Feng Hong

Interleukin 22 (IL‐22) plays a protective role in T cell‐mediated murine hepatitis: IL‐22 is a survival factor for hepatocytes via STAT3 activation

The central role of T cell activation in hepatocellular injury has been well documented. In this article, we provide evidence suggesting that T cells may also play a protective role in liver disease by releasing interleukin‐22 (IL‐22), a recently identified T cell‐derived cytokine whose biological significance is unclear. IL‐22 messenger RNA and protein expression are significantly elevated in T cell‐mediated hepatitis induced by concanavalin A (ConA) but are less extensively elevated in the carbon tetrachloride‐induced liver injury model. Activated CD3+ T cells are likely responsible for the production of IL‐22 in the liver after injection of ConA. The IL‐22 receptor is normally expressed at high levels by hepatocytes and further induced after ConA injection. IL‐22 blockade with a neutralizing antibody reduces signal transducer and activator of transcription factor 3 (STAT3) activation and worsens liver injury in T cell‐mediated hepatitis, whereas injection of recombinant IL‐22 attenuates such injury. In vitro treatment with recombinant IL‐22 or overexpression of IL‐22 promotes cell growth and survival in human hepatocellular carcinoma HepG2 cells. Stable overexpression of IL‐22 in HepG2 cells constitutively activates STAT3 and induces expression of a variety of antiapoptotic (e.g., Bcl‐2, Bcl‐xL, Mcl‐1) and mitogenic (e.g., c‐myc, cyclin D1, Rb2, CDK4) proteins. Blocking STAT3 activation abolishes the antiapoptotic and mitogenic actions of IL‐22 in hepatic cells. In conclusion, the T cell‐derived cytokine IL‐22 is a survival factor for hepatocytes; this suggests that T cell activation may also prevent and repair liver injury by releasing hepatoprotective cytokine IL‐22 in addition to its previously documented central role in hepatocellular injury. (HEPATOLOGY 2004;39:1332–1342.)

Opposing roles of STAT1 and STAT3 in T cell–mediated hepatitis: regulation by SOCS

T cell-mediated fulminant hepatitis is a life-threatening event for which the underlying mechanism is not fully understood. Injection of concanavalin A (Con A) into mice recapitulates the histological and pathological sequelae of T cell-mediated hepatitis. In this model, both signal transducer and activator of transcription factor 1 (STAT1) and STAT3 are activated in the liver. Disruption of the STAT1 gene by way of genetic knockout attenuates liver injury, suppresses CD4(+) and NK T cell activation, and downregulates expression of proapoptotic interferon regulatory factor-1 protein and suppressor of cytokine signaling-1 (SOCS1), but enhances STAT3 activation and STAT3-controlled antiapoptotic signals. Studies from IFN-gamma-deficient mice indicate that IFN-gamma not only is the major cytokine responsible for STAT1 activation but also partially accounts for STAT3 activation. Moreover, downregulation of STAT3 activation in IL-6-deficient mice is associated with decreased STAT3-controlled antiapoptotic signals and expression of SOCS3, but upregulation of STAT1 activation and STAT1-induced proapoptotic signals and exacerbation of liver injury. Taken together, these findings suggest that STAT1 plays a harmful role in Con A-mediated hepatitis by activation of CD4(+) and NK T cells and directly inducing hepatocyte death, whereas STAT3 protects against liver injury by suppression of IFN-gamma signaling and induction of antiapoptotic protein Bcl-X(L). STAT1 and STAT3 in hepatocytes also negatively regulate one another through the induction of SOCS.

