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目的研究FRK是否通过调节表皮生长因子受体(EGFR)-Y1173磷酸化促进胶质瘤细胞的凋亡。方法应用PolyJet~(TM)将FRK质粒转染入脑胶质瘤U251细胞中,western blot(WB)检测质粒转染效果及EGFR蛋白水平和EGFR-Y1173磷酸化水平的变化,流式细胞术检测脑胶质瘤细胞凋亡的变化;转染EGFR(WT)、EGFR(Y1173F)质粒,或共转FRK和EGFR(WT)、EGFR(Y1173F)质粒,流式细胞术检测脑胶质瘤U251细胞凋亡的变化。结果成功将FRK质粒转染入脑胶质瘤U251细胞,过表达FRK促进脑胶质瘤U251细胞的凋亡。过表达FRK增加EGFR-Y1173磷酸化水平,而对EGFR蛋白水平无影响。过表达EGFR(WT)、EGFR(Y1173F)质粒,细胞凋亡均减少,分别减少了25%和75%。WB结果显示,共转FRK和EGFR(WT)、EGFR(Y1173F)质粒成功;流式细胞术结果显示,与对照组相比,过表达FRK促进了细胞凋亡,EGFR抑制了细胞凋亡,且转染EGFR(Y1173F)质粒组的作用比转染EGFR(WT)的效果更明显。此外,转染EGFR(Y1173F)质粒可逆转过表达FRK对胶质瘤细胞的促凋亡作用。结论 FRK可以通过促进EGFR-Y1173磷酸化从而调节胶质瘤细胞的凋亡。
Objective To investigate whether FRK can promote the apoptosis of glioma cells by regulating the phosphorylation of epidermal growth factor receptor (EGFR) -Y1173. Methods The FRK plasmid was transfected into glioma U251 cells with PolyJet ™. The transfection efficiency and the EGFR protein level and phosphorylation of EGFR-Y1173 were detected by Western blot (WB). Flow cytometry Glioma cells were transfected with EGFR (WT) and EGFR (Y1173F) plasmids or co-transfected with FRK and EGFR (WT) and EGFR (Y1173F) plasmids. Flow cytometry was used to detect glioma U251 cells Apoptosis changes. Results FRK plasmid was successfully transfected into glioma U251 cells and overexpression of FRK promoted the apoptosis of glioma U251 cells. Overexpression of FRK increased EGFR-Y1173 phosphorylation, but had no effect on EGFR protein level. Overexpression of EGFR (WT) and EGFR (Y1173F) plasmids reduced apoptosis by 25% and 75%, respectively. WB results showed that the co-transfected FRK and EGFR (WT) and EGFR (Y1173F) plasmids were successful. The results of flow cytometry showed that overexpression of FRK promoted apoptosis and EGFR inhibited apoptosis compared with the control group The effect of transfecting the EGFR (Y1173F) plasmid group was more pronounced than transfection of EGFR (WT). In addition, transfection of the EGFR (Y1173F) plasmid reverses the pro-apoptotic effect of FRK over glioma cells. Conclusion FRK can regulate glioma cell apoptosis by promoting EGFR-Y1173 phosphorylation.