Emergence of highly profibrotic and proinflammatory Lrat + Fbln2 + HSC subpopulation in alcoholic hepatitis

Background and Aims: Relative roles of HSCs and portal fibroblasts in alcoholic hepatitis (AH) are unknown. We aimed to identify subpopulations of collagen type 1 alpha 1 (Col1a1)–expressing cells in a mouse AH model by single‐cell RNA sequencing (scRNA‐seq) and filtering the cells with the HSC (lecithin retinol acyltransferase [Lrat]) and portal fibroblast (Thy‐1 cell surface antigen [Thy1] and fibulin 2 [Fbln2]) markers and vitamin A (VitA) storage. Approach and Results: Col1a1–green fluorescent protein (GFP) mice underwent AH, CCl 4 , and bile duct ligation (BDL) procedures to have comparable F1‐F2 liver fibrosis. Col1a1‐expressing cells were sorted via FACS by VitA autofluorescence and GFP for single‐cell RNA sequencing. In AH, approximately 80% of Lrat+Thy1−Fbln2− activated HSCs were VitA‐depleted (vs. ~13% in BDL and CCl 4 ). Supervised clustering identified a subset co‐expressing Lrat and Fbln2 (Lrat+Fbln2+), which expanded 44‐fold, 17‐fold, and 1.3‐fold in AH, BDL, and CCl 4 . Lrat+Fbln2+ cells had 3–15‐times inductions of profibrotic, myofibroblastic, and immunoregulatory genes versus Lrat+Fbln2− cells, but 2–4‐times repressed HSC‐selective genes. AH activated HSCs had up‐regulated inflammatory (chemokine [C‐X‐C motif] ligand 2 [Cxcl2], chemokine [C‐C motif] ligand 2), antimicrobial (Il‐33, Zc3h12a), and antigen presentation (H2‐Q6, H2‐T23) genes versus BDL and CCl 4 . Computational deconvolution of AH versus normal human bulk‐liver RNA‐sequencing data supported an expansion of LRAT+FBLN2+ cells in AH; AH patient liver immunohistochemistry showed FBLN2 staining along fibrotic septa enriched with LRAT+ cells; and in situ hybridization confirmed co‐expression of FBLN2 with CXCL2 and/or human leukocyte antigen E in patient AH. Finally, HSC tracing in Lrat‐Cre;Rosa26mTmG mice detected GFP+FBLN2+ cells in AH. Conclusion: A highly profibrotic, inflammatory, and immunoregulatory Lrat+Fbln2+ subpopulation emerges from HSCs in AH and may contribute to the inflammatory and immunoreactive nature of AH.