Myocardial Delivery Of Modified Mrna For Exosomal Protein Induces Cardiac Regeneration

Permanent loss of cardiomyocytes (CM) after myocardial infarction (MI) and limited cardiac regenerative capacity leads to heart failure. Recent studies from zebrafish, newt, and even 1-day old neonatal mice showed that they could regenerate their heart through inducing existing CM proliferation. Attempts have been made to transiently reconstitute embryonic signaling in adult hearts, including overexpression of cell cycle activating genes with limited success. iPSC-derived extracellular vesicles (EV)/exosomes have been shown to improve cardiac function and some degree of CM renewal. However, the iPSC-EVs-mediated cardiac regeneration mechanism remains unclear and largely pertains to microRNAs and other RNAs, but not the proteins. Our hypothesis is the myocardial delivery of iPSC-EV-specific protein improves cardiac function and remodeling post-MI by activating pro-proliferative and anti-oxidative stress molecular pathways. Our preliminary studies showed that hiPSC-EVs induced the CM cell cycle in mice post-MI, and by employing a proteomic approach, we found a novel protein exclusively expressed in iPSC-EVs. The overexpression of hiPSC-EV enriched protein in the form of modRNA (modified mRNA) induces a robust CM cell cycle in rat neonatal CMs and in heart post-MI. By employing cardiomyocyte specific MADM (Mosaic Analysis of Double Markers) mice we showed that EV-protein induced robust cardiomyocyte division post-MI. This increase in the CMs proliferation by the modRNA was associated with reduced scar size, improved cardiac function, and mice survival 28 days post-MI. Moreover, we show that the YAP, a master regulator of CM proliferation and cardiac regeneration, binds to the promoter of the protein and induces its expression. Furthermore, using the siRNA and modRNA (inhibition and over-expression respectively) approach, we found that the protein-induced Yap1-β-catenin molecular pathway stimulates CM proliferation. Furthermore, the overexpression protein inhibited the oxidative stress or ROS analyzed by HPLC and DNA damage response post-MI. As a result of it, we have seen a significant reduction in CM apoptosis post-MI. Taken together we see the myocardial injection of iPSC-EV specific protein through a highly therapeutic modRNA tool improve cardiac function by inducing CMs proliferation, inhibiting oxidative stress, and reactivating cardiac regeneration post-injury.