The Role Of Z39Ig In The Pathogenesis Of Chronic HBV Infection And Its Immunoregulatory Function In Hepatic Immune Tolerance

The liver is the largest digestive gland in the adult body and is responsible for intaking, metabolism, detoxification and storage of nutritional substances obtained from food. In addition, liver is also a special immune organ, which is responsible for the removal of pathogenic microorganisms invaded from gastrointestinal tract and some metabolic components. All kinds of antigenic material is processed by antigenic presenting cells (APCs) and presented to circulating T cells in liver all the time, which decides there must be precise regulatory mechanisms in controlling T cell immune response. Under physiological conditions, T cell function in normal to maintain liver and even systemic immune balance. Under pathological conditions, T cells response to self antigens will result in the occurrence of autoimmune hepatic disease, if not an effective response to foreign antigens will lead to chronic infection or cancer.The liver is the organ susceptible to immune tolerance. So far, liver transplantation is the most likely to survive in organ transplant. Even in the very early, it was found that the transplanted liver can survive without any pretreatment of immunosuppressants instead of occurring tansplant rejection. Strikingly, the transplantation of a kidney and a liver from the same donor enhanced the survival of the kidney. However, the liver susceptibe to immune tolerance is characterized by the double-edged sword. When the liver is infected with some viral such as HBV and HCV, and parasites such as schistosomiasis, antigen-specific T cells often occurs tolerance in liver, which makes the infections often become chronicity, eventually leading to severe liver pathological changes.It is accepted that T cell differentiation, activation and effects requires not only T cell receptor (TCR)-mediated antigen-specific signaling but also a costimulatory signal provided by antigen presenting cells (APCs). Accumulating evidence have shown that co-stimulatory signal play an improtant role in regulating T cell response. Reasonably closing or enhancing these costimulatory pathway has been applied to tumor, viral and parasitic infections, transplantation rejection and other fields of experimental treatment research.Costimulatory molecules can be divided into two categories, one can induce and enhance T cell immune response to antigens, such as B7.1 and B7.2, and the other can induce T cell tolerance, also known as co-inhibitory molecules such as B7-H1 and B7-DC,etc. The role co-inhibitory molecules in induction of immune tolerance in liver has become the topic research in recent years.Z39Ig (also called CRIg or VSIG4), a recently identified member of the B7 superfamily, is a negative regulator of T cell activation. Z39Ig protein is restricted to the surface of peripheral blood monocyte-derived macrophages (MDMs) and macrophages in certain tissues, especially expressed on Kupffer cells. Z39Ig can inhibits CD4+ as well as CD8+ T cell proliferation and IL-2 production through ligating an unidentified receptor, which exerts a greater inhibitory effect on the T cell response than both PD-L1 and PD-L2. In animal model of autoimmune disease, the level of IFN-γproduced by antigen-specific T cells was decreased by giving Z39Ig-Fc protein, which suggested that Z39Ig maybe play a role in inducing peripheral tolerance. Then, whether Z39Ig plays a role in maitaining immune tolerance of liver or not?In this study, we investigated the role of Z39Ig in the pathogenesis of chronic HBV infection firstly. Thirty-nine liver biopsy specimens from CHB patients were collected.We found Z39Ig expression was restricted to macrophages, and was absent on T cells, B cells, dendritic cells and sinusoidal endothelial cells. Immunihistochemical staining showed that Z39Ig expression was decreased in the liver tissues of CHB patients compared with healthy controls, especially in IA patients who exhibit more severe inflammation in liver. Statistical analysis showed that macrophage-associated Z39Ig expression was positively correlated with plasma HBV load but had a negative relation to serum alanine aminotransaminase levels, which suggested Z39Ig maybe invovled in the pathogenesis of CHB and its expression was regulated by inflammatory stimuli. Then, we constructed lentivirus expressing Z39Ig and then infected 293FT. We investigated the effects of Z39Ig expressing 293FT on T cell proliferation and cytokines(IL-2 and IFN-γ) production. We found membrane bound Z39Ig inhibited T cell response indeed. We further explored the factors caused Z39Ig expression reduction. We found IFN-γwas the most obvious cytokine downregulated Z39Ig expression on MDMs in vitro. Further, we analyzed the relationship between Z39Ig mRNA expression and IFN-γin liver tissues of IA patients by quantitative real-time PCR, the results showed a negative correlation between them. The cytokine blocking experiments suggested that IFN-γoriginated from activated T cells maybe responsible for Z39Ig expression reduction. This part of results suggested Z39Ig expressed on macrophages in liver maybe involved in the pathogenesis of chronic HBV infection, and T cells can opposite T cell hyporesponsiveness through dampening Z39Ig inhibitory signals from macrophages and thus maintain their antiviral function in CHB.In order to clarify the possible mechanisms of Z39Ig involved in liver tolerance, we compared T lymphocytes from Z39Ig knockout mice and wild-type mice in the distribution and phenotype, the results showed there was no significant difference between them. Then, we established ConA-induced mouse model of autoimmune hepatitis in KO and WT mice. Whether the level of serum transaminase or the degree of liver injury, there was no significant difference between KO mice and WT mice, we also found no significant difference in the activity or the proportion of T cell subsets, which suggested the effect of Z39Ig expression was not obvious on immune status in acute hepatic inflammation. Further, we established ConA-induced mouse model of hepatic immune tolerance. We found tolerance was established in WT mice, resulting in diminished necrotic liver damage, ConA could still provoke infiltration of leukocytesinto liver tissues. But in KO mice, necrotic liver injury was pronounced and resulted in extended bridging necroses. There was an significant elevation of serum transaminase in KO mie compared with WT mice. Then, we found the ability of T cells in liver of KO mice in producing IL-2, TNF-αand IFN-γwas significantly increased compared with WT mice, the result of cytokines detected by quantitative real-time PCR corroborated the findings. This part of results suggestd that Z39Ig maybe play a role in liver immune tolerance. This study found macrophage-associated Z39Ig maybe play a role in the induction of liver immune tolerance, which maybe one of the factors leading to chronic HBV infection. To clarify the precise regulatory mechanism of Z39Ig signal maybe provide a new clue to understand the formation of immune tolerance of liver.