Immune Function Of LSECtin In HBV Infected Mice Model

The liver, as an immunologically privileged organ, bears the tolerance and immune dualfunction. The delicate and finely tuned balance between immunity and tolerance resultsdirectly from the complex interactions between the various liver-resident immune sentinelsand other peripheral leukocyte populations. Though immune tolerance (liver transplants) andimmune hyporesponsiveness (chronic viral infection) have been long recognized, only noware we beginning to understand the specific mechanisms. These properties render the liver anattractive target site for pathogens. Some pathogens (such as HBV) can escape immunecontrol and persist in hepatocytes.The liver serves as a target organ for hepatotropic viruses HBV, which can leads to chronichepatitis, cirrhosis and hepatocellular carcinoma, all of which pose a serious threat to humanhealth worldwide. Although the virus itself has no direct cytopathic effects on the hepatocytes.However, immune reaction, range from early inadequacy to later disorder, induced by chronicviral infection leads to great damage to liver. Thus, it is generally considered that the immuneresponse against the virus is a major cause of liver disease. On the other hand, the clearance ofthe virus is completely mediated by the immune response. Since the immune response is adouble-edged sword, we should consider more to maintain equilibrium when we intervenechronic infections.After HBV infection, active and effective removal of virus is dependent on the regulation ofinnate and secondary immune. CD4+T cells serves as master regulators of the adaptiveimmune response to HBV; CD8+T cells are the key cellular effectors mediating HBVclearance from the infected liver via cytotoxic and non-cytotoxic ways. Therefore, thefunction and status of CD8+T cells play a pivotal role in determining the viral infectionprogress. Unfortunately,during many persisting infections, CD8+T cells are often in a stateof dysfounction. The current study found that a variety of immunoregulatory pathways duringchronic viral infection. First, T cell intrinsic expression of inhibitory receptors such as PD-1and others can negatively regulate T cell function and limit the effectiveness of antiviralactivity. Second, immunoregulatory cytokines such as IL-10, TGF-β, and possibly additionalfactors produced by other cells in the environment can modulate and suppress antiviral T cellresponses. Third, regulatory cells can modulate antiviral effector T cells. These regulatorycells could include traditional FoxP3+natural or adaptive CD4+T regulatory cells (Tregs),regulatory CD8+T cells, and myeloid-derived suppressor cells. These cells could act throughcell-to-cell contact, inhibition of APC maturation, production of immunoregulatory cytokines, or direct inhibition of CD8+T cell effector function. Finally, changes in antigen-presentingcell (APC) usage or function could impact virus-specific CD8+T cells. The quality of T cellstimulation can also be impacted by changes in dendritic cell function, as well as differencesin antigen presentation by professional versus nonprofessional APC during persisting viralinfection. Integrated system composed of cells and molecules, weaving a complex set ofinhibitory networks, which result in CD8+T cells exhaustion, thereby contribute the persistentviral infection.The increasingly recognized therapeutic strategy for HBV infection is a combination ofan immune modulator and antiviral drugs,which may have potential to help these patientswith chronic infection. Merely inhibition viral replication can not recover the damagedimmune system. Reducing viral load, improving immunity, can break immune tolerance torestore the patient’s antiviral immune response and ultimately achieve lasting clearance ofvirus and restore the body’s protective immunity. Immunomodulatory methods including DCvaccine and cytokine-induced killer (CIK) and blockade of immune inhibitory molecules(PD-1, IL-10, LAG-3, TIM-3etc.). HBV-specific T cell dysfunction is one of the key reasonsto the failure to clearance pathogens. Since multiple cell-surface inhibitory receptors as acrucial factor cause the T cells exhaustion, it is an effective way to target immune inhibitorymolecules and blodckade them in order to restore the T cells function, particularly CD8+Tcells antiviral effect.LSECtin, typeⅡtransmembrane glycoprotein, located on human chromosome19p13.3, is anew member of the C-type lectin family, which including CD23, DC-SIGN and DC-SIGNR.LSECtin binds to Ebola virus, SARS