Mechanistic role of RND3-regulated IL33/ST2 signaling on cardiomyocyte senescence

Abstract

Abstract Objective: Hyperinflammatory responses are pivotal in the pathophysiology of cardiomyocyte senescence, with IL33 serving as a crucial pro-inflammatory mediator. Our previous findings highlighted RND3's suppressive effect on IL33 expression. This study delves into the influence of RND3 on IL33/ST2 signaling activation and cardiomyocyte senescence. METHODS: AC16 cardiomyocytes were subjected to treatments involving recombinant IL33, NF-κB inhibitor PDTC, or ST2 antibody Astegolimab. SA-β-gal and γH2AX staining were utilized to monitor alterations in cell senescence and DNA damage, respectively. Western blot analysis was conducted to ascertain the expression of Senescence-Associated Secretory Phenotype (SASP) and NF-κB activation. Utilizing CRISPR/Cas9 technology, the RND3 gene was knocked out in H9C2 cells, followed by senescence analysis and sST2 level detection in the culture medium supernatant via ELISA. Post-AAV9 injection overexpressing RND3in SD rats, IL33/ST2 and SASP expression in heart tissues, and serum IL33 and sST2 changes were evaluated using ELISA. RESULTS: Exogenous IL-33 significantly induced IL-1α, IL6, and MCP1 expression, increased the p-p65/p65 ratio, and the proportion of SA-β-gal and γH2AX positive cells in AC16 cells. PDTC and Astegolimab application mitigated these effects. RND3 knockout in H9C2 cells led to increased intracellular IL33, ST2L, IL1 α, IL6, and MCP1 expression, decreased sST2 in the supernatant, and increased SA-β-gal and γH2AX positive cells. RND3 overexpression suppressed IL33, ST2L, IL-1α, IL6, and MCP1 expression in heart tissues, decreased serum IL33, and increased sST2 levels. CONCLUSION: RND3 expression in cardiomyocytes modulates cell senescence by negatively regulating the IL33/ST2/NF-κB signaling pathway, underscoring its potential as a therapeutic target in cardiovascular senescence.