Cannabinoid-2 receptor mediates protection against hepatic ischemia/reperfusion injury

Hepatic ischemia‐reperfusion (I/R) injury continues to be a fatal complication that can follow liver surgery or transplantation. We have investigated the involvement of the endocannabinoid system in hepatic I/R injury using an in vivo mouse model. Here we report that I/R triggers several‐fold increases in the hepatic levels of the endocannabinoids anandamide and 2‐arachidonoylglycerol, which originate from hepa‐tocytes, Kupffer, and endothelial cells. The I/R‐in‐duced increased tissue endocannabinoid levels positively correlate with the degree of hepatic damage and serum TNF‐α, MIP‐1α, and MIP‐2 levels. Furthermore, a brief exposure of hepatocytes to various oxidants (H2O2 and peroxynitrite) or inflammatory stimuli (endotoxin and TNF‐α) also increases endocannabinoid levels. Activation of CB2 cannabinoid receptors by JWH133 protects against I/R damage by decreasing inflammatory cell infiltration, tissue and serum TNF‐α, MIP‐1a and MIP‐2 levels, tissue lipid peroxidation, and expression of adhesion molecule ICAM‐1 in vivo. JWH133 also attenuates the TNF‐a‐induced ICAM‐1 and VCAM‐1 expression in human liver sinusoidal endothelial cells (HLSECs) and the adhesion of human neutrophils to HLSECs in vitro. Consistent with the protective role of CB2 receptor activation, CB2_/_ mice develop increased I/R‐induced tissue damage and proinflammatory phenotype. These findings suggest that oxidative/nitrosative stress and inflammatory stimuli may trigger endocannabinoid production, and indicate that targeting CB2 cannabinoid receptors may represent a novel protective strategy against I/R injury. We also demonstrate that CB2−/− mice have a normal hemodynamic profile.—Bátkai, S., Osei‐Hyiaman, D., Pan, H., El‐Assal, O., Rajesh, M., Mukhopadhyay, P., Hong, F., Harvey‐White, J., Jafri, A., Hasko, G., Huffman, J. W., Gao, B., Kunos, G., Pacher, P. Cannabi‐noid‐2 receptor mediates protection against hepatic ischemia/reperfusion injury. FASEB J. 21, 1788–1800 (2007)

Interleukin 6 alleviates hepatic steatosis and ischemia/reperfusion injury in mice with fatty liver disease

Fatty liver, formerly associated predominantly with excessive alcohol intake, is now also recognized as a complication of obesity and an important precursor state to more severe forms of liver pathology including ischemia/reperfusion injury. No standard protocol for treating fatty liver exists at this time. We therefore examined the effects of 10 days of interleukin 6 (IL‐6) injection in 3 murine models of fatty liver: leptin deficient ob/ob mice, ethanol‐fed mice, and mice fed a high‐fat diet. In all 3 models, IL‐6 injection decreased steatosis and normalized serum aminotransferase. The beneficial effects of IL‐6 treatment in vivo resulted in part from an increase in mitochondrial β oxidation of fatty acid and an increase in hepatic export of triglyceride and cholesterol. However, administration of IL‐6 to isolated cultured steatotic hepatocytes failed to decrease lipid contents, suggesting that the beneficial effects of IL‐6 in vivo do not result from its effects on hepatocytes alone. IL‐6 treatment increased hepatic peroxisome proliferator‐activated receptor (PPAR) α and decreased liver and serum tumor necrosis factor (TNF) α. Finally, 10 days of treatment with IL‐6 prevented the susceptibility of fatty livers to warm ischemia/reperfusion injury. In conclusion, long‐term IL‐6 administration ameliorates fatty livers and protects against warm ischemia/reperfusion fatty liver injury, suggesting the therapeutic potential of IL‐6 in treating human fatty liver disease. Supplementary material for this article can be found on the Hepatology website (http://interscience.wiley.com/jpages/0270‐9139/suppmat/index.html). (HEPATOLOGY 2004;40:933–941.)

Host factors and failure of interferon‐α treatment in hepatitis C virus

Failure of interferon‐α (IFN‐α) treatment in patients with chronic hepatitis C virus (HCV) infection is a challenging obstacle for clinical and experimental hepatology. Both viral and host factors have been implicated in reducing responsiveness to IFN‐α therapy. The role of viral factors has been studied extensively and has been summarized in several review articles; however, much less attention has been paid to host factors. In this paper, we review evidence of host factor involvement in IFN‐α treatment failure. We discuss possible underlying mechanisms responsible for these effects. Potential therapeutic strategies to enhance the effectiveness of IFN‐α therapy for HCV are also proposed. (HEPATOLOGY 2004;39:880ndash;890.)

