(102) TLR9 and Type II IFN signaling in monocyte-derived macrophages orchestrate CD8 T cell activity in DNaseII deficiency-induced autoimmune hepatitis

Introduction/Rationale: Aberrant accumulation of nucleic acids underlies a variety of sterile inflammation–induced liver diseases, as evidenced by patients with hypomorphic mutations in DNase2. DNaseII⁻/⁻ IFNαR⁻/⁻ double-knockout (DKO) mice recapitulate key features observed in these patients. However, it remains unclear which nucleic acid sensors are responsible for triggering this autoimmune hepatitis (AIH) phenotype and how they do so. In this study, we aim to examine how nucleic acid–induced signaling alters the heterogeneity of liver macrophages and shapes CD8⁺ T cell function in autoimmune hepatitis.

Methods: Deep profiling of liver immune cells was performed by spectral flow cytometry and immunofluorescence staining, scRNAseq and bulk RNAseq analysis. The roles of endosomal TLRs and IFNγ are explored using genetically ablation and use of bone marrow radiation chimeras.

Results: We observed extensive expansion of myeloid cells and CD8+ tissue resident memory T in the DKO liver that depends on TLR9. The infiltrating Ly6Chi inflammatory monocytes rapidly differentiate into monocyte-derived Kupffer cells and eventually replace the original embryonic Kupffer cells. These infiltrating monocyte-lineage cells exhibit a stronger inflammatory signature marked by elevated expression of genes related to antigen-presentation. Notably, TLR9 is hihgly expressed in monocyte-derived macrophages and is required for the differentiation process. Transcriptome analysis comparing monocyte-lineage cells sorted from DKO and TLR9 TKO livers reveal a robust TLR9-dependent IFN signature that is induced by IFNγ. The initial recruitment of TLR9 expressing monocytes, subsequent activation of lymphocytes, and other aspects of hepatic inflammation in DKO mice are completely dependent on IFNγ signaling

Conclusion: Taken together, these findings highlight a critical synergy between IFNg and TLR9 signaling in liver infiltrating monocyte-lineage cells that promotes activation of CD8+ T cells in the pathogenesis of AIH.