<i>Zeb2os</i> Hinders Cardiac Healing by Suppressing ZEB2 Reactivation and Cardiomyocyte Dedifferentiation.

Abstract

Long noncoding RNAs (lncRNA) have emerged as critical regulators in cardiovascular biology, influencing cardiac development, remodeling, and regeneration. <i>Zeb2os</i> (zeb2 opposite strand), a natural antisense transcript of the <i>Zeb2</i> (zinc finger E-box-binding homeobox 2) gene, has been linked to these processes in various organs. Although ZEB2 promotes cardiac repair, the role <i>of Zeb2os</i> in these processes remains unclear. This study investigates the role of <i>Zeb2os</i> in modulating ZEB2 expression and cardiac remodeling after ischemic injury. We used adeno-associated virus vectors to overexpress <i>Zeb2os</i> in mouse models of cardiac IR (ischemia/reperfusion) injury. RNA sequencing, immunofluorescence, and high-resolution respirometry were used to evaluate the effects of <i>Zeb2os</i> delivery on gene expression, ZEB2 reactivation, cardiomyocyte phenotype, scar composition, and mitochondrial function. Experiments in cultured cardiomyocytes under hypoxia further explored the regulatory dynamics between <i>Zeb2os</i> and <i>Zeb2</i>. We identified <i>Zeb2os</i> as a hypoxia-responsive lncRNA that displays an inverse and oscillatory expression pattern with <i>Zeb2</i> in both in vitro and in vivo models of cardiac injury. Functional experiments revealed that <i>Zeb2os</i> negatively regulates ZEB2 expression, impairing the cardiomyocyte dedifferentiation and metabolic remodeling necessary for effective repair. Adeno-associated virus-mediated delivery of <i>Zeb2os</i> resulted in preserved sarcomere structure, altered scar composition, reduced expression of regenerative genes, and diminished cardiac function following injury. In contrast, silencing of <i>Zeb2os</i> increased ZEB2 protein expression, suggesting a potential therapeutic strategy to enhance repair. Mechanistically, modulation of <i>Zeb2os</i> levels inversely regulated ZEB2 protein expression, whereas ZEB2 modulation did not affect <i>Zeb2os</i> levels, indicating a unidirectional regulatory axis between the 2 transcripts. Our findings identify <i>Zeb2os</i> as a stress-responsive inhibitor of ZEB2 reactivation that limits cardiomyocyte plasticity and hinders repair following ischemic injury. Given its specific activity under ischemic conditions, targeting <i>Zeb2os</i> may represent a novel therapeutic strategy to enhance endogenous cardiac regeneration.