Crucial Role of IL-4/STAT6 in T Cell-Mediated Hepatitis: Up-Regulating Eotaxins and IL-5 and Recruiting Leukocytes

T cell-mediated immune responses are implicated in the pathogenesis of a variety of liver disorders; however, the underlying mechanism remains obscure. Con A injection is a widely accepted mouse model to study T cell-mediated liver injury, in which STAT6 is rapidly activated. Disruption of the IL-4 and STAT6 gene by way of genetic knockout abolishes Con A-mediated liver injury without affecting IFN-γ/STAT1, IL-6/STAT3, or TNF-α/NF-κB signaling or affecting NKT cell activation. Infiltration of neutrophils and eosinophils in Con A-induced hepatitis is markedly suppressed in IL-4 −/− and STAT6−/− mice compared with wild-type mice. IL-4 treatment induces expression of eotaxins in hepatocytes and sinusoidal endothelial cells isolated from wild-type mice but not from STAT6−/− mice. Con A injection induces expression of eotaxins in the liver and elevates serum levels of IL-5 and eotaxins; such induction is markedly attenuated in IL-4−/− and STAT6−/− mice. Finally, eotaxin blockade attenuates Con A-induced liver injury and leukocyte infiltration. Taken together, these findings suggest that IL-4/STAT6 plays a critical role in Con A-induced hepatitis, via enhancing expression of eotaxins in hepatocytes and sinusoidal endothelial cells, and induces IL-5 expression, thereby facilitating recruitment of eosinophils and neutrophils into the liver and resulting in hepatitis.

Elevated interleukin-6 during ethanol consumption acts as a potential endogenous protective cytokine against ethanol-induced apoptosis in the liver: involvement of induction of Bcl-2 and Bcl-x(L) proteins.

Elevation of serum interleukin-6 (IL-6) levels is always associated with alcoholic liver disease (ALD), but the significance of such elevation is not clear. Here we show that chronic ethanol consumption induces significant apoptosis in the liver of IL-6 (−/−) mice but not IL-6 (+/+) mice. IL-6 (−/−) hepatocytes are more susceptible to ethanol- and tumor necrosis factor α- (TNFα-) induced apoptotic killing, which can be corrected by IL-6. Expression of both anti-apoptotic (such as Bcl-2 and Bcl-xL) and proapoptotic (such as Bax) proteins is markedly elevated in the liver of human ALD and chronically ethanol-fed IL-6 (+/+) mice. On the contrary, induction of Bcl-2 and Bcl-xL is not observed in the liver of chronically ethanol-fed IL-6 (−/−) mice, whereas expression of Bax protein remains elevated. Injection of IL-6 markedly induces expression of Bcl-2 and Bcl-xL but not Bax in the liver. Finally, high concentrations of ethanol inhibit IL-6-activated anti-apoptotic signal, but increasing the concentrations of IL-6 is able to overcome such inhibitory effect. These findings suggest that elevated serum IL-6 levels in ALD may overcome the inhibitory effect of ethanol on IL-6-mediated anti-apoptotic signals and prevent alcohol-induced hepatic apoptosis by induction of Bcl-2 and Bcl-xL.

Additive activation of hepatic NF-kappaB by ethanol and hepatitis B protein X (HBX) or HCV core protein: involvement of TNF-alpha receptor 1-independent and -dependent mechanisms.