coronavirus, filovirus, Lassa virus, and confers cellsusceptibility to Japanese encephalitis virus. Preliminary results from our laboratory showedthat LSECtin is a negative regulator of T-cell immune response. In a calcium-dependentmanner, LSECtin binds to N-linked glycoprotein on the activated T cell surface, which leadsto inhibition of T cell proliferation and decreased secretion of IFN-γ, IL-2. Thus negativelyregulates T cells immune response. This prompted us to investigate the immune roles ofLSECtin during HBV infection.We plan to develop acute HBV infection and chronic infection in a mouse model. Using thismodel, we can overexpress and block LSECtin proteins to address its immune responseduring HBV infection.HBV infected host tropism bias: human and chimpanze, and HBV transgenic mice borncentral tolerance, all of which have limited a systematic examination of the host-virus interactions during HBV infection in a small animal model. Until recently, Francis V. Chisariteam and Taiwan Pei-Jer Chen team were first developed a mouse model of acute HBVinfection and chronic HBV infection respectively. These models provided a invaluable tool.We try to establish these models. Afte optimization dose and testing, we have successfullyestablished a mouse model of acute HBV infection, in which Balb/c mice carried HBsAg fortwo weeks. At the same time, we also established a longer duration acute infection model thatantigen lasted up to1month in Balb/c mice. Finally, we have established a chronic infectionmode. In30%of the injected C57BL/6mice, HBV surface antigenemia persisted for>1year.Balb/c LSECtin-/-mice were employed to address the founction in the two HBV acuteinfection models. After HDI HBV plasmid, LSECtin-deficient mice had lower serum levels ofsurface antigen and E antigen and lesser core antigen positive hepatocytes and lower surfaceantigen positive rate. Complementary to KO, we overexpressed LSECtin in mice, whichdelayed clearance of antigen. In the context of a chronic model, C57BL/6LSECtin-/-miceaccelerated the clearance of antigen and had lower positive rate of serun HBsAg. Thephenotype was consistent with acute models’. In short, LSECtin can delay the clearance ofvirus antigens, may be as an accomplice of virus persistent infection.What mechanism(s) likely responssible for accelerated viral clearance during HBV infectionin LSECtin-deficient mice? As a negative regulator of adaptive immunity, LSECtin couldbind activated T cells, which may inhibition of T cells proliferation, decreasing T cells tosecrete IFN-γ, IL-2. Since the CD8+T cells have a crucial role in determining thedevelopment of viral infection, we proposed a hypothesis that whether LSECtin inhibitedCD8+T cells antiviral function. After HDI HBV plasmid, compared WT mouse intrahepaticCD8+T cells phenotype with that of KO mouse, we found that CD8+T cells had CD25highCD62LlowCD127lowphenotype, which exert antiviral properties, in LSECtin KO mice.ELISpot experiments further suggested that LSECtin can suppressed the viral antigen-specificT cells to secrete IFN-γ. These results suggested that the suppressed effctor T cells(CD8+T)may partly contribute to the delayed clearance of HBV from the blood and infectedhepatocytes in the HDI-HBV-treated model.Solid Phenotye and preliminary mechanisms directed us to try some applications aboutLSECtin. Small molecule compounds GlcNAcβ1-2Man was high affinity ligands of LSECtin.Ki was3.5+0.4uM. Recent studies have found that, Ebola virus and Lassa virus binding toLSECtin-expressing cells were inhibited by GlcNAcβ1-2Man efficiently. In addition,LSECtin-mediated Japanese encephalitis virus infection was reduced by treatment with GlcNAcβ1-2Man. Even so, whether the sugar has any blockade ability to LSECtin in vivo hasnot been reported internationally. The pAAV-HBV1.2plasmid was injected into C57BL/6mice using HDI. Three days later after injection, the mice were intraperitoneally injected13uM of GlcNAcβ1-2Man. Once a day for six weeks. surprisingly, sugar injection group hada lower surface antigen and E antigen levels than PBS group, also a lower surface antigenpositive rate. According to the results, it was suggested that GlcNAcβ1-2Man may be as apotential inhibitor to LSECtin in vovo.In summary, this study demonstrated that inhibitory receptor LSECtin is an importantpathway regulating CD8+T cell dysfunction during HBV infection in mice. Small moleculecompounds GlcNAcβ1-2Man is a potential target ligands for LSECtin. The blockade of theLSECtin may open new therapeutic approaches for chronic viral infection.