Abstract P2063: Circular RNA, circREGEN, Alters Cardiac Regeneration Through Cardiomyocyte Proliferation and Regulation of Oxidative Stress

Introduction: A subtype of non-coding RNA, circular RNAs (circRNAs), formed through backsplicing, are emerging as important regulators of cardiac development and cardiovascular diseases. CircRNAs are known to regulate gene expression through a multitude of functions. Prevalent circRNAs involved in cardiac development and potential regeneration can be determined by focusing on their differential expression in regeneration competent versus non-competent neonatal mouse hearts. We hypothesized that a circRNA that was both highly expressed in day 0.5 tissue and was upregulated under hypoxic conditions (simulating the uterine environment) would have a high potential for regeneration. Methods and Results: RNA-seq was performed in day 1 and day 7 mouse hearts and circREGEN was found to be downregulated on day 7 both in RNA-seq analysis and in neonatal mouse tissue samples using RT-PCR. Further circRNA validation was also performed and tissue distribution was measured, showing high expression levels in the heart, specifically, neonatal cardiomyocytes. HL-1 cells, neonatal mouse and rat CMs, neonatal living myocardial tissue, and human induced pluripotent stem cell (iPSC)-derived CMs were subjected to hypoxia (simulating this low oxygen uterine environment) and observed an increase in expression. This data showed that in addition to having high sequence conservation, circREGEN was functionally conserved across species as well. We decided to further examine the expression of circREGEN in living myocardial tissue, an ex-vivo model, after cryoinjury and saw an up-regulation. Further experiments were performed to functionally validate circREGEN using siRNAs and overexpression plasmids in neonatal mouse and rat CMs that showed a decrease in proliferation after circREGEN knockdown, and a reduction in ROS after overexpression, showing potential protective and regenerative effects of circREGEN. Conclusion: In conclusion, we have promising in vitro data, which warrants further functional investigation both in vitro, ex-vivo, and in vivo , with the ultimate goal to determine if circREGEN has therapeutic potential to regenerate damaged heart tissue after a myocardial infarct by stimulating the proliferation of CMs in the adult heart.