Alcohol consumption and viral hepatitis infection synergistically accelerate liver injury, but the underlying mechanism is not fully understood. Here we have examined the effects of ethanol on hepatitis B protein X (HBX)- or hepatitis C core protein (HCV core protein)-mediated activation of NF-kappaB, a critical signal in hepatic injury, regeneration, and tumor transformation. Acute ethanol or acetaldehyde exposure potentiates HBX or HCV core protein activation of NF-kappaB in primary mouse hepatocytes. Such potentiation can be abolished by blocking ethanol metabolism or overexpression of dominant negative NF-kappaB-inducing kinase (NIK), IkappaB kinase (IKK), or IkappaB. Moreover, pertussis toxin attenuates NF-kappaB activation induced by acetaldehyde but not by HBX or HCV core protein, whereas HBX or HCV core protein-mediated activation of NF-kappaB is abolished completely in tumor necrosis factor a receptor 1 (TNFR1) (-/-) hepatocytes. Finally, chronic ethanol consumption induces hepatic CYP2E1 protein expression and potentiates HBX or HCV core protein activation of NF-kappaB in the liver. These findings suggest that ethanol activates hepatic NF-kappaB via its metabolism and that HBX or HCV core protein activates hepatic NF-kappaB via TNFR1. With the essential role of TNFR1 in alcoholic liver injury, targeting TNFR1 by hepatitis viral proteins could contribute to cooperative effects of alcohol consumption and viral hepatitis on liver disease.

Tumor necrosis factor α attenuates interferon α signaling in the liver: involvement of SOCS3 and SHP2 and implication in resistance to interferon therapy1

Although interferon α (IFN‐α) has been used for a decade to treat viral hepatitis, a disease that affects millions of people worldwide, more than 60% of viral hepatitis patients respond poorly. It has been reported that high levels of tumor necrosis factor α (TNF‐α) correlated highly with resistance to IFN‐α therapy. Here we demonstrate that injection of TNF‐α suppresses IFN‐α signaling and markedly induces expression of suppressor of cytokine signaling 3 (SOCS3) and SH2 containing protein‐tyrosine phosphatase 2 (SHP2) in the liver. TNF‐α induction of SOCS3 and SHP2 remains unchanged while induction of STAT1 protein expression is completely abolished in IL‐6‐deficient mice. Immunoprecipitation experiments show that injection of TNF‐α increases SHP2 association with JAKs. Overexpression of SOCS3 and SHP2 inhibits IFN‐α signaling in hepatic cells. Injection of carbon tetrachloride, which is known to induce TNF‐α in the liver, attenuates IFN‐α signaling in the liver. This attenuation is also observed in TNF‐α receptor II‐ (TNF‐R2‐) deficient mice but is markedly diminished in TNF‐R1‐deficient mice. Taken together, these findings suggest that TNF‐α may be involved in resistance to IFN‐α therapy by induction of SOCS3 and SHP2, and they could be therapeutic targets for improving the efficacy of IFN‐α therapy.

Additive activation of hepatic NF-kB by ethanol and HBX or HCV core protein: involvement of TNF-a receptor I-independent and -dependent mechanisms

Alcohol consumption and viral hepatitis infection synergistically accelerate liver injury, but the underlying mechanism is not fully understood. Here we have examined the effects of ethanol on hepatitis B protein X (HBX)- or hepatitis C core protein (HCV core protein)-mediated activation of NF-kappaB, a critical signal in hepatic injury, regeneration, and tumor transformation. Acute ethanol or acetaldehyde exposure potentiates HBX or HCV core protein activation of NF-kappaB in primary mouse hepatocytes. Such potentiation can be abolished by blocking ethanol metabolism or overexpression of dominant negative NF-kappaB-inducing kinase (NIK), IkappaB kinase (IKK), or IkappaB. Moreover, pertussis toxin attenuates NF-kappaB activation induced by acetaldehyde but not by HBX or HCV core protein, whereas HBX or HCV core protein-mediated activation of NF-kappaB is abolished completely in tumor necrosis factor a receptor 1 (TNFR1) (-/-) hepatocytes. Finally, chronic ethanol consumption induces hepatic CYP2E1 protein expression and potentiates HBX or HCV core protein activation of NF-kappaB in the liver. These findings suggest that ethanol activates hepatic NF-kappaB via its metabolism and that HBX or HCV core protein activates hepatic NF-kappaB via TNFR1. With the essential role of TNFR1 in alcoholic liver injury, targeting TNFR1 by hepatitis viral proteins could contribute to cooperative effects of alcohol consumption and viral hepatitis on liver disease.