Protection Of Glycyl-L-glutamine Against Myocardial Ischemia-reperfusion Injury

OBJIECTIVE:To study the effect of Glycyl-L-glutamine(Gly-Gln) on the heart function,mitochondrial membrane potentials,apoptosis,the release of myocardial enzymes of cardiomyocytes suffering from ischemia/reperfusion injury.And to investigate the protective mechanism of Gly-Gln in ischemia/reperfusion injury.METHODS:1.To investigate the effect of Gly-Gln on the cultured cardiomyocytes injured by H/R:A cardiac H/R model in vitro was established with primary cultured neonate rat cardiomyocytes. There were four experimental groups:control group;H/R group;H/R+Gly-Gln group;Gly-Gln group.The viability of cardiomyocytes was observed by MTT assay.The contents of lactate dehydrogenase(LDH) and creatine kinase(CK) in the culture solution of various groups were determined.The apoptosis rate of cardiomyocytes was analysed by AnnexinV /PI double staining.The mitochondrial membrane potential(△ψm) of cardiomyocytes was measured by JC-1 as a fluorescent molecular probe.2.To investigate the effect of Gly-Gln on isolated I/R heart:A model of myocardial ischemia-reperfusion injury was established with a Langendorff apparatus to observe the effect of Gly-Gln on the heart function,myocardial enzymes and monophasic action potentials in cardiomyocytes.30 male SD rats were randomly divided into four groups:control group;I/R group;I/R+Gly-Gln group;Gly-Gln group.We measured the left ventricular end-diastolic pressure(LVEDP),left ventricular developed pressure(LVDP),±dp/dt_max,heart rates(HR), monophasic action potentials(MAP) during perfusion and determined the level of lactic dehydrogenase(LDH) and creatine kinase(CK) in the coronary effluent fluid at different times.3.To investigate the effect of Gly-Gln on cardiac I/R model in vivo:An in vivo cardiac I/R model was established in mice to observe the effect of Gly-Gln on the myocardial function.40 mice were randomly divided into four groups:control group;I/R group;I/R+Gly-Gln group; Gly-Gln group.The heart rate(HR),left ventricular short axis fractional shortening(FS),ejection fraction(EF),cardiac output(CO),stroke volume(SV) and ratio E/A were detected with Vevo770 High-Resolution.RESULTS:1.Gly-Gln increased viability of cardiomyocytes suffering H/R injury remarkably(P＜0.01), inhibited the release of LDH and CK which induced by H/R injury(P＜0.05),Gly-Gln reduced the apoptosis rate of cardiomyocytes significantly(P＜0.01).The effect of Gly-Gln was in a dose-dependent manner within certain concentration range,The△ψm of cardiomyocyte was obviously lower in H/R group than that in control group(P＜0.01).But,the△ψm was higher in H/R + Gly-Gln group than that in H/R group(P＜0.01).2.The isolated rat heart function decreased severely after 20 min ischemia and 40 min reperfusion(I/R):the LVEDP increased and the LVDP and±dp/dtmax decreased.But the LVEDP decreased and the LVDP,±dp/dtmax increased in I/R+Gly-Gln group than I/R group. Moreover,the activities of LDH and CK in the coronary effluent fluid decreased significantly in I/R+Gly-Gln group compared with I/R group.3.Gly-Gln reduced the decrease of SV,CO,LV Vol d/s,ratio E/A and the increase of LVEF, LVFS which induced by I/R injury in vivo rat.CONCLUSION:1.Gly-Gln promotes the survival of cardiomyocytes subjecting to H/R injury,reducing the release of myocardial enzymes induced by H/R injury,and prevents the apoptosis and decrease in mitochondrial membrane potential of cardiomyocytes suffering H/R injury.2.Gly-Gln exerts protective effects against myocardial I/R injury in isolated rat hearts by effectively relieving the decrease in left ventricular function and decreasing the activities of LDH and CK in the coronary effluent fluid.3.Gly-Gln improves the cardiac function of mice against myocardial I/R injury in vivo.