Tat蛋白对人脑内皮细胞闭锁蛋白表达的影响  

Effect of Tat protein on occludin in human brain endothelial cells

作　　者：徐瑞芬[1] 赵清侠[1] 吴刚[2] 郭姣[1] 侯敏娜[1] 冯旭阳[3] 

机构地区：[1]陕西省人民医院麻醉科,西安710068 [2]西安交通大学医学院第二附属医院麻醉科,陕西西安710004 [3]第四军医大学附属西京医院心脏内科,陕西西安710032

出　　处：《现代医药卫生》2017年第1期37-39,共3页Journal of Modern Medicine & Health

基　　金：陕西省社会发展科技攻关项目(2016SF-045)

摘　　要：目的探讨重组Tat蛋白对人脑内皮细胞闭锁蛋白的影响。方法采用Weksler′s方法分离和处理人脑血管内皮细胞。采用一步法定量反转录酶-聚合酶链式反应(RT-PCR)分析基因的m RNA转录水平。依据Reyes′s法,利用蛋白免疫印迹实验检测闭锁蛋白的表达,并进行免疫荧光检测,荧光显微镜观察分析图像。结果重组Tat蛋白可以降低闭锁蛋白的转录和蛋白的表达。在重组Tat蛋白处理30 min后闭锁蛋白的m RNA水平开始出现明显抑制(P<0.05),重组Tat蛋白处理1、2、6、12、24 h时闭锁蛋白的m RNA水平分别为0.66、0.55、0.58、0.54、0.53,与对照组(重组Tat蛋白处理即刻,0 min)比较,差异有统计学意义(P<0.05)。闭锁蛋白水平在6 h内保持不变,12、24 h时分别为0.71、0.61(P<0.05)。同时,荧光显微镜也观察到了组织学改变。未处理组闭锁蛋白的染色强且均匀。细胞经重组Tat蛋白处理24 h后,闭锁蛋白的染色明显减弱。结论重组Tat蛋白能够破坏血-脑脊液屏障的完整性,可以抑制闭锁蛋白的表达。Objective To investigate the effect of recombinant Tat protein on occludin in brain endothelial cells.Methods Human brain microvascular endothelial cells（HBMECs） were isolated and treated according to the Weksler′s method.The levels of gene mRNA transcripts were analyzed by one-step quantitative reverse transcriptase-polymerase chain reaction（RTPCR）. Occludin expression was detected by Western blot according to Reyes′s method. Immunofluorescence detection was conducted. The images were observed and analyzed by fluorescence microscope. Results The recombinant Tat protein could reduce occludin transcription and protein expression. The occludin mRNA level began to appear obvious inhibition after 30 min of re combinant Tat protein treatment（P〈0.05）. The occludin mRNA levels at 1,2,6,12,24 h of recombinant Tat protein treatment were 0.66,0.55,0.58,0.54,0.53 respectively,showing statistical differences compared with the control group（P〈0.05）. The occludin level maintain unchanged within 6 h,which was 0.71 and 0.61 at 12,24 h（P〈0.05）. Meanwhile,the histological alterations of occludin were also observed by fluorescence microscope. In the untreated group,occludin staining was strong and uniformly. After Tat treating cells for 24 h,the occludin staining was weakened obviously. Conclusion Recombinant Tat protein can disrupt blood brain barrier（BBB） integrity and inhibits occluding expression.

关 键 词：血脑屏障 基因产物 tat 蛋白质类 脑 内皮细胞 

分 类 号：R743[医药卫生—神经病学与精神病学]