Abstract Rationale Hepatopulmonary syndrome (HPS) is a severe vascular complication of chronic liver disease characterized by hypoxemia and pulmonary vasodilation. The common bile duct ligation (CBDL) mouse model recapitulates key pathological features of HPS, providing an experimental platform to investigate hepatic cellular mechanisms driving disease onset and progression. We applied single-cell RNA sequencing (scRNA-seq) to comprehensively define hepatic immune, stromal, and parenchymal responses underlying HPS pathogenesis. Methods Male mice were underwent CBDL or sham surgery at week 12. Three weeks after surgery, oxygen saturation and intrapulmonary shunting assays confirmed successful HPS model establishment. Liver tissues were processed for scRNA-seq using the 10x Genomics platform. Data was analyzed with Cell Ranger and Seurat for integration, clustering, and differential expression analyses, followed by pathway and ligand receptor network interrogation. Results Three weeks after CBDL, mice exhibited clinical and physiological manifestations of HPS, including jaundice, 20-30 % weight loss, and significantly increased liver-to-body and spleen-to-body weight ratios. Arterial oxygen saturation was markedly reduced, whereas intrapulmonary shunting was elevated. scRNA-seq of 50 000 liver cells revealed pronounced inflammation and endothelial activation with a prominent ductular reaction. Kupffer cells and SACR-related (SPP1-associated repair) macrophages (Spp1+, Apoe+, Lyz2+) expanded markedly, reflecting an activated innate-immune landscape. The inflammatory milieu was associated with cholangiocyte proliferation and activation of hepatic stellate cells (HSCs), accompanied by fibroblast expansion and matrix remodeling. Pericentral hepatocytes were depleted, indicating localized parenchymal loss and disrupted hepatic zonation. Hallmark GSEA demonstrated upregulation of inflammatory response, epithelial mesenchymal transition (EMT), and fibrosis pathways in hepatocytes and cholangiocytes. Kupffer cells exhibited unfolded protein response and oxidative stress signatures, whereas hepatic stellate cells showed enhanced hypoxia and EMT signaling, consistent with fibrogenic activation. Collectively, these findings define a coordinated hepatic response linking immune activation, ductular expansion, and progressive fibrosis during HPS development. Conclusions Single-cell analysis of the CBDL mouse liver delineates distinct cellular and molecular programs underlying HPS, highlighting Kupffer cell-driven inflammation and HSCs activation as central drivers and potential therapeutic targets. This abstract is funded by: NIH
Zheng et al. (Fri,) studied this question.