羧基末端连接蛋白反应蛋白对脑血管平滑肌细胞氧化损伤的修复作用  

The effect of CtBP-interacting protein on the oxidative damage of cerebrovascular smooth muscle cells

作　　者：杜郭佳[1] 赛力克·对山拜[1] 更·党木仁加甫 苏日青[1] 范雁东[1] Du Guojia;Sai Like· Dui Shanbai;Geng· Dang Murenjiafu;Su Riqing;Fan Yandong(Department of Neurosurgery, the First Teaching Hospital of Xinjiang Medical University, Xinjiang 830054, China)

机构地区：[1]新疆医科大学第一附属医院神经外科,830054

出　　处：《中华实验外科杂志》2019年第8期1365-1367,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金项目(81760222);新疆维吾尔自治区自然科学基金项目(2019D01A87).

摘　　要：目的观察羧基末端连接蛋白反应蛋白(CtIP)对脑平滑肌细胞氧化损伤的功能作用,并探讨其作用机制.方法通过添加三丁基过氧化氢(TBHP)刺激诱导脑平滑肌细胞氧化应激,采用过表达和干扰慢病毒技术制备CtIP基因表达和沉默表达细胞株,α-平滑肌肌动蛋白(α-SMA)免疫荧光检测细胞的损伤程度,蛋白质印迹法(Western blot)检测细胞CtIP、α-SMA蛋白的表达,实时定量反转录聚合酶链反应(Real-time RT-PCR)检测CtIP信号通路基因的表达,SPSS软件Student-t检验分析组间基因表达差异显著性.结果免疫荧光和Western blot检测结果显示,过表达CtIP后,α-SMA的表达减弱,血管平滑肌细胞损伤程度减轻.而干扰CtIP表达后,α-SMA表达显著增加,血管平滑肌细胞损伤程度提高.Real-time RT-PCR结果显示,CtIP基因显著上调BRCA1,ZBRK1和CDK4基因的表达,分别增加了2.04倍(t=-12.976,P<0.01)、2.72倍(t=-5.976,P<0.01)和2.08倍(t=-7.980,P<O.01),而抑制下调了p21基因的表达近2.05倍(t=10.936,P<0.01).结论 CtIP基因对损伤的脑平滑肌细胞具有显著的修复作用,并确定了CtIP基因与BRCA1,ZBRK1,CDK4和p21基因在损伤过程中的调控关系.Objective To explore the effect of CtBP-interacting protein (CtIP) on oxidative damage of cerebral smooth muscle cells and its mechanism. Methods Cerebral smooth muscle cells were stimulated by TBHP to induce oxidative damage. The cell line of CtIP gene were prepared by over-expression and interfering lentivirus technology. Cell damage was detected by immunofluorescence assay of alpha smooth muscle actin (α-SMA). The expression of CtIP and α-SMA protein was detected by Western blotting, and the related genes of CtIP signaling pathway were detected by Real-time RT-PCR. Student-t test of SPSS software was used to analyze the difference of gene expression between groups. Results The results of immunofluorescence and Western blotting showed that overexpression of CtIP inhibited the level of α-SMA expression with the decreased injury degree of vascular smooth muscle cells. Interfering with CtIP expression resulted in the α-SMA expression increased significantly and vascular smooth muscle cells were damaged more severely. Real-time RT-PCR data showed that expression of CtIP significantly increased the expression of BRCA1, ZBRK1 and CDK4 genes by 2.04 times (t=-12.976, P<0.01), 2.72 times (t=-5.976, P<0.01) and 2.08 times (t=-7.980, P<0.01) respectively, while the expression of p21 gene was decreased by 2.05 times (t=10.936, P<0.01). Conclusion It is confirmed that CtIP gene has the significantly repair effect on injured cerebral smooth muscle cells, and the regulatory relationship between CtIP gene and BRCA1, ZBRK1, CDK4 and p21 genes in the process of injury are determined.

关 键 词：羧基末端连接蛋白反应蛋白基因 氧化损伤 脑血管平滑肌细胞 修复 

分 类 号：R743[医药卫生—神经病学与精神病学]