Abstract 15756: Vascular Guidance of Cardiomyocyte Proliferation During Growth and Regeneration

Abstract

Introduction: Stimulating cardiomyocyte (CM) proliferation is a major strategy for achieving therapeutic heart regeneration. However, heart regeneration requires coordinated interactions of multiple cell types. Because a hallmark of advanced heart failure is vascular rarefaction, the requirement of cardiac endothelial cells (CECs) for cardiac growth and regeneration is of particular importance. Hypothesis: We hypothesized that CECs are required for CM proliferation during growth and regeneration. Methods and Results: We performed a large-scale histologic assessment of neonatal mouse hearts and found the rate of CEC proliferation to shadow CM proliferation over the first 10 days of life. Using a nearest neighbor analysis, we found the fraction of proliferating CECs to be significantly enriched around cycling CMs compared to non-cycling CMs, suggesting that CEC and CM expansion are coupled within a myovascular niche. Single cell sequencing of neonatal mouse hearts after cryoinjury revealed that a majority of these proliferating CECs also express Vegfr2 . To functionally link CEC and CM proliferation, we generated Cdh5-CreER T2 ; Vefgr2 flox/flox mice to genetically delete Vegfr2 from CECs. Compared to mice with intact Vegfr2 , loss of Vegfr2 from CECs in neonatal mice leads to loss of CECs and severely dampens CM proliferation by 4 days (7.01±0.88% vs 0.39±0.35%, p = 7.4x10-4, n = 9),. Interestingly, CM proliferation is attenuated when Vegfr2 is deleted from CECs despite an increase of hypoxia indicators in CMs, signifying that hypoxia-induced CM proliferation is dependent on CECs. In contrast to CEC depletion, treatment of cryoinjured neonatal hearts with AAV encoding the master angiogenic factor, Vegfa can enhance heart regeneration with increased CM cycling in the borderzone (12.6±2.2% vs 5.4±0.4%, p = 0.02, n = 8), reduced scarring of the left ventricle (3.4±1.4% vs 7.6±1.2%, p = 03, n = 16), and improved fractional shortening (51.7±2.5% vs 36.7±4.3%, p = 0.007, n = 14). Conclusions: CEC and CM expansion are spatiotemporally coupled in a myovascular niche during cardiac growth. CECs play a critical role to support CM proliferation and are likely to provide instructive cues that may be leveraged for therapeutic heart regeneration.