Cardiomyocyte‐Specific Overexpression of Activated Yes‐Associated Protein Modified‐RNA Promotes Cardiomyocyte Proliferation and Myocardial Regeneration

Abstract

Background The proliferative capacity of cardiomyocytes in adult mammalian hearts is far too low to replace the cells that are lost to myocardial infarction. Both cardiomyocyte proliferation and myocardial regeneration can be improved via the overexpression of a constitutively active variant of YAP5SA (Yes‐associated protein, 5SA [active] mutant), but persistent overexpression of proliferation‐inducing genes could lead to hypertrophy and arrhythmia, whereas off‐target expression in fibroblasts and macrophages could increase fibrosis and inflammation. Methods and Results Transient overexpression of YAP5SA or GFP (green fluorescent protein; control) was targeted to cardiomyocytes via our cardiomyocyte‐specific modified mRNA translation system ( YAP5SA CM‐SMRTs or GFP CM‐SMRTs, respectively). YAP5SA‐cardiomyocyte specificity was confirmed via in vitro experiments in cardiomyocytes and cardiac fibroblasts that had been differentiated from human induced‐ pluripotent stem cells and in human umbilical‐vein endothelial cells, and the regenerative potency of YAP5SA CM‐SMRTs was evaluated in a mouse myocardial infarction model. In cultured human induced‐pluripotent stem cells‐cardiomyocytes, YAP was abundantly expressed for 3 days after YAP5SA CM‐SMRTs administration and was accompanied by increases in the expression of markers for proliferation, before declining to near‐background levels after day 7, whereas GFP fluorescence remained high from days 1 to 3 after GFP CM‐SMRTs treatment and then slowly declined. GFP fluorescence was also observed in human induced‐pluripotent stem cells‐cardiac fibroblasts and human umbilical‐vein endothelial cells on day 1 after GFP CM‐SMRTs administration but declined substantially by day 3. In the mouse myocardial infarction model, echocardiographic assessments of left‐ventricular ejection fraction and fractional shortening were significantly greater, whereas infarct sizes were significantly smaller in YAP5SA CM‐SMRTs–treated mice than in vehicle‐treated control animals, and YAP5SA CM‐SMRTs appeared to promote cardiomyocyte proliferation. Conclusions The CM‐SMRTs can be used to transiently and specifically overexpress YAP5SA in cardiomyocytes, and this treatment strategy significantly promoted cardiomyocyte proliferation and myocardial regeneration in a mouse myocardial infarction model.