Single-cell Profiling of Intrahepatic Immune Cells Reveals an Expansion of Tissue-resident Cytotoxic CD4+ T Lymphocyte Subset Associated With Pathogenesis of Alcoholic-associated Liver DiseasesSummary

Background & Aims: The immunological mechanisms underpinning the pathogenesis of alcoholic-associated liver disease (ALD) remain incompletely elucidated. This study aims to explore the transcriptomic profiles of hepatic immune cells in ALD compared with healthy individuals and those with metabolic dysfunction-associated steatotic liver disease (MASLD). Methods: We utilized single-cell RNA sequencing to analyze liver samples from healthy subjects and patients with MASLD and ALD, focusing on the immune cell landscapes within the liver. Key alterations in immune cell subsets were further validated using liver biopsy samples from additional patient cohorts. Results: We observed a significant accumulation of CD4+ T cells in livers of patients with ALD, surpassing the prevalence of CD8+ T cells, in contrast to patients with MASLD and healthy counterparts, whereas natural killer (NK) cells and γδT cells exhibited reduced intrahepatic infiltration. In-depth transcriptional and developmental trajectory analyses unveiled that a distinct CD4+ subset characterized by granzyme K (GZMK) expression, displaying a tissue-resident signature and terminal effector state, prominently enriched among CD4+ T cells infiltrating the livers of patients with ALD. Subsequent examination of an independent ALD patient cohort corroborated the substantial enrichment of GZMK+CD4+ T lymphocytes, primarily within liver fibrotic zones, suggesting their potential involvement in disease progression. Additionally, we noted shifts in myeloid populations, with expanded APOE+ macrophage and FCGR3B+ monocyte subsets in ALD samples relative to MASLD and healthy tissues. Conclusions: In summary, this study unravels the intricate cellular diversity within hepatic immune cell populations, highlighting the pivotal immune pathogenic role of the GZMK+CD4+ T lymphocyte subset in ALD pathogenesis.