNF-induced RIPK1-dependent necroptosis and RIPK1-dependent apoptosis. Finally, Sib protects mice from concanavalin A-induced hepatitis. These results reveal the small-molecule Sib as a new RIPK1 inhibitor potentially of interest for the treatment of immune-dependent hepatitis.

Chlorogenic acid ameliorated concanavalin A-induced hepatitis by suppression of Toll-like receptor 4 signaling in mice.

Chlorogenic acid (CGA), one of the most abundant dietary polyphenolic compounds, has been reported to exhibit anti-inflammatory ability. However, the hepatoprotective effects and molecular mechanisms of CGA on concanavalin A (Con A)-induced hepatitis have not been explored. In the present study, we found that pretreatment with CGA dose-dependently inhibited the elevation of plasma aminotransferases and alleviated hepatic pathological damage as well as hepatocyte apoptosis in Con A-exposed mice. Additionally, CGA markedly suppressed the production of serum tumor necrosis factor (TNF)-α and interferon (IFN)-γ, alleviated the infiltration of hepatic macrophages, neutrophils, and activated CD4+ T lymphocytes in Con A-primed mice. Moreover, CGA downregulated Con A-induced hepatic expression of adhesion molecules (ICAM-1, VCAM-1 and ELAM-1) mRNA and protein, and inhibited Con A-activated Toll-like receptor (TLR) 4 signal molecules including TLR4, p-IRAK1, p-IκB, and p-p38. Finally, our results also showed that CGA exhibited a therapeutic effect, which CGA posttreatment improved hepatic damage at 1, 3, and 6h after Con A. Taken together, these data suggested that CGA could effectively prevent mice from Con A-induced hepatitis, which might be through suppressing the activation of TLR4 signaling, downregulating the expression of adhesion molecules, and alleviating the infiltration and activation of hepatic leukocytes and the production of pro-inflammatory cytokines.

Celecoxib attenuates depressive-like behavior associated with immunological liver injury in C57BL/6 mice through TNF-α and NF-κb dependent mechanisms.

AIM: Depression associating patients with chronic liver diseases is a major treatment goal. This study aimed to evaluate the potential hepatoprotective and antidepressant effects of celecoxib in a model of experimental autoimmune hepatitis (EAH) and depressive-like behavior in C57BL/6 mice. MAIN METHODS: EAH was induced by immunization with S-100 liver antigen emulsified in complete Freund's adjuvant (CFA). Mice were randomly allocated to 5 groups; control phosphate buffered saline group; control CFA group; EAH group, and 2 groups of EAH plus celecoxib (7.5 or 15mg/kg/d respectively). Mice were assessed behaviorally by novelty-suppressed test, tail suspension test, locomotor assessment and forced swimming tests. Serum liver enzymes and hepatic hydroxyproline content were biochemically analyzed. Histopathological analysis for liver and brain sections and immunohistochemical studies for hepatic and hippocampal tumor necrosis factor (TNF-α), nuclear factor Kappa-B (NF-κB) and caspase-3 were performed. KEY FINDINGS: EAH group exhibited significant depressive-like changes, increase in liver enzymes and hepatic hydroxyproline content. Signs of autoimmune hepatitis and structural changes in hippocampus were confirmed by histopathological studies. Immunohistochemical examination revealed overexpression of hepatic and hippocampal TNF-α, NF-κB and caspase-3 positive cells. Celecoxib (7.5mg/kg/d) significantly ameliorated hepatic biochemical changes, hepatic and hippocampal histopathological and immunohistochemical changes induced in EAH group. Celecoxib (15mg/kg/d) significantly ameliorated the behavioral changes, histopathological and immunohistochemical changes in hippocampus, with non-significant change in hepatic biochemical profile, histopathological and immunohistochemical changes induced in EAH group. SIGNIFICANCE: The celecoxib (7.5mg/kg/d) through its anti-inflammatory effect may represent a new therapeutic approach to treat autoimmune hepatitis associated with depressive symptoms.

Cre-inducible human CD59 mediates rapid cell ablation after intermedilysin administration.

Cell ablation is a powerful tool for studying cell lineage and/or function; however, current cell-ablation models have limitations. Intermedilysin (ILY), a cytolytic pore-forming toxin that is secreted by Streptococcus intermedius, lyses human cells exclusively by binding to the human complement regulator CD59 (hCD59), but does not react with CD59 from nonprimates. Here, we took advantage of this feature of ILY and developed a model of conditional and targeted cell ablation by generating floxed STOP-CD59 knockin mice (ihCD59), in which expression of human CD59 only occurs after Cre-mediated recombination. The administration of ILY to ihCD59+ mice crossed with various Cre-driver lines resulted in the rapid and specific ablation of immune, epithelial, or neural cells without off-target effects. ILY had a large pharmacological window, which allowed us to perform dose-dependent studies. Finally, the ILY/ihCD59-mediated cell-ablation method was tested in several disease models to study immune cell functionalities, hepatocyte and/or biliary epithelial damage and regeneration, and neural cell damage. Together, the results of this study demonstrate the utility of the ihCD59 mouse model for studying the effects of cell ablation in specific organ systems in a variety of developmental and disease states.

