Effect Of Visfatin On Rat Heart And Its Mechanism

Visfatin is a novel visceral fat factor. It has aroused much attention since it wasfirst reported in2005. Visfatin play an important physiological and pathologicalroles in diabetes, obesity and other metabolic and endocrine diseases,inflammatory diseases, breast cancer, ischemic stroke and ischemic heart disease.It is reported that visfatin play a role of myocardial protection by reducing themyocardial infarct size induced by myocardial ischemia and reperfusion injury（IRI） in mice. Study of the role and mechanism of ectogenous visfatin in the heartis therefore of great significance. However, the direct effect on the isolatedperfused rat heart of ectogenous visfatin, the roel of visfatin inischemia-reperfusion myocardial injury, and its impact on [Ca²⁺]iand the ionchannel currents of single rat ventricular myocytes have not been reported.ObjectiveThe purpose of this work is to investigate the direct roles of ectogenousvisfatin on cardiac function in an isolated perfused rat heart model, and the effectof ectogenous visfatin on myocardial IRI in a rat heart model. Inaddition, theeffects of ectogenous visfatin on [Ca²⁺]i, sodium-calcium exchange current （INa/Ca）,L-type calcium channel current （ICa, L） and ATP sensitive potassium channelcurrent （IK, ATP） were investigated in a rat model of acutely isolated singleventricular myocytes, in an attempt to identify the direct cardiac function ofectogenous visfatin on the isolated rat heart and its effect on myocardial IRI, thusgaining insights into the role of visfatin on the rat heart and the underlying mechanisms.MethodsIsolated SD rat hearts were perfused with KH solution by the Langendorffsystem at a constant temperature （37°C） and a constant voltage （80cm H2O）no-circulating perfusion, and a myocardial IRI rat model was established by globalischemial when the cardiac function of the isolated rat heart was stable. Leftventricular systolic pressure （LVSP）, left ventricular diastolic pressure （LVEDP）,left ventricular developed pressure （LVDP） left ventricular pressure increase anddecrease rate （±dp/dt）, coronary flow （CF） and heart rate （HR） were observedunder different concentrations of ectogenous visfatin of the isolated rat heart andthe ischemia-reperfusion injury rat heart model and the IRI rat model. Single ratventricular myocytes were obtained from the SD rat heart by a enzymolysisprocedure; using laser scaning confocal microscopy [Ca²⁺]iwere observed in thepresent of visfatin; using whole cell patch clamp technique, INa/Ca, ICa, L, and IK, ATPof the single rat ventricular myocytes were observed in the presence of differentconcentrations of ectogenous visfatin.Results（1）. Visfatin concentration-dependently inhibited LVSP, LVDP,±dp/dt, HRand CF of the isolated perfused rat hearts. After perfusion for15min with50nmol/l （n=8）,100nmol/l （n=7） and200nmol/l visfatin （n=8）, LVSP decreased by25.7±4.0%,29.8±2.6%and39.0±3.6%, which were significantly higher than9.1±1.4%in the control group （P<0.01vs. control, n=8）; LVDP reduced by33.8±3.6%,33.3±3.2%and43.3±6.1%, which were higher than9.7±1.4%in thecontrol （P<0.01vs. control）;+dp/dt decreased by29.4±5.3%,25.7±5.0%and40.7 ±7.7%, higher than7.9±1.9%in the control group （P<0.01vs. control）;-dp/dtdecreased by31.7±4.9%,32.5±5.9%and46.5±6.2%, higher than9.8±3.1%in thecontrol group （P<0.01vs. control）; HR and CF also decreased in varying degrees.These results suggest that visfatin played a negative inotropic effect on theisolated perfused rat heart.