二甲双胍通过诱导大鼠泌乳素瘤MMQ细胞发生自噬而促进细胞凋亡

Metformin promotes apoptosis of rat prolactinoma MMQ cells by inducing autophagy

目的 :探讨自噬在二甲双胍诱导的垂体泌乳素瘤细胞凋亡中的作用。方法 :用不同浓度的二甲双胍处理泌乳素瘤MMQ细胞不同时间后,CCK-8法检测细胞增殖,FCM法检测细胞凋亡,透射电子显微镜检测细胞中自噬溶酶体的形态及数量变化。用二甲双胍和(或)自噬抑制剂3-甲基腺苷(3-methyladenine,3-MA)处理MMQ细胞后,蛋白质印迹法检测哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,m TOR)-自噬-凋亡信号通路相关蛋白的表达。结果:二甲双胍明显抑制MMQ细胞增殖,并诱导细胞凋亡(P值均<0.05)。二甲双胍能够诱导MMQ细胞自噬,使细胞中的自噬小体数量明显增加(P<0.001);而自噬抑制剂3-MA处理可显著抑制二甲双胍诱导的细胞自噬和凋亡相关蛋白微管相关蛋白轻链3(microtubule-associated protein light chain 3,LC3)-Ⅱ、Beclin 1和聚腺苷酸二磷酸核糖转移酶[poly(ADP-ribose)polymerase,PARP]表达(P值均<0.05)。二甲双胍明显抑制MMQ细胞中mTOR及其下游蛋白P70核糖体蛋白S6激酶(P70 ribosomal protein S6 kinase,P70S6K)和真核细胞翻译起始因子4E结合蛋白(eukaryotic initiation factor 4E binding protein 1,4EBP1)的表达活性(P值均<0.05)。结论:二甲双胍可能通过下调mTOR活性,诱导细胞自噬,从而促进垂体泌乳素瘤细胞发生凋亡。

Objective： To investigate the role of autophagy in metformin-induced apoptosis of pituitary prolactinoma cells. Methods： The different concentrations of metformin was used to treat prolactinoma MMQ cells for different times. CCK-8 assay was used to detect the proliferation of MMQ cells. FCM was used to detect the apoptosis of MMQ cells. The morphology and quantity of autolysosomes in MMQ cells were detected by transmission electron microscopy. After MMQ cells were treated with metformin and/or autophagy inhibitor 3-methyladenine（3-MA）, the expressions of mammalian target of rapamycin（m TOR）-autophagy-apoptotic signaling pathway-related proteins were detected by Western blotting.Results： Metformin significantly inhibited the proliferation of MMQ cells（P〈0.05）, and induced the apoptosis of MMQ cells（P〈0.05）. Metformin induced autophagy, and the number of autophagic bodies in metformin-treated MMQ cells was significantly increased（P〈0.05）. Autophagy inhibitor 3-MA significantly inhibited the expressions of microtubuleassociated protein light chain 3-Ⅱ（LC3-Ⅱ）, Beclin 1 and poly（ADP-ribose） polymerase（PARP） induced by metformin（all P〈0.05）. Metformin also significantly inhibited the expressions of phosphorylated m TOR（p-m TOR） and its downstream proteins phosphorylated P70 ribosomal protein S6 kinase（p-P70 S6 K） and phosphorylated eukaryotic initiation factor 4 E binding protein 1（p-4 EBP1）（all P〈0.05）.Conclusion： Metformin maybe promote the apoptosis of pituitary prolactinoma cells by downregulating m TOR activity and inducing autophagy.