Investigation Of HESC-derived Cardiomyocyte Therapy In Different Myocardial Injury Models

Ischemic cardiomyopathy is a kind of heart disease caused by ischemia and hypoxia.The main manifestations of ischemic cardiomyopathy are cardiomyocytes death,fibrosis and impaired cardiac function,which finally lead to heart failure.As the death toll increases year by year,ischemic cardiomyopathy has become one of the main killers of human health.It is urgent to prevent and treat ischemic cardiomyopathy effectively.Pluripotent stem cells have broad application prospects in the treatment of ischemic cardiomyopathy because of their ability of self-renewal and multidirectional differentiation.However,the indication,efficacy and mechanism of stem cell therapy for ischemic cardiomyopathy still need to be further studied.The aim of this study was to investigate the efficacy of human embryonic stem cell-derived cardiomyocytes(ESC-Rep-CMs)in the treatment of myocardial injuries caused by permanent ischemia(Permanent ischemia,PI)and ischemia-reperfusion(Ischemic reperfusion,IR)in mice.In this study,a human embryonic stem cell line(ESC-Rep),which can be traced in vivo,was constructed by gene editing technique.The cells express three reporter genes,copGFP,HSVtk and Fluc,driven by CAG promoter.In vitro studies have shown that ESC-Rep cells maintained the potential for self-renewal and multidirectional differentiation and can efficiently differentiate into beating cardiomyocytes(ESC-Rep-CMs)expressing all three reporter genes.A positive linear correlation was observed in vitro between the signal intensity of luciferase activity and cardiomyocyte number,indicating that ESC-Rep-CMs could be well traced by in vivo bioluminescence imaging(BLI)after cell transplantation.After cell transplantation,we firstly confirmed the retention of ESC-Rep-CMs in recipient hearts of both PI and IR mouse by the methods of BLI and immunofluorescence.There is no significant difference in the survival rate of transplanted cells between PI and IR models.The results of echocardiography showed that cardiac contractile function indexes,including ejection fraction(Ejection fraction,EF)and shortening fraction(Fractional shortening,FS),were significantly increased in the PI group post ESC-Rep-CM transplantation.Cardiac diastolic function indicators,including the ratio of the peak velocity of early(E)and late filling waves(A),as well as the ratio of early diastolic lateral mitral annulus velocity(E’)and late diastolic lateral mitral annulus velocity(A’),were also significantly up-regulated in ESC-Rep-CMs-injected PI mice.However,the effect of cardiomyocyte transplantation on cardiac function of IR mice was not obvious.The results of Masson staining showed that ESC-Rep-CM transplantation could significantly decrease the infarct size in PI group,but had no significant effect on that in IR group.In order to explore the reasons for different therapeutic effects between the two models,we measured the expression of inflammatory factors in serum and the activity of myeloperoxidase(MPO)in heart tissue of mice.We found that cardiomyocyte transplantation could significantly inhibit the expression of proinflammatory factors(TNF-α,IL-6)and promote the expression of anti-inflammatory factor(IL-10)in PI mice.However,these effects of cardiomyocyte transplantation were not observed in IR mice.We further confirmed that the pro-inflammatory factor TNF-α significantly promoted the fibrosis and the anti-inflammatory factor IL-10 had an obvious inhibitory effect on the degree of fibrosis by using the model of myocardial fibrosis in vitro.Therefore,the results of this study indicated that hESC-CMs could improve the systolic and diastolic function of the heart in PI mice,but had no obvious therapeutic effect on the damaged heart of IR mice.Meanwhile,hESC-CMs may slow down the process of fibrosis by reducing the degree of inflammation after cardiac ischemia,thus improving cardiac function.