Single-cell multimodal analysis reveals the dynamic immunopathogenesis of HBV-ACLF progression

Background Acute-on-chronic liver failure (ACLF) is a life-threatening syndrome involving dysfunction of multiple immune cell types. Objective This study aimed to comprehensively depict the dynamic trajectory of immune responses throughout the disease course of HBV-related ACLF (HBV-ACLF). Design Single-cell RNA sequencing and single-cell proteomics were performed on the peripheral blood mononuclear cells of 45 samples from 17 patients who were hospitalised (progressive/stable/recovering course of HBV-ACLF, 6/5/6) and 15 control subjects (liver cirrhosis, chronic hepatitis B and healthy controls, 5/5/5). Functional and mechanistic experiments were validated in vivo and in vitro. Results Single-cell multiomics analysis revealed specific changes in the peripheral immune response in ACLF. VCAN + CD14 + -monocytes with activated interferon-stimulated genes and enhanced inflammatory functions, stimulated by HBV relapse and expanded in ACLF-1, fuelling early inflammatory storm. The subsequent apoptotic hepatocytes predominantly induce hyperinflammatory C-X-C motif chemokine receptor 2 (CXCR2) + -neutrophils and CD163 + -monocytes, enriching in patients with progressive ACLF and serving as significant markers of disease deterioration. Cytotoxic T-cells were functionally impaired and significantly decreased in progressive patients. CXCR2 + -neutrophils exhibited immunosuppressive activity and induced the exhaustion of cytotoxic T-cells. Pharmacological inhibition of CXCR2 significantly reduced neutrophils infiltration, restored cytotoxic T-cells and showed therapeutic effect in ACLF mice. Six immune cellular modules (CMs) were identified for patient stratification, with CM2 and CM6 showing strong predictive value for disease outcomes, and CM3 indicating a potential early therapeutic window. Conclusion Our longitudinal multiomics study revealed the dynamic evolution of the immune response in HBV-ACLF and characterised diverse immune patterns for the future precise management and therapeutic intervention.