Opposing role of tumor necrosis factor receptor 1 signaling in T cell-mediated hepatitis and bacterial infection in mice.

UNLABELLED: Death receptor (DR) ligands such as tumor necrosis factor (TNF) have been identified as fundamental mediators of liver damage both in mouse models and in humans. While the essential site of function of DR signaling is conceivably the hepatocyte, a systematic analysis is missing. Using mice with conditional gene ablation, we analyzed the tissue-specific function of DR signaling in T cell-dependent (concanavalin A) and independent (lipopolysaccharide/galactosamine) hepatitis and in models of bacterial infection (Listeria monocytogenes, lipopolysaccharide). We report that lipopolysaccharide/galactosamine-induced liver injury depends on hepatocyte-intrinsic TNF receptor 1 (p55, TNFR1). In contrast, we show that T cell-induced hepatitis was independent of TNFR1 signaling in hepatocytes, T cells, or endothelial cells. Moreover, T cell-induced hepatitis was independent of hepatocyte-intrinsic Fas-associated protein with death domain, TNF-related apoptosis-inducing ligand receptor, or Fas signaling. Instead, concanavalin A-induced hepatitis was completely prevented in mice with myeloid-derived cell (MDC)-specific deletion of TNFR1. Significantly, however, mice lacking TNFR1 in MDCs succumbed to listeria infection, although they displayed similar sensitivity toward endotoxin-induced septic shock when compared to control mice. These results suggest that TNFR1 signaling in MDCs is a critical mediator of both the detrimental and the protective functions of TNF in T cell-induced hepatitis and bacterial infection, respectively. CONCLUSION: The critical site of action of DRs is completely dependent on the nature of hepatitis; the data specify MDCs as the essential cell type of TNFR1 function in T cell-mediated hepatitis and in the response to listeria, thereby identifying the opposing role of MDC TNFR1 in autoimmunity and bacterial infection. (Hepatology 2016;64:508-521).

MORPHOLOGICAL CHANGES IN MICE LIVER IN DYNAMICS OF CONCANAVALIN A - INDUCED HEPATITIS.

The injure of the liver tissue and its infiltration by cells of the innate and adaptive immunity in dynamics of Con A-induced hepatitis in mice was studied. The semiquantitative method of damage rate of microcirculation channel and liver parenchyma was used, leukocyte liver infiltration and cellular composition of infiltrates were investigated also. Primary liver reaction to the Con-A was the inflammatory changes in the vascular bed, followed by disturbances in the parenchyma.The sufficient increasing of leukocyte migration to the liver was revealed. Besides, the neutrophile infiltration was increased first with a maximum at 6 hours of the experiment (63,9 ±4,6%, p<0,001 to the control level) ,and then the lymphocyte infiltration was increased with creation of manycellular lymphocytemacrophage infiltrates (62% at 48 hours comparing to 6 hours of experiment) and sufficient quantity of plasma cells population (4,9%, p<0,05 comparing to 6 hours of experiment). The obtained data gives the base to suggest that the elevated infiltration of liver tissue by leukocytes, particularly by lymphocytes and monocytes, together with necrotic death increasing creats the conditions for effective intracellular interaction and immune response to autoantigenes. This can be the essential pathogenic mechanism of development of autoimmune liver deseases.

STK4 regulates TLR pathways and protects against chronic inflammation-related hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is frequently associated with pathogen infection-induced chronic inflammation. Large numbers of innate immune cells are present in HCCs and can influence disease outcome. Here, we demonstrated that the tumor suppressor serine/threonine-protein kinase 4 (STK4) differentially regulates TLR3/4/9-mediated inflammatory responses in macrophages and thereby is protective against chronic inflammation-associated HCC. STK4 dampened TLR4/9-induced proinflammatory cytokine secretion but enhanced TLR3/4-triggered IFN-ß production via binding to and phosphorylating IL-1 receptor-associated kinase 1 (IRAK1), leading to IRAK1 degradation. Notably, macrophage-specific Stk4 deletion resulted in chronic inflammation, liver fibrosis, and HCC in mice treated with a combination of diethylnitrosamine (DEN) and CCl4, along with either LPS or E. coli infection. STK4 expression was markedly reduced in macrophages isolated from human HCC patients and was inversely associated with the levels of IRAK1, IL-6, and phospho-p65 or phospho-STAT3. Moreover, serum STK4 levels were specifically decreased in HCC patients with high levels of IL-6. In STK4-deficient mice, treatment with an IRAK1/4 inhibitor after DEN administration reduced serum IL-6 levels and liver tumor numbers to levels similar to those observed in the control mice. Together, our results suggest that STK4 has potential as a diagnostic biomarker and therapeutic target for inflammation-induced HCC.