摘要：
Objective To investigate the effect of diazoxide (DZ) postconditioning mediating the expression of phospho glycogen synthase kinase-3β,Bcl-2,Bax and the cell viability during hypoxia/reoxygenation (H/R) in cultured adult rat cardiac myocytes.Methods The model of isolated adult rat cardiac myocytes was established and randomly divided into 5 groups (n=6):① Normal group:caridocytes were incubated at 37 °C in a humidified atmosphere of 5％ carbon dioxide CO2) and 95％ air for 6 h; ② H/R group:cardiocytes were exposed to 3 h of hypoxia followed hy 3 h of reoxygenation; ③ DZ group:cardiocytes were exposed to 3 h of hypoxia followed by 3 hof reoxygenation,while 100 μ mol/L DZ was added in medium 5 min after reoxygenation; ④ DZ+5-hydroxydecanoate(5-HD) group:cardiocytes were exposed to 3 h of hypoxia followed by 3 h of reoxygenation,while 100 μmol/L 5-HD was added in medium rightly after 3 h' hypoxia and 100 μmol/L DZ was added in medium 5 min after reoxygenation; ⑤ 5-HD group:3 h of hypoxia followed by 3 h of reoxygenation,while 100 μmol/L 5-HD was added in medium after 3 h' hypoxia.The cell viability was assayed by the rate of Rod-shaped cells; the expression of pGSK-3β,Bcl-2 and Bax were assessed by western blot 3 h after reoxygenation.Results After 3 h reoxygenation,contraction of the single myocyte is (13.12±0.19)％ in normal group.Compared with the normal group,the cell viability was gready decreased in the other 4 groups [(7.97±0.22)％ for H/R group,(10.48± 0.20)％ for DZ group,(7.97±0.19)％ for the DZ+5-HD group,(8.22±0.22)％ for the 5-HD group](P＜0.05).The cell viability in DZ group was(64±5)％.Compared with the H/R group,it was significantly increased(P＜0.05).The content of Bcl-2 and pGSK-3β in DZ group were higher(P＜0.05),while the expression of Bax was lower(P＜0.05).But these effects were abolished with administration of 5-HD rightly after hypoxia(P＞0.05); There was no statistical difference between H/R group and 5-HD group(P＞0.05).Conclusions DZ could alleviate hypoxia/reoxygenation injury through mediating the expression of pGSK-3β,Bax,Bcl-2 proteins via opening of mitoKATP channel.%目的 研究线粒体ATP-敏感性钾通道(mitochondrial ATP-sensitive potassium channel,mitoKATP通道)开放剂二氮嗪(diazoxide,DZ)后处理对成年大鼠心肌细胞缺氧/复氧(hypoxia/reoxygenation,H/R)后细胞存活及对磷酸化糖原合成激酶-3β(phospho glycogen synthase kinase-3β,pGSK-3β),Bcl-2,Bax表达的影响. 方法 体外培养原代成年大鼠(30只)心肌细胞建立H/R损伤模型,按随机数字表法随机分为5组(每组6只):①Normal组:在二氧化碳(CO2)培养箱中持续培养6h组；②H/R组:缺氧3h复氧3h组；③DZ组:缺氧3h复氧3h,复氧5 min时给予100 μmol/L DZ组；④DZ+5-HD组:缺氧3h复氧3h,缺氧末给予100 μmol/L 5-羟葵酸盐(5-hydroxydecanoate,5-HD),复氧5 min时给予100μmol/L DZ组；⑤5-HD组:缺氧3h复氧3h,缺氧末给予100 μmol/L 5-HD组.复氧3h末通过计数细胞长杆率测定存活率,免疫印迹法测定心肌细胞内pGSK4β,Bcl-2和Bax的表达. 结果 复氧3h末,Normal组单个心肌细胞收缩幅度为(13.12±0.19)％,与Normal组比较,其他各组单个心肌细胞收缩幅度明显降低[H/R组为(7.97±0.22)％,DZ组为(10.48±0.20)％,DZ+5-HD组为(7.97±0.19)％,5-HD组为(8.22±0.22)％](P＞0.05),心肌细胞内Bcl-2表达水平降低,Bax表达水平升高(P＜0.05).DZ后处理组心肌细胞存活率为(64±5)％,与H/R组比较明显增高(P＜0.05),心肌细胞内Bcl-2、pGSK4β表达水平升高,Bax表达水平降低,而缺氧末即刻给予5-HD可以逆转DZ后处理的这些作用(P＜0.05)；5-HD组与H/R组比较差异无统计学意义(P＞0.05). 结论 DZ后处理可能通过激活mitoKATP通道、上调pGSK-3β及Bcl-2,下调Bax的表达增加H/R后成年大鼠心肌细胞的存活.
