摘要
目的探讨钙离子/钙调蛋白依赖性蛋白激酶II(CaMK II)在H9c2心肌细胞缺氧/复氧(H/R)模型中的作用。方法将H9c2心肌细胞分为4组:正常对照组(Control)、H/R组、CaMK II抑制剂KN-93+H/R组和KN-93组。H/R组是对H9c2心肌细胞进行缺氧4 h,再复氧6 h处理;后两组是先用KN-93(终浓度1μmol/L)预处理2 h后再进行/不进行H/R损伤。采用MTT法检测各组细胞活力,碘化丙啶(PI)染色观察细胞坏死情况,Western blot法检测受体相互作用蛋白激酶(RIPK)3、磷酸化混合谱系蛋白激酶样结构域蛋白(pMLKL)和pCaMK II蛋白的表达。结果与Control组比较,H/R组细胞活力显著降低,PI阳性细胞数明显增多,RIPK3、pMLKL和pCaMK II的表达水平显著增加(P<0.01);与H/R组比较,KN-93+H/R组细胞活力显著增加,PI阳性细胞数明显减少,RIPK3、pMLKL和pCaMKII蛋白表达明显下调(P<0.01);而KN-93组的各项指标与Control组相比无显著性差异(P>0.05)。结论 CaMK II可通过程序性细胞坏死通路加重H9c2心肌细胞的H/R损伤。
Objective To investigate the role of Ca2+/calmodulin-dependent protein kinase II(CaMK II) in the hypoxia/reoxygenation(H/R) injury model of H9c2 cardiomyocytes. Methods H9c2 cells were divided into control, H/R(hypoxia for 4 hours and reoxygenation for 6 hours), CaMK II inhibitor KN-93+H/R(pretreated with KN-93 for 2 hours before H/R injury) and KN-93(treated with KN-93 for 2 hours only) groups. The activity of H9c2 cardiomyocytes was detected by MTT assay, cell necrosis was observed by propidium iodide(PI) staining. The expressions of receptor interacting protein kinase(RIPK) 3 and phosphorylated mixed lineage kinase domain-like protein(pMLKL) and pCaMK II protein were detected by Western blot. Results Compared with the control group, cell viability was dramatically decreased, the number of PI-positive cells was significantly increased, and the expression levels of RIPK3 and pMLKL and pCaMK II were distinctly increased(P<0.01) in H/R group. However, the cell viability increased markedly, the number of PI-positive cells decreased obviously, and the expressions of RIPK3 and pMLKL and pCaMK II were significantly down-regulated in KN-93+H/R group than in H/R group(P<0.01), but with no significant difference between KN-93 group and control group(P>0.05). Conclusion CaMK II aggravates H/R injury of H9c2 cardiomyocytes via programmed cell necrosis pathway.
作者
杨吉平
费琳
钟钰西
秦泗佳
柴学军
YANG Ji-ping;FEI Lin;ZHONG Yu-xi;QIN Si-jia;CHAI Xue-jun(Institute of Basic Medical Sciences,Shaanxi Key Laboratory of Ischemic Cardiovascular Disease,Xi'an Medical University,Xi'an 710021;Departments of Psychiatry,the First Affiliated Hospital of Xi'an Jiaotong University,Xi'an 710061,China)
出处
《解剖科学进展》
CAS
2021年第5期581-584,共4页
Progress of Anatomical Sciences
基金
陕西省科技厅一般项目(2021JM-492)
陕西省教育厅重点实验室项目(19JS061)
西安医学院博士科研启动基金(2018DOC05)。