PKC在枸橼酸舒芬太尼预处理大鼠心肌细胞缺氧/复氧损伤模型中的作用  

Role of protein kinase C in rat myocardial cells with anoxia-reoxygenation injury preconditioned with sufentanil

作　　者：高小茹[1] 王萌[1] 张冬梅[2] 宋华丽[1] 

机构地区：[1]宁夏医科大学临床学院,银川医学硕士研究生750004 [2]宁夏医科大学总医院麻醉科,银川750004

出　　处：《医学研究生学报》2016年第7期688-692,共5页Journal of Medical Postgraduates

基　　金：国家自然科学基金(81260029)

摘　　要：目的蛋白激酶C(protein kinase C,PKC)信号通路是预处理早期保护机制细胞信号传递的共同通路,PKC是预处理早期保护的必备信号传递物质。文中探讨PKC在枸橼酸舒芬太尼预处理大鼠心肌细胞缺氧/复氧损伤模型中的作用。方法原代心肌细胞分离培养。将5 d的心肌细胞分成对照组、缺氧/复氧组、舒芬太尼组和佛波醇+枸橼酸舒芬太尼组。对照组细胞培养不予任何处理,建立培养乳鼠心肌细胞缺氧/复氧损伤模型,舒芬太尼组加入舒芬太尼至终浓度0.0003μmol/L,佛波醇+舒芬太尼组使用佛波醇干预10 min后再给予枸橼酸舒芬太尼,并在各组相应处理后取材检测细胞增殖情况。免疫荧光共聚焦技术观察Connexin43(Cx43)蛋白的平均光密度,流式细胞术检测各期的凋亡率,Western blot检测细胞缝隙连接蛋白Cx43总蛋白。结果与对照组相比,缺氧/复氧组、舒芬太尼处理组、佛波醇+舒芬太尼组细胞增殖明显增多(P<0.05);与缺氧/复氧组增殖(0.325 8±0.023 5)相比,舒芬太尼组(0.498 0±0.0432 4)、佛波醇+舒芬太尼组(0.724 0±0.123 4)的细胞增殖增多(P<0.05)。与舒芬太尼组增殖(0.498 0±0.043 24)相比,佛波醇+舒芬太尼组(0.724 0±0.123 4)的细胞增殖增多(P<0.05)。流式细胞术检测显示,与对照组相比,缺氧/复氧组、舒芬太尼组、佛波醇+舒芬太尼组早期凋亡率、晚期凋亡率和总凋亡率升高(P<0.05);与缺氧/复氧组早、晚、总凋亡率[(4.96±0.59)%、(18.77±0.92)%、(23.73±0.51)%]相比,舒芬太尼组[(5.86±0.38)%、(10.37±0.38)%、(16.23±0.32)%]、佛波醇+舒芬太尼处理组[(5.71±0.58)%、(5.54±0.43)%、(11.24±0.62)%]均显著降低(P<0.05),其中舒芬太尼处理组较佛波醇+舒芬太尼处理组晚期总凋亡率均低,差异有统计学意义(P<0.05)。结论枸橼酸舒芬太尼对缺氧复氧损伤的心肌具有保护作用,PKC激动剂联合枸橼酸舒芬太尼对心肌细胞缺氧/复氧损伤有叠加的保护�Objective The protein kinase C（PKC） is an essential signaling substance in the early protection of cells in preconditioning.This study was to investigate the role of PKC in the myocardial cells of the rat model of anoxia-reoxygenation（A-R） injury preconditioned with sufentanil.Methods Primary myocardial cells isolated and cultured for 5 days were allocated to a control,an A-R,a sufentanil preconditioning（SF）,and a phorbol＋sufentanil preconditioning（PMA＋SF） group.The A-R injury model was established with cultured myocardial cells from neonatal rats,which were preconditioned with sufentanil at the concentration of0.000 3 μmol/L in the SF group or phorbol followed by sufentanil 10 minutes later in the PMA＋SF group.Then the proliferation of the cells was detected,the optical density of the Cx43 protein observed by immunofluorescence confocal technology,the apoptosis rate of the cells determined by flow cytometry,and the total Cx43 proteins calculated by Western blot.Results Cell proliferation was significantly increased in the A-R,SF,and PMA＋SF groups as compared with the control（P〈0.05）,higher in the SF and PMA＋SF than in the A-R group（0.498 0 ± 0.0432 4 and 0.7240 ± 0.1234 vs 0.325 8 ± 0.023 5,P〈0.05）,and in the SF than in the PMA＋SF group（P〈0.05）.Flow cytometry showed significantly increased rates of early,late,and total cell apoptosis in the A-R（[4.96 ± 0.59],[18.77 ± 0.92],and [23.73 ± 0.51]%）,SF（[5.86 ± 0.38],[10.37 ± 0.38],and [16.23 ± 0.32]%）,and PMA＋SF group（[5.71 ± 0.58],[5.54 ± 0.43],[11.24 ± 0.62]%） as compared with the control（P〈0.05）,remarkably lower in the SF and PMA＋SF than in the A-R group（P〈0.05）,and the late and total cell apoptosis rates markedly lower in the SF than in the PMA＋SF group（P〈0.05）.Conclusion Sufentanil has a protective effect and PKC agonists combined with sufentanil may add to the effect on myocardial cells in A-R injury.

关 键 词：蛋白激酶C 枸橼酸舒芬太尼 缺氧/复氧损伤 CX43 

分 类 号：R96[医药卫生—药理学]