佛波酯处理人胃癌MGC80-3细胞过程中磷酯酶C-γ2的意义

目的 探讨佛波酯(TPA)处理人胃癌MGC80-3细胞过程中磷酯酶C-γ2(PLC-γ2)的意义.方法 通过DAPI染色,荧光显微镜观察分析TPA对胃癌MGC80-3细胞的影响;借助核浆分离手段获得胃癌细胞核浆蛋白,并通过免疫印迹(Western blotting)检测TPA对PLC-γ2蛋白表达水平影响;通过免疫荧光技术处理,激光扫描共焦显微镜观察胃癌细胞内PLC-γ2蛋白的定位和转运;用PLC-γ2的抑制剂(U73122)预处理细胞,免疫印迹和激光扫描共焦显微镜分别检测其对TPA作用胃癌细胞内PLC-γ2的影响;以及荧光显微镜下观察分析U73122是否影响TPA对胃癌细胞的作用.结果 TPA诱导胃癌MGC80-3细胞凋亡;同时TPA提高PLC-γ2蛋白表达水平,并诱导其发生核浆转运;其抑制剂(U73122)可以降低TPA对PLC-γ2蛋白表达水平的作用,但并不能影响TPA诱导胃癌细胞凋亡;而TPA诱导的PLC-γ2核浆转运却没有被PLC-γ2抑制剂(U73122)阻止.结论 尽管TPA提高了胃癌MGC80-3细胞中PLC-γ2表达水平,但PLC-γ2表达水平和TPA诱导的胃癌MGC80-3凋亡没有直接关系,而其核浆转运可能与TPA诱导的胃癌细胞凋亡相关.

THE IMPLICATION OF PLC-γ2 IN GASTRIC CANCER MGC80-3 CELLS TREATED BY TPA

Objective To study the implication of phosphoinositide specific phospholipase Cγ2(PLC-γ2) in gastric cancer MGC80-3 cells treated by TPA. Methods The effect of 12 O tetradecanoylphorbol 1, 3 acetate(TPA) on gastric cancer MGC80-3 cells was detected and analyzed by DAPI staining under fluorescence microscope. The effect of TPA on the expression level of PLC-γ2 protein was detected by Western blotting, when nuclear and cytoplasmic protein fractions were prepared through lysis of cell and centrifugation. Localization and translocation of PLC-γ2 were observed under laser scanning confocal microscope with immune fluorescence technique. After MGC80-3 cells were pretreated by U73122(PLC-γ2 inhibitor), the effect of TPA on PLC-γ2 was detected by Western blotting and laser scanning confocal microscope with immunefluorescence technique, and the effect of U73122 on MGC80-3 cells followed by TPA was observed and analyzed by DAPI staining under fluorescence microscope. Results TPA induced the apoptosis of gastric cancer MGC80-3 cells. Meanwhile, TPA increased the expression level of PLC-γ2 protein and induced its translocation from cytoplasm to nucleus. U73122 inhibited the effect of TPA on the expression level of PLC-γ2 protein, but it did not affect on the cell apoptosis induced by TPA. However, the translocation of PLC-γ2 protein induced by TPA was not inhibited by its inhib