TNFR1与PDC-E2对胶质瘤细胞凋亡影响的相关性研究

摘要

Objective:To investigate the expression variation of TNFR1,PDC-E2 and the relative mechanism during the process of apoptosis of glioma cells,and investigate the role of TNFR1 and PDC-E2 during the process.Methods:To transfect TNFR1 siRNA into U87MG cells to detect the affect of glioma apoptosis and the interactions of TNFR1 and PDC-E2 in vitro by immune coprecipitation.To test the cell location of TNFR1 and PDC-E2 in U87MG by immunofluorescence.By transfecting with TNFR1 siRNA glioma cells,effect of the interactions of TNFR1 and PDC-E2 to Caspase-3 was tested.Results:When TNFR1 siRNA were transfected into U87MG cells respectively,the results showed that TNFR1 could inhibit the apoptosis of glioma cells,and the inhibition effect of TNFR1 on the apoptosis of glioma cells was decreased.There was interaction of TNFR1 and PDC-E2 in HEK293T cells and glioma by immune coprecipitation and immunofluorescence.The interaction of TNFR1 and PDC-E2 was detected in glioma cells to promote the expression of Caspase-3.Conclusion:TNFR1 and PDC-E2 could inhibit the apoptosis of glioma cells,and the synergistic effect to apoptosis was more obvious than the individual effect.TNFR1 could interact with PDC-E2 and then co-increase the protein level of Caspase-3.%目的:胶质瘤是一种最常见的中枢神经系统恶性肿瘤,生长迅速,恶性程度高.研究胶质瘤细胞凋亡过程中肿瘤坏死因子受体1(TNFRl)和丙酮酸脱氢酶复合物E2(PDC-E2)蛋白的表达变化,探讨TNFR1和PDC-E2在此过程中的作用.方法:培养TNFR1的小干扰细胞,将干扰掉TNFR1的质粒转染入U87MG细胞,检测TNFR1和PDC-E2对胶质瘤细胞凋亡的影响.通过免疫共沉淀方法检测TNFR1和PDC-E2在体外的相互作用;通过免疫荧光检测两者在U87MG细胞内的共定位;在转染干扰掉TNFR1的胶质瘤细胞中,检测TNFR1和PDC-E2的相互作用对凋亡标记物Caspase-3蛋白表达量的调节作用.结果:将TNFR1的小干扰质粒转染入U87MG细胞,结果显示未干扰掉的TNFR1对胶质瘤细胞凋亡有抑制作用,干扰掉TNFR1时对胶质瘤细胞凋亡的抑制作用减弱.体外HEK293T细胞及胶质瘤细胞中的免疫共沉淀以及免疫荧光显示TNFR1和PDC-E2存在相互作用;在胶质瘤细胞中检测到TNFR1和PDC-E2的相互作用协同促进Caspase-3的表达.结论:TNFR1和PDC-E2抑制胶质瘤细胞的凋亡,协同作用比单独对凋亡的影响更加明显;TNFR1和PDC-E2存在相互作用,两者协同作用更能增高作用底物Caspase-3的表达.