Role Of Myeloid Progenitors And Tissue Resident Memory CD8~+T Cells In Autoimmune Liver Disease

First part:Immunosuppressive activityof myeloid progenitors via IFN-?-STAT1-iNOS in Con A induced AIH modelAutoimmune hepatitis(AIH)is a CD4+T cell mediated autoimmune liver disease.Mononuclear cells infiltration surrounding the portal vein,liver injury,as well as elevated level of serum aspartate aminotransferase(AST),alanine aminotransferase(ALT)and auto-antibodies can be detected in AIH patients.Standard treatment of AIH is immunesuppression therapy including glucocorticoids.Autoimmune diseases often induce dysregulated hematopoiesis,characterized by altered number and function of hematopoietic stem and progenitor cells(HSPCs).Compared with healthy controls,HSPC percentagein the bone marrow of AIH patients is higher,with increased number of colony forming units(CFU)in vitro.Besides,stromal cells show actively supportive function.HSPCs is the source of all kinds of blood cells and immune cells while they can also respond to pathogens directly.On the other hand,inflammatory cytokines such as IFN-? could regulate the number and function of HSPCs.However,there are limited studies on the direct regulation of HSPCs on T cells,which are often detrimental to autoimmunity.Here we found that in a murine model of Concanavalin A(Con A)induced AIH,LSK(Lineage-Sca-1+c-Kit+)-like cells accumulated in liver,spleen and bone marrow,which were myeloid progenitors(Lineage-Sca-1-c-Kit+)that upregulated Sca-1 expression upon T cell derived IFN-? stimulation.Strikingly,bone marrow LSK-like cells from mice with Con Ainduced autoimmune hepatitis or alternatively myeloid progenitors from wild type mice possessed strong in vitro suppressive ability.Their suppressive function depended on T cell derived IFN-? in a paracrine fashion,which induced STAT1 phosphorylation,inducible Nitric Oxide Synthase(NOS)expression and nitric oxide production.Blocking IFN-?/IFN-? receptor interaction,knockout of STAT1 or iNOS inhibition abrogated their suppressive function.In addition,the suppressive function was independent of differentiation as mitomycin C treated myeloid progenitors maintained T cell suppressive ability in vitro.In summary,our data demonstrate a mechanism of inflammation induced suppressive function of myeloid progenitors which may participate directly in suppressing T cell mediated immunopathology.Second part:Role of CD8+ tissue resident memory T cells in primary billilary cholangitisPrimary billilary cholangitis(PBC)is an autoimmune liver disease which maily affect middle-aged women.PBC initiates from break of immune tolerance to self mitochondrial antigens and immune attack of intrahepatic small bile duct by autoreactive cells.Bile duct damage results to liver cholestasis and inflammation,which probably further develops into liver fibrosis even cirrhosis.Due to limited knowledge of immunopathogenesis of PBC,there is hardly any effective therapy.The approved drugs for PBC,ursodeoxycholic acid and obeticholicacid,aim to modulate cholestasis induced liver toxicity but can not deal with the autoimmune attack.Previously we have found that IL-12p40-/-IL-2R?-/-(DKO)mice can be used as a murine PBC model,in which the liver inflammation,bile duct damage and fibrosis ressemble clinical PBC symptoms.In order to identify the main contributors of PBC disease in DKO mice,we crossed CD4-/-or CD8?-/-mice with DKO mice.It showed that after knock out of CD8a,the liver inflammation and bile duct damage almost disappeared while knock out of CD4 could ameliorate disease slightly.Besides,CD8+T cell depletion by antibody treatment could significantly alleviate liver lesions.These results demonstrated the pathogenic role of CD8+T cells in DKO mice.Furtherly,transcriptional analysis of CD8+ T cells from liver or spleen of WT or DKO mice was performed.Liver CD8+T cells showed a high expression level of tissue resident and T cell activation related genes than other CD8+T cell subsets.We comfirmed the existence of CD8+ tissue memory cells in liver of DKO mice,maily distributed in the portal inflammatory infiltration and sinosoidal by flow cytometry and immunohistochemistry.Importantly,the percentage of CD69+ Trm cells in CD8+T cells increased in the liver of PBC patients compared with healthy control.In order to investigate the role of Trmcells in the pathogenesis of PBC,the transcriptome of CD8+Tcm/em/rmcells was analysed.We found higher expression of tissue resident and activation related genes in Trm cells compared to other CD8+T cell subsets.CD8+ Trm cells also shows low level of proliferation and apoptosis.According to previous reports and our analysis,we hypothesized that Trm cells might be the earliest activated cells during the break of immune tolerance due to liver residency,and could produce a large amout of cytokines to recruit other circulating immune cells to liver tissues thus to promote inflammation and liver injury.In addition,we are screening some special molecule or transcriptional factors of Trm cells,which might be used as target in a potential Trm cells based therapy.In summary,our data demonstrate a higher CD8+Trm cell percentage in liver of PBC patients and mice model,and CD8+Trmcells in liver of DKO mice exhibit highly activated transcriptome status which might promote liver inflammation and tissue damage of PBC.