MCM2 mediates post-MI cardioprotection by promoting the pro-angiogenic cardiosome signaling

AbstractO_ST_ABSBackgroundC_ST_ABSIn the past decade, induced cardiac rejuvenation has emerged as a leading approach to repair cardiac injury. Recent studies demonstrate that promoting cell cycle reentry in adult cardiomyocytes (CM) enhances cardiac rejuvenation by influencing paracrine signaling. We previously demonstrated that the inhibition of two cell cycle inhibitors, Retinoblastoma 1 (Rb1) and Meis homeobox 2 (Meis2), in the adult CM enhances angiogenesis and cardiac function following ischemic injury, but the underlying mechanisms have yet to be elucidated. The goal of this study is to determine the mechanisms by which inhibition of Rb1 and Meis2 promotes cardiac rejuvenation in a mouse model of myocardial infarction. MethodsMyocardial infarction was induced in adult C57/Bl6 mice via permanent LAD occlusion followed by direct injection of either control or Rb1+Meis2 siRNA cocktail to the ischemic myocardium. MCM2 overexpression done via direct myocardial injection of an MCM2-plasmid DNA expression cassette. Cardiac function and LV wall motion was assessed via echocardiography, and fibrosis and CM hypertrophy were assessed via histology. RNA-sequencing was performed on isolated adult murine CMs with siRNA-mediated Rb1+Meis2 knockdown to delineate the downstream mechanisms. Further identification of Rb1, Meis2, and MCM2-dependent mechanisms were done using in vitro techniques in isolated CMs and HUVEC cells. ResultsWe show that siRNA-mediated knockdown of Rb1 and Meis2 in vivo reduces pathological LV remodeling and preserves cardiac structure and function in adult mouse hearts post-MI. RNA-seq analyses revealed MCM2 as a potential downstream target of Rb1/Meis2 to enhance protective paracrine signaling in primary adult CMs. Indeed, re-expression of MCM2, which is developmentally lost from neonatal to adult CM in the heart, improves cardiac function and LV wall motion while reducing myocyte hypertrophy and fibrotic scar size post-MI. Mechanistically, re-expression of MCM2 promotes the secretion of pro- angiogenic factors from adult CM, and transfer of conditioned media from MCM2 expressing CM induced vasculogenesis in HUVEC cells. Proteomic analysis of the MCM2 interactome confirmed a significant enrichment of angiogenic mediators and suggests an MCM2-dependent protein packaging of pro- angiogenic factors in CM-derived small extracellular vesicles (cardiosomes). ConclusionRe-expression of MCM2 in adult CM promotes the secretion of pro-angiogenic cardiosomes that induce paracrine revascularization of endothelial cells and mitigates cardiac injury post-MI. SUMMARY DIAGRAM O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/628232v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@17eaeforg.highwire.dtl.DTLVardef@40f0f5org.highwire.dtl.DTLVardef@18b7664org.highwire.dtl.DTLVardef@109cb1c_HPS_FORMAT_FIGEXP M_FIG C_FIG