（2）. Pretreatment with visfatin improved the heartfunction of the IRI rat heart, ischemia-reperfusion rat heart, including LVEDP,LVDP and±dp/dt. After reperfusion, LVEDP of the isolated perfused rat heart rosesharply, but50nmol/l,100nmol/l and200nmol/l visfatin reduced the increase ofLVEDP after reperfusion. After reperfusion for5min,10min,15min and20minin the200nmol/l visfatin group, LVEDP decreased to42.4±1.5mmHg,44.3±2.5mmHg,43.2±2.2mmHg and42.4±2.8mmHg respectively, which weresignificantly lower than those in the control group （p<0.05vs. control, n=6）. Afterreperfusion for5min,10min,15min and20min in the200nmol/l visfatin group,LVDP was37±6%,42±6%,45±6%and44±5%respectively;+dp/dt was65±4%,77±6%,69±3%and66±5%respectively;-dp/dt was72±7%,76±3%,74±6%and71±6%, respectively, which were all significantly higher than those in the controlgroup at the same time （p<0.05or p<0.01, n=6）. No statistical difference in LVSP,HR and CF was observed at each time point of reperfusion in50nmol/l,100nmol/l and200nmol/l visfatin groups, as compared with the control group. Theabove results show that200nmol/l visfatin pretreatment significantly improvedthe cardiac function of the IRI isolated rat heart.（3）. Effects of ectogenousvisfatin on [Ca²⁺]i: In the visfatin group, exposure to200nmol/l visfatin resultedin a significant decrease in systolic [Ca²⁺]i; after300s systolic [Ca²⁺]idecreasedfrom38.54±2.67to23.59±8.47; diastolic [Ca²⁺]ialso decreased insignificantly,thus decreasing the amplitude of [Ca²⁺]itransients of single rat ventricularmyocytes.（4） The effects of visfatin on ICa, L, INa/Caand IK, ATPof single rat ventricular myocytes: Visfatin concentration dependently decreased ICa, Lof the ratventricular myocytes. After perfusion with20nmol/l,100nmol/l and500nmol/lvisfatin for5min, ICa, Ldecreased by9.3±5.0%（n=6）,22.2±8.2%（n=6） and47.0±8.2%（n=6） respectively; the figures of the the latter two groups weresignificantly higher than3.6±1.7%in the control group （n=8）（p<0.05）. Visfatinconcentration-dependently increased the inwardINa/Caand outwardINa/Ca. Afterstable perfusion with extracellular fluid for5min, inwardINa/Caand outwardINa/Cain the control group （n=8） decreased by5.3±1.6%and2.8±2.4%respectively.Perfusion with20nmol/l （n=6）,100nmol/l （n=6） and500nmol/l （n=5） visfatin for5min, inwardINa/Caincreased by19.6±4.8%,24.0±2.5%and35.3±2.4%respectively, which were significantly higher than those of the control group（P<0.01）. Perfusion with100nmol/l and500nmol/l visfatin for5min increasedthe outwardINa/Caby5.7±1.7%and15.7±2.7%respectively, which weresignificantly higher than those of the control group （P<0.05）, while there was nosignificant changeand after5-min perfusion with20nmol/l visfatin, as comparedto the control group. In addition, IK, ATPin the membrane of single rat ventricularmyocytes was measured using whole-cell voltage-clamp technique. But even whenthe concentration of visfatin was increased to500nmol/l, the IK, ATPcurveunderwent no significant alteration after perfusion with visfatin for5min,indicating that visfatin could not open the quiescent IK, ATP. Further work is neededto identify the actual effect of visfatin on IK, ATPin the membrane of rat ventricularmyocytes.Conclusion: Visfatin concentration-dependently inhibited LVDP, LVSP and±dp/dt of the isolated perfusion rat heart, playing a negative inotropic effect.Visfatin concentration-dependently improved IRI in the isolated perfusion ratheart by improving the systolic and diastolic function, playing the role of myocardial protection. Visfatin decreased [Ca²⁺]itransients of single ratventricular myocytes, concentration-dependently inhibited L-type calcium channelcurrent in rat ventricular myocytes, increased inward sodium-calcium exchangecurrent, reduced the intracellular calcium ion concentration, and reduced calciumoverload, thus weakening the contractile function of myocardial cells, playing arole in protecting the myocardium if the IRI rat heart.