The Ets2 super-enhancer modulates endothelial-mesenchymal transition during cardiac aging

Abstract Aims Cardiac aging is characterized by endothelial dysfunction and associated cardiovascular pathologies, often involving endothelial-to-mesenchymal transition (EndoMT) in cardiac endothelial cells. While the transcription factor Ets2, a member of the Ets family, is known to regulate endothelial cell survival and function, its role in EndoMT and cardiac aging remains poorly understood. Methods and Results To investigate this, we utilized single-nucleus RNA sequencing (snRNA-Seq) in Ets2-super-enhancer (Ets2-SE)-deficient mice to examine the regulation of Ets2 expression across various cardiac cell types. We assessed the relationship between Ets2 expression and heart aging, and evaluated the characteristics of cardiac aging in Ets2-SE-deficient mice. Furthermore, we generated endothelial cell-specific Ets2 knockout mice (ECKO) to investigate the role of Ets2 in EndoMT of cardiac endothelial cells both in vitro and in vivo. Our results establish a link between decreased Ets2 expression and the development of aging-associated cardiac pathological remodeling. Through data analyses, we identified a super-enhancer (Ets2-SE) that regulates Ets2 expression in the heart. Ets2-SE-deficient mice exhibited significantly lower Ets2 expression in cardiac tissues and displayed advanced aging phenotypes, including increased cardiac fibrosis and dysfunction, compared to wild-type controls. SnRNA-Seq analyses revealed a remarkable downregulation of Ets2 in endothelial cells, correlating with the activation of EndoMT. Furthermore, endothelial-specific deletion of Ets2 exacerbated aging and myocardial infarction-induced cardiac fibrosis and heart dysfunction. Mechanistic studies demonstrated that silencing ETS2 in human umbilical vein endothelial cells (HUVECs) promotes EndoMT by transcriptionally suppressing the endothelial marker gene TIE1. This transition is accompanied by endothelial cell senescence and the activation of the senescence-associated secretory phenotype (SASP), which contributes to myocardial fibrosis and cardiac aging, partially mediated by Serpine1. These findings identify Ets2 as a critical regulator of EndoMT in the context of cardiac aging. Conclusions Our findings reveal that the Ets2 super-enhancer regulates Ets2 expression in cardiac endothelial cells, modulating heart aging and EndoMT. Ets2's regulation of endothelial marker genes, especially TIE1, plays a pivotal role in mitigating EndoMT and preventing senescence in cardiac vascular endothelial cells, suggesting potential therapeutic targets for addressing cardiovascular aging.