R-(α)-甲基组胺对AtT20垂体瘤细胞Pomc及其转录因子表达的影响  

Effect of R-(α)-MeHA on Expression of Pomc and Transcription Factors in Pituitary Tumor AtT20 Cells

作　　者：王晓敏[1] 潘志强[1] 梁龙龙[1] 方肇勤[1] 卢文丽[1] 刘小美[1] 

机构地区：[1]上海中医药大学基础医学院,上海201203

出　　处：《中国肿瘤》2017年第8期650-655,共6页China Cancer

基　　金：上海市进一步加快中医药事业发展三年行动计划(2014年-2016年)(ZY3-CCCX-3-3010);上海市健康辨识与评估重点实验室建设项目(13DZ2261000)

摘　　要：[目的]研究R-(α)-甲基组胺[R-(α)-MeHA]对AtT20垂体瘤细胞Pomc及其转录因子表达的影响。[方法]以小鼠垂体瘤细胞(AtT20)为实验对象,分别采用0.02~12.5μmol/L R-(α)-MeHA干预AtT20细胞24h,通过MTT检测药物对细胞增殖的影响;采用0.1μmol/L R-(α)-MeHA干预AtT20细胞30min、1、2、4、8、12h等相应时间,运用ELISA方法检测细胞分泌至培养液的促肾上腺皮质激素(ACTH)含量,RT-q PCR方法检测相关基因mRNA表达,Western blot方法检测相关蛋白表达。[结果]0.02~12.5μmol/L的R-(α)-MeHA对AtT20细胞的增殖无抑制作用更无毒性作用。0.1μmol/L R-(α)-MeHA对AtT20细胞ACTH分泌功能无明显作用。与对照组比较,0.1μmol/L R-(α)-MeHA干预AtT20细胞4h后,可增强阿片促黑素皮质素原(Pomc)基因表达、0.5μmol/L R-(α)-MeHA促进Pomc蛋白表达;此外0.1μmol/L R-(α)-MeHA干预AtT20细胞30min^12h不同时点后Pou1f1基因表达均显著上调,其中以4h最为明显,上调达3倍(P<0.05);同样,0.1μmol/L R-(α)-MeHA作用4h也显著促进Egr1基因表达上调约2倍(P<0.05);作用1h显著增强了Prop1基因表达,上调近4倍(P<0.05);干预4h、8h均显著上调Foxl2基因表达,上调约3倍(P<0.05),对Neurod1、Pitx1和Tbx19基因表达无明显影响。[结论]R-(α)-MeHA具有促进Pomc基因以及其转录因子Pou1f1、Egr1、Prop1和Foxl2的表达作用。[Purpose] To study the effect of R-（α）-MeHA on the expression of Pomc and transcription factors in mouse pituitary tumor AtT20 cells. [Methods] Mouse pituitary tumor AtT20 cells were treated with 0.02 -12.5μmol/L R-（α）-MeHA for 24 h,and cell proliferation was detected by MTT assay. Then,AtT20 cells were treated with 0.1μmol/L R-（α）-MeHA for 30 min,1,2,4,8,12 h,and cell culture supernatants were collected and ACTH were tested by ELISA kit. The cells total RNA was extracted using TRIzol kit and was reverse transcribed to obtain the c DNA,which was amplificated by real time quantitative PCR（RT-q PCR）. The cells were lysed with RIPA lysis buffer and protein expression was detected by Western blot. [Results] Compared with the control group,the proliferation of AtT20 cells was not affected by 0.02-12.5μmol/L R-（α）-MeHA and ACTH level was slight changed by 0.1μmol/L R-（α）-MeHA. However,R-（α）-MeHA promoted pomc mRNA and POMC protein expression after treatment for 4h. Pou1f1 and Egr1 mRNA expression was significantly increased after 0.1μmol/L R-（α）-MeHA treatment for 4h（P〈0.05）; the Prop1 mRNA expression was significantly increased after treatment for 1h（P〈0.05）; and Foxl2 mRNA expression was increased significantly after treatment for 4h and 8h（P〈0.05）. However,Neurod1,Pitx1 and Tbx19 mRNA expression were not affected by R-（α）-MeHA treatment. [Conclusion] Pomc and its transcription factors Pou1f1,Egr1,Prop1 and Foxl2 gene expressions are regulated by R-（α）-MeHA in mouse pituitary tumor AtT20 cells.

关 键 词：AtT20垂体瘤细胞 R-(α)-甲基组胺 基因表达 POMC Acth POU1F1 

分 类 号：R739[医药卫生—肿瘤]