p53上调凋亡调控因子对不同p53表型胶质瘤细胞生长的影响  

Influence of p53-upregulated modulator of apoptosis on the growth of glioma cells with different p53 phenotypes

作　　者：王新星[1] 苗旺[1] 王宏勤[1] 刘晓东[1] 范益民[1] 

机构地区：[1]山西医科大学第一医院神经外科,太原030001

出　　处：《肿瘤研究与临床》2013年第6期385-388,共4页Cancer Research and Clinic

基　　金：山西省科技攻关项目（20110313011-3）;山西省卫生厅科技攻关项目（20100105、20100224）;山西省基础研究计划（青年科技研究基金）（2012021035-4）

摘　　要：目的观察p53上调凋亡调控因子（PUMA）对不同p53表型人类脑胶质瘤细胞系生长的影响，并探讨其可能的作用机制。方法构建PUMA腺病毒（Ad—PUMA）及空载体腺病毒（Ad．DsRed），分别转染胶质瘤细胞系U251（p53突变型）及SHG-44（p53野生型），CCK-8法检测各组细胞增殖能力；流式细胞术检测各组细胞凋亡率；Westernblot法检测PUMA及凋亡相关蛋白bcl-2、Bax的表达；Caspase活性检测试剂盒检测Caspase-3、Caspase-8、Caspase-9的活性。结果在p53状态不同的胶质瘤细胞系U251及SHG-44中，Ad．PUMA转染组与空载体组及空白对照组比较，均显现出细胞增殖抑制效应抑制率分别为（50．89±4．73）％和（44．45±5．33）％，P〈O．05]以及促凋亡作用凋亡率分别为（44．89±5．08）％和（31．67±7．32）％，P〈0．051；Westernblot结果显示，Ad—PUMA组胶质瘤细胞转染后，可见PUMA蛋白表达升高，同时伴随抗凋亡蛋白bcl-2表达降低及促凋亡蛋白Bax表达增强（P〈0．05）；Caspase活性检测结果显示Ad—PUMA组Caspase-3、Caspase-9活性增加（P〈0．05），而Caspase-8活性未见明显改变（P〉0．05）。结论无论p53表型如何，PUMA均可抑制胶质瘤细胞增殖，促进其凋亡，其机制可能是通过线粒体凋亡途径即上调Bax，抑制bcl-2表达，进而激活Caspase-9实现的，Ad—PUMA有望成为胶质瘤基因治疗的新靶点。Objective To observe the influence of p53-upregulated modulator of apoptosis （PUMA） on the growth of human brain glioma cell lines U251 （p53 mutant） and SHG-44 （p53 wild type）, and to explore its possible mechanism. Methods Construct the adenovirus PUMA （Ad-PUMA） and vector of adenovirus （Ad- DsRed） which were respectively transfected into glioma cell lines U251 and SHG-44. Cells proliferation rates were measured with cell counting kit-8 （CCK-8）. The apoptotic ratios were detected by flow cytometry. The expression of PUMA and apoptosis associated proteins （bcl-2, Bax） were determined with Western blot analysis. Caspase-3, Caspase-g, Caspase-9 activity were measured by Caspase activity assay kit. Results Compared with vector group and blank control group, Ad-PUMA transfected group showed strong cell proliferating inhibition effects [the inhibition rates were （50.89＋_.4.73） % and （44.45＋5.33） % respectively, P 〈 0.05] and pro-apoptotic effects [apoptotic rates were （44.89＋5.08） % and （31.67＋7.32） %, P 〈 0.05] in different p53 glioma cell lines U251 and SHG-44. Western blot analysis showed that PUMA protein expression increased after Ad-PUMA transfection, accompanied by the reduced expression of the anti-apoptotic protein bcl-2 and the increased expression of pro-apoptotic protein Bax. The activity of Caspase testing results showed that the Caspase-3, Caspase-9 activity increased significantly, while the Caspase-8 activity changed little. Conclusion No matter how p53 phenotype, PUMA can inhibit glioma proliferation, promote apoptosis, and its mechanism may be through the mitochondrial apoptotic pathways, upregulation of Bax and inhibition of bcl-2 expression, which activated Caspase-9. Ad-PUMA is expected to become a new target for gene therapy of gliomas.

关 键 词：神经胶质瘤 p53上调凋亡调控因子 P53 细胞增殖 细胞凋亡 

分 类 号：R73[医药卫生—肿瘤]