TGF-β对胃癌细胞干性因子Oct4、Nanog表达的影响及机制

摘要

目的 探讨TGF-β对胃癌细胞干性因子Oct4、Nanog表达的影响.方法 将培养的胃癌MKN45细胞分为对照组、TGF-β(10 ng/mL)组和TGF-β+BAPTA-AM组,对照组不进行任何处理,其余两组分别采用相应试剂刺激24 h.采用qRT-PCR、Western blotting技术检测胃癌相关干性因子Nanog、Oct4的mRNA和蛋白表达水平.高速离子成像系统检测加入正常的PSS液(空白对照组)、加入TGF-β(20 ng/mL)(单独TGF-β组)及加入TGF-β+BAPTA-AM(联合组)作用后胃癌细胞MKN45内Ca2+浓度的变化.结果 与对照组比较,TGF-β组Nanog和Oct4 mRNA及蛋白的表达上调(P均<0.05),与TGF-β组比较,TGF-β+BAPTA-AM组Nanog和Oct4 mRNA及蛋白的表达下调(P均<0.05).与空白对照组比较,TGF-β组细胞内钙增加(P<0.05);与单独TGF-β 组比较,联合组细胞内钙减少(P<0.05).结论 TGF-β可能通过增加细胞内钙浓度而使胃癌细胞干性因子Oct4、Nanog的表达增强.%Objective To investigate the influence of transforming growth factor-β (TGF-β)on the expression of gas-tric cancer stem genes Oct4 and Nanog in gastric cancer cells. Methods The cultured gastric cancer cells MKN45 were divided into the control group,TGF-β (10 ng/ mL)group,and TGF-β + BAPTA-AM group. The control group was not treated and the other two groups were stimulated with the corresponding reagent for 24 h. The mRNA and protein expression levels of Nanog and Oct4 were detected by qRT-PCR and Western blotting. High-speed ion imaging system was used to ex-amine the changes of intracellular Ca2 + concentrations in gastric cancer MKN45 cells with normal PSS solution (control group),TGF-β (20 ng/ ml)(TGF-β group)and intracellular calcium chelator BAPTA-AM (BAPTA-AM group). Results Compared with the control group,the mRNA and protein expression levels of Nanog and Oct4 in the TGF-β group were up-regulated (both P < 0. 05);compared with the TGF-β group,the mRNA and protein expression levels of Nanog and Oct4 in the TGF-β + BAPTA-AM group were down-regulated (both P < 0. 05). Compared with the control group,the in-tracellular calcium increased in the TGF-β group (P < 0. 05);compared with the TGF-β group,the intracellular calcium decreased in the BAPTA-AM group (P < 0. 05). Conclusion TGF-β could enhance the expression of cancer stem genes Oct4 and Nanog in gastric cancer cells by increasing the intracellular calcium concentration.