地奥心血康对培养H9c2心肌细胞缺氧/复氧损伤的保护机制

The protective effect of di'aoxinxuekang on cultured H9c2 cardiomyocytes treated by hypoxia/reoxygenation

目的:探讨地奥心血康对培养H9c2心肌细胞缺氧/复氧损伤的保护作用机制。方法:建立缺氧/复氧损伤心肌细胞模型,用地奥心血康进行干预。将培养的H9c2心肌细胞随机分为6组,正常对照组(C组);缺氧/复氧模型组(H/R组);缺氧/复氧模型+吡那地尔(pinacidil,Pin)阳性对照组(H/R+Pin组);缺氧/复氧模型+地奥心血康(DAXXK)低剂量、中、高剂量(10mg/L、20 mg/L、40mg/L)干预组(H/R+L、M、H组)。MTT法测定细胞存活率;用罗丹明123(Rh123)荧光探针标记线粒体,在荧光显微镜下采集被Rh123染色的各组心肌细胞荧光图片;收集培养细胞上清液测定乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)的活性及丙二醛(MDA)的含量。结果:与正常组比较,缺氧/复氧模型组细胞存活率、线粒体膜电位及SOD活性均下降,而LDH活性、MDA含量均升高。地奥心血康用药组可有效提高心肌细胞存活率、升高线粒体膜电位,同时降低受损心肌LDH的释放量,并有效提高SOD的活性,减少MDA的生成量。结论:地奥心血康对缺氧/复氧造成的H9c2心肌细胞损伤具有保护作用,其机制与保护细胞线粒体膜、增强细胞清除氧自由基能力、减少脂质过氧化物的产生有关。

Objective： To study the mechanisms of the protective effects of Di’aoxinxuekang on cultured H9c2 cardiomyocytes treated by hypoxia/reoxygenation.Methods： the hypoxia/reoxygen（H/R） injury cardiomyocyte model was established in H9c2 cell strain with or without Di’aoxinxuekang treaed.Cultured H9c2 myocardial cells were randomly divided into six groups：control group（C group）;H/R group;H/R ＋ pinacidil（H/R＋Pin） group;H/R ＋ Di’aoxinxuekang low concentration group（10mg/L）,H/R ＋ Di’aoxinxuekang middle concentration group（20mg/L）,H/R＋ Di’aoxinxuekang high concentration group（40 mg/L）.Cell survival rate was determined using the 3-（4,5-dimethylthiazol-2-y1）2,5-diphenyltetrazolium bromide（MTT） assay.Mitochondrial membrane potential（△Ψm） changing was estimated using Rhodamine 123（Rho123） which is a fluorescent explorer.the fluorescence imaging of H9c2 cardiomyocyte stained with Rho123 was obtained from the fluorescence microscopy.The activity of lactate dehydrogenase（LDH）,superoxide dismutase（SOD） and the content of malondialdehyde（MDA） was measured respectively in cellular supernate.Results： Compared with C group,the cell survival rate,mitochondrial membrane potential（△Ψm） and the activity of SOD of H/R model group were all decreased.The activity of LDH and the content of MDA were all increased.The groups being treated with Di’aoxinxuekang could effectively increase the cell survival rate and mitochondrial membrane potential（△Ψm） and decrease the release of LDH.Moreover,they could increase the activity of SOD and decrease the production of MDA.Conclusion： It is suggested that Di’aoxinxuekang has a protective effect on H9c2 cardiomyocyte treated by H/R injury.The mechanisms of the protective effects of Di’aoxinxuekang are related to protect mitochondria membrane from hypoxia/reoxygen（H/R） injury and to enhance the capability of the cell clearing the oxygen free radical and to decrease the production of lipid peroxidation.