STING mediates hepatocyte pyroptosis in liver fibrosis via epigenetic activating NLRP3 inflammasome. STING mediates hepatocyte pyroptosis in liver fibrosis via epigenetic activating NLRP3 inflammasome

Background and Aims: The activation of stimulator of interferon genes (STING) and NOD-like receptors protein 3 (NLRP3) inflammasomes-mediated pyroptosis signaling pathways represent two distinct central mechanisms in liver disease. However, the interconnection between these two pathways and the epigenetic regulation of the STING-NLRP3 axis in hepatocyte pyroptosis during liver fibrosis remain unknown and is the focus of this study. Approach and Results: Liver fibrosis was induced in Sting knockout, Gasdermin D (Gsdmd) knockout mice, and in mice with hepatocyte-specific Nlrp3 deletion. RNA-sequencing, metabolomics, epigenetic compound screening system, and chromatin immunoprecipitation were utilized. STING and NLRP3 inflammasome signaling pathways were activated in cirrhotic livers but were suppressed by Sting knockout. Sting knockout also ameliorated hepatic pyroptosis, inflammation, and fibrosis in the murine cirrhotic model. In vitro, STING induced pyroptosis in primary murine hepatocytes via activating the NLRP3 inflammasome. H3K4-specific histone methyltransferase WD repeat-containing protein 5 (WDR5) and DOT1-like histone H3K79 methyltransferase (DOT1L) were identified to regulate NLRP3 expression in STING-overexpressed AML12 hepatocytes. WDR5/DOT1L-mediated histone methylation enhanced interferon regulatory transcription factor 3 (IRF3) binding to the Nlrp3 promoter and promoted STING-induced Nlrp3 transcription in hepatocytes. The RNA-sequencing and metabolomics analysis in murine livers and primary hepatocytes showed that metabolic reprogramming might participate in NLRP3-mediated hepatocyte pyroptosis and liver fibrosis. Moreover, hepatocyte-specific Nlrp3 deletion and downstream Gsdmd knockout attenuated hepatic pyroptosis, inflammation, and fibrosis in murine cirrhotic models. Conclusions: This study describes a novel epigenetic mechanism by which the STING-WDR5/DOT1L/IRF3-NLRP3 signaling pathway enhances hepatocyte pyroptosis and hepatic inflammation in liver fibrosis. Overall design: 1.The primary murine hepatocytes were stimulated with LPS (1 µg/mL) for 6 hours and nigericin (10 µM) for 30 minutes. The NLRP3 inhibitor MCC950 (10 µM) were added 2 hours before stimulation 2. One liver fibrosis model was induced by intraperitoneal injection of thioacetamide (TAA) in mice with hepatocyte-specific Nlrp3 deletion and wild type for 8 weeks. Control mice received the same volume of normal saline (NS). Another model was induced by intraperitoneal injection of carbon tetrachloride (CCl4) in mice with Sting knock out and wild for 8weeks. Control mice received the same volume of olive oil.