诱导型一氧化氮合酶对β-肾上腺受体兴奋诱导老年大鼠心脏功能和心肌细胞凋亡的影响

Effect of inducible nitric oxide synthase on the cardiac function and myocardial apoptosis in rats induced by beta-adrenergic receptor stimulation

目的:探讨心脏中诱导性一氧化氮合酶(induciblenitricoxidesynthase,iNOS)与心肌缺血性损伤和β-肾上腺受体兴奋之间的关系。方法:实验选用雄性Fisher大鼠50只。随机将其中20只大鼠(鼠龄3～5个月)分为青年大鼠对照组和青年大鼠干预组各10只;20只老年大鼠(鼠龄24～25个月)分为老年大鼠对照组和老年大鼠干预组各10只。余下年轻和老年大鼠各5只,处死后,取心肌组织作一氧化氮合酶Westernblot研究。青年大鼠对照组大鼠和老年大鼠对照组大鼠实验前接受生理盐水干预,青年大鼠干预组和老年大鼠干预组大鼠接受特异性iNOS阻断剂-1400W干预。4组大鼠的离体心脏分别接受生理盐水和1400W干预及50%正常冠状动脉流量灌注30min后,泵入异丙肾上腺素(isoproterenol,Iso)30min。结果:心肌梗死后30min,所有组左心室形成压leftventriculardeveloped(pressure,LVDP),左室内压最大上升下降速率rateofriseofleftventric-(ularpressure,±dp/dt)均较基础值下降13%～45%。Iso泵入青年大鼠对照组和青年大鼠干预组大鼠30min后,心室功能部分恢复,表现为LVDP和±dp/dt上升。与此明显对比,Iso泵入老年大鼠对照组大鼠后,非但未改善缺血所致的心功能不全,反倒进一步加重心功能损伤。表现为LVDP和±dp/dt进一步下降43%～60%。

AIM:To investigate the relationship of inducible nitric oxide synthase(iNOS) in heart with myocardial ischemic injury and beta adrenergic receptor(β AR) stimulation. METHODS:Fifty male Fisher rats were used in this study.Twenty rats(3 to 5 months old) were randomly divided into young control group(n=10) and young interventional group(n=10),another 20 aging rats(24 to 25 months old) were randomized into aging control group(n=10) and aging interventional group(n=10).The myocardial tissues of the rest 5 young rats and 5 aging rats were taken after being killed for Western blot study of nitric oxide synthase(NOS).Rats in the young and aging control groups were treated with saline before experiment,while those in the young and aging interventional groups received intervention of specific iNOS blocker- 1 400 W.After interventions of saline and 1 400 W and perfusion of 50% normal coronary flow reduction for 30 minutes,the isolated hearts were pumped with isoproterenol(Iso) stimulation for 30 minutes. RESULTS:Thirty minutes after myocardial infarction, left ventricular developed pressure(LVDP) and rate of rise and decrease of left ventricular pressure(± dp/dt) in all the groups were decreased by 13% to 45% as compared with the basic values.Thirty minutes after Iso was pumped into the rats of the young control and interventional groups,the ventricular functions were recovered partially,which was manifested by increase of LVDP and± dp/dt.In contrast, after Iso was pumped into the rats of the aging control group,the cardiac insufficiency induced by ischemia was not ameliorated,but the cardiac injury was aggregated further,which was manifested that LVDP and ± dp/dt were further decreased by 43% to 60% . Western blot studies showed a phenotypic change of increase in iNOS expression in the aging heart,the total nitric oxide(NO) production after ischemia for 60 minutes in the four groups were(134.78± 10.55),(138.78± 19.10),(280.50± 29.58),(127.84± 13.58) μ mol/L respectively,it was significantly higher in the aging control group than in the other three groups(P< 0.01).The 1 400 W treatment stopped the dysfunctional response to Iso stimulation, blocked the synthesis of NO and 3 nitrotyrosion, and reduced the release and activity of myocardial apoptosis caspase 3.The content of 3 nitrotyrosin in the aging interventional group[(18.72± 0.56) μ mol/g] was obviously lower than that in the aging control group[(43.94± 1.01) μ mol/g](P< 0.01),but had insignificant difference from that in the young control group[(14.12± 0.14) μ mol/g](P >0.05).The myocardial content of caspase 3(DEVD pNA) was obviously lower in the aging interventional group[(189.57± 12.18) mmol/g] than in the aging control group[(524.52± 13.76) mmol/g](P< 0.01). CONCLUSION:Aging induces increased levels of iNOS expression in heart.Upregulation of iNOS can release NO rapidly in responses to ischemia and Iso stimulation.Consequently,the increased nitrative stress results in activation of apoptotic process,and predisposes the aging heart to cardiac dysfunction and myocardial ischemic injury.