Abstract / Summary
Abstract Sepsis-induced cardiomyopathy (SCM) is a life-threatening complication with limited therapeutic options, characterized by a pronounced inflammatory cytokine storm. Although NLRP3 inflammasome–dependent macrophage pyroptosis has emerged as a critical contributor to cytokine release, its upstream regulatory mechanisms remain poorly understood. This study investigated whether calpain promotes macrophage pyroptosis via NLRP3 inflammasome activation, thereby exacerbating SCM. A cecal ligation and puncture (CLP) rat model was employed for in vivo studies, and THP-1-derived macrophages stimulated with LPS plus ATP (LA) were used in vitro. CLP rats developed severe cardiac dysfunction and elevated myocardial IL-1β and IL-18 levels, all of these pathological alterations were markedly reversed by calpain inhibitor MDL28170. Notably, CD68⁺GSDMD⁺calpain⁺ macrophages were detected in cardiac tissues of CLP rats. In THP-1 cells, LA triggered GSDMD-dependent pyroptosis, as evidenced by reduced viability, increased IL-1β/IL-18 secretion, and enhanced plasma membrane localization of GSDMD-NT; these effects were abrogated by GSDMD siRNA or MDL28170. Knockdown of CAPN2 (m-calpain subunit), but not CAPN1, significantly rescued LA-induced pyroptotic phenotypes. Mechanistically, CAPN2 silencing suppressed LA-induced ASC upregulation, cleaved caspase-1 production, and caspase-1 enzymatic activity, with affecting NLRP3 expression, indicating that CAPN2 acts upstream of canonical NLRP3 inflammasome activation to trigger caspase-1-mediated GSDMD cleavage. Functionally, conditioned medium from LA-stimulated macrophages markedly reduced H9c2 cell viability, an effect prevented by calpain inhibition in macrophages. Collectively, our findings provide the first evidence that calpain-2 (CAPN2) drives macrophage pyroptosis via the NLRP3 inflammasome/caspase-1/GSDMD axis to exacerbate septic cardiomyopathy, highlighting calpain as a promising therapeutic target for SCM.