高表达RNA结合模序蛋白5促进胶质瘤A172细胞凋亡  

High-expressed RNA binding motif protein-5 promotes the apoptosis of A172 glioblastoma multiforme cells

作　　者：黄浩洋[1] 钟东[1] 张福安[1] 吕东[1] 李炯[1] 石爽[1] Huang Haoyang;Zhong Dong;Zhang Fuan;Lyu Dong;Li Jiong;Shi Shuang(Department of Neurosurgery,the First Affiliated Hospital of Chongqing Medical University,Chongqing 400016,China)

机构地区：[1]重庆医科大学附属第一医院神经外科,400016

出　　处：《中华神经外科杂志》2018年第8期836-840,共5页Chinese Journal of Neurosurgery

基　　金：财政部、卫生部国家临床重点专科建设项目[财社（2011）170号];重庆市科委自然科学基金（cstc2011jjA10091）

摘　　要：目的 探讨RNA结合模序蛋白5（RBM-5）的高表达促进A172胶质瘤细胞株凋亡的机制.方法 回顾性纳入重庆医科大学附属第一医院神经外科2015年8月至2017年6月手术切除的19例脑胶质瘤标本,采用免疫组织化学染色方法检测RBM-5的表达.采用定量实时聚合酶链反应（qRT-PCR）检测RBM-5在人脑胶质瘤细胞株A172、U87、U251及SHG-44的表达.构建稳定感染高表达RBM-5慢病毒的A172细胞株,采用qRT-PCR验证RBM-5在3组细胞株中（分别为实验组、阴性对照组和空白对照组）mRNA水平的表达;进一步采用蛋白质印迹法检测RBM-5、BAX、Bcl-2和Cleaved Caspase-9在3组细胞株中的表达;最后采用流式细胞术检测3组细胞株的凋亡情况.结果 免疫组织化学染色结果显示,RBM-5在低级别胶质瘤（WHO Ⅰ～Ⅱ级,A值分别为0.021±0.011、0.004 ±0.002）中的表达高于高级别胶质瘤（WHOⅢ～Ⅳ级,A值均为0.000±0.000）（F=7.410,P＜0.05）.qRT-PCR结果显示,RBM-5在A172、U251、U87和SHG-44细胞株中的表达依次为0.494±0.186、1.168 ±0.775、2.437±1.281和3.547±1.177,其中RBM-5在A172中表达最低,在SHG-44中表达最高（F=6.023,P＜0.05）.蛋白质印迹实验证实,高表达RBM-5的A172细胞株中RBM-5 （202.854±6.573）、BAX（208.000±10.324）和Cleaved Caspase-9（112.811±1.684）的表达均较阴性对照组（分别为117.156 ±0.357、152.105±2.819和62.873±1.313）和空白对照组（分别为119.283±1.356、153.159 ±0.571和73.281 ±4.480）高;而Bcl-2的表达（60.420±1.986）均较阴性对照组（107.157 ±2.056）和空白对照组（99.563±3.997）低.流式细胞术结果显示,实验组细胞凋亡率[（20.88 ±0.72）％]高于阴性对照组[（9.51±0.57）％,P＜0.05]和空白对照组[（9.49±0.49）％,P＜0.05].结论 在A172胶质瘤细胞株中高表达的RBM-5可能通上调BAX和CleavedCaspaseO蛋白、下调Bcl-2蛋白促进其凋亡,提示RBM-5可能成为脑胶质瘤临床治疗的新靶点.Objective To investigate the apoptotic mechanism of A172 cells with high-expressed RNA binding motif protein （RBM-5）.Methods Nineteen glioma specimens,gathered from Neurosurgery Department of the First Affiliated Hospital of Chongqing Medical University from August 2015 to June 2017,were retrospectively brought into this research and immunohistochemistry was performed to detect the expression of RBM-5.Quantitative real-time polymerase chain reaction （qRT-PCR） was used to measure the discrepant expression of RBM-5 in A172,U87,U251 and SHG-44 human glioma cells.The lentiviral synthesized in vitro was performed to transfect A172 cell line.Afterwards,the expression of RBM-5 in mRNA level of cells among three groups （namely experimental group,control group and blank group） was evaluated using qRT-PCR.RBM-5,BAX,Bcl-2 and Cleaved-caspase-9 in cells of 3 groups were assessed using Western Blot.Finally,the apoptotic effects of cell lines among 3 groups were investigated using flow cytometry.Results Immunohistochemistry demonstrated that RBM-5 in low-grade glioma tissues （WHO Ⅰ-Ⅱ,A values respectively represented 0.021 ± 0.011 and 0.004 ± 0.002） displayed significantly more expression than that in high-grade glioma tissues （WHO Ⅲ-Ⅳ,A values represented 0.000 ± 0.000） （F =7.410,P ＜ 0.05）.The qRT-PCR results showed that the expression values of RBM-5 in A172,U251,U87 and SHG-44 cells were 0.494 ± 0.186,1.168 ± 0.775,2.437 ± 1.281 and 3.547 ± 1.177,respectively,which implied that RBM-5 displayed the least expression in A172 cell line and,on the contrary,the most in SHG-44 cell line （F =6.023,P ＜ 0.05）.The results of Western Blot verified that RBM-5 （202.854 ± 6.573）,BAX （208.000 ± 10.324） and Cleaved-Caspase-9 （112.811 ± 1.684） of A172 cell line with high-expressed RBM-5 represented obviously more expression than those of control group （117.156 ± 0.357,152.105 ± 2.819 and 62.873 ± 1.313,respectively） and those of blank group （119.283 ± 1.356,153.159 ±0.5

关 键 词：神经胶质瘤 细胞凋亡 RNA结合模序蛋白5 

分 类 号：R739.4[医药卫生—肿瘤]