ABSTRACT
BACKGROUND AND AIMS: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by biliary inflammation and fibrosis. We showed an elevated interferon γ response in patients with primary sclerosing cholangitis and in multidrug resistance protein 2-deficient ( Mdr2-/- ) mice developing sclerosing cholangitis. Interferon γ induced expression of the cytotoxic molecules granzyme B (GzmB) and TRAIL in hepatic lymphocytes and mediated liver fibrosis in sclerosing cholangitis. APPROACH AND RESULTS: In patient samples and Mdr2-/- mice, we identified lymphocyte clusters with a cytotoxic gene expression profile using single-cell RNA-seq and cellular indexing of transcriptomes and epitopes by sequencing analyses combined with multi-parameter flow cytometry. CD8 + T cells and NK cells showed increased expression of GzmB and TRAIL in sclerosing cholangitis. Depletion of CD8 + T cells ameliorated disease severity in Mdr2-/- mice. By using Mdr2-/- × Gzmb-/- and Mdr2-/- × Tnfsf10-/- mice, we investigated the significance of GzmB and TRAIL for disease progression in sclerosing cholangitis. Interestingly, the lack of GzmB resulted in reduced cholangiocyte apoptosis, liver injury, and fibrosis. In contrast, sclerosing cholangitis was aggravated in the absence of TRAIL. This correlated with elevated GzmB and interferon γ expression by CD8 + T cells and NK cells enhanced T-cell survival, and increased apoptosis and expansion of cholangiocytes. CONCLUSIONS: GzmB induces apoptosis and fibrosis in sclerosing cholangitis, whereas TRAIL regulates inflammatory and cytotoxic immune responses, subsequently leading to reduced liver injury and fibrosis.
ABSTRACT
BACKGROUND AND AIMS: Primary sclerosing cholangitis (PSC) is an idiopathic, chronic cholestatic liver disorder characterized by biliary inflammation and fibrosis. Increased numbers of intrahepatic interferon-γ- (IFNγ) producing lymphocytes have been documented in patients with PSC, yet their functional role remains to be determined. METHODS: Liver tissue samples were collected from patients with PSC. The contribution of lymphocytes to liver pathology was assessed in Mdr2-/- x Rag1-/- mice, which lack T and B cells, and following depletion of CD90.2+ or natural killer (NK)p46+ cells in Mdr2-/- mice. Liver pathology was also determined in Mdr2-/- x Ifng-/- mice and following anti-IFNγ antibody treatment of Mdr2-/- mice. Immune cell composition was analysed by multi-colour flow cytometry. Liver injury and fibrosis were determined by standard assays. RESULTS: Patients with PSC showed increased IFNγ serum levels and elevated numbers of hepatic CD56bright NK cells. In Mdr2-/- mice, hepatic CD8+ T cells and NK cells were the primary source of IFNγ. Depletion of CD90.2+ cells reduced hepatic Ifng expression, NK cell cytotoxicity and liver injury similar to Mdr2-/- x Rag1-/- mice. Depletion of NK cells resulted in reduced CD8+ T cell cytotoxicity and liver fibrosis. The complete absence of IFNγ in Mdr2-/-x Ifng-/- mice reduced NK cell and CD8+ T cell frequencies expressing the cytotoxic effector molecules granzyme B and TRAIL and prevented liver fibrosis. The antifibrotic effect of IFNγ was also observed upon antibody-dependent neutralisation in Mdr2-/- mice. CONCLUSION: IFNγ changed the phenotype of hepatic CD8+ T cells and NK cells towards increased cytotoxicity and its absence attenuated liver fibrosis in chronic sclerosing cholangitis. Therefore, unravelling the immunopathogenesis of PSC with a particular focus on IFNγ might help to develop novel treatment options. LAY SUMMARY: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by biliary inflammation and fibrosis, whose current medical treatment is hardly effective. We observed an increased interferon (IFN)-γ response in patients with PSC and in a mouse model of sclerosing cholangitis. IFNγ changed the phenotype of hepatic CD8+ T lymphocytes and NK cells towards increased cytotoxicity, and its absence decreased liver cell death, reduced frequencies of inflammatory macrophages in the liver and attenuated liver fibrosis. Therefore, IFNγ-dependent immune responses may disclose checkpoints for future therapeutic intervention strategies in sclerosing cholangitis.
