新蛋白质合成介导知母活性成分对M_2受体稳定性的调节作用  

ZMS regulation of M_2 muscarinic receptor stability mediated by de novo synthesis of protein

作　　者：张永芳[1] 夏宗勤[1] 胡雅儿[1] 

机构地区：[1]上海交通大学基础医学院细胞调控研究室,上海200025

出　　处：《上海交通大学学报（医学版）》2009年第11期1324-1327,共4页Journal of Shanghai Jiao tong University:Medical Science

基　　金：上海市卫生局基金(054038)~~

摘　　要：目的探讨知母活性成分ZMS提高CHOm2细胞毒蕈碱样乙酰胆碱2型受体(M2受体)mRNA表达的机制。方法体外培养至80%～90%融合的CHOm2细胞分为ZMS1组(单纯添加1×10-5mol/LZMS作用24h)、ZMS2组(添加1×10-5mol/LZMS作用24h后加入1μg/mL环己酰亚胺作用12h)、ZMS3组(加入1μg/mL环己酰亚胺预处理4h后加入1×10-5mol/LZMS作用24h),以上各组分别设立对照组(以等体积DMSO替代ZMS,其他处理与相应ZMS组相同)。各组细胞培养基中均加入放线菌素D抑制mRNA合成,于不同时间点收集CHOm2细胞,Real-time PCR检测M2受体mRNA相对表达量并计算半衰期。结果与相应对照组比较,ZMS1组和ZMS2组CHOm2细胞M2受体mRNA的半衰期明显延长,分别为(4.75±0.54)hvs(2.13±0.23)h和(5.43±1.13)hvs(2.46±0.09)h(均P<0.05);添加1μg/mL环己酰亚胺预处理的ZMS3组CHOm2细胞M2受体mRNA半衰期的(3.06±0.23)h与其相应对照组的(3.00±0.20)h比较,差异无统计学意义(P>0.05)。结论ZMS提高M2受体mRNA的稳定性需要有新蛋白质合成的参与。Objective To explore the mechanism of ZMS regulation of M 2 muscarinic receptor mRNA expression. Methods In vitro cultured CHOm2 cells were divided into ZMS 1 group （treatment with 1×10 -5 mol/L ZMS for 24 h）,ZMS 2 group （treatment with 1×10 -5 mol/L ZMS for 24 h and 1 μg/mL cycloheximide for 12 h） and ZMS 3 group （treatment with 1 μg/mL cycloheximide for 4 h and 1×10 -5 mol/L ZMS for 24 h）,and their corresponding control groups were also established （substitution of ZMS by DMSO）. Actinomycin D was added to cultured CHOm2 cells of each group to inhibit the synthesis of mRNA. CHOm2 cell samples were taken at different time points,the relative expression of M 2 receptor mRNA was detected by Real-time PCR,and half life of M 2 receptor mRNA was calculated. Results Compared with corresponding control groups,the half life of M 2 receptor mRNA of CHOm2 cells in ZMS 1 group and ZMS 2 group was significantly prolonged ／[（4.75h±0.54） h vs （2.13±0.23） h,P 0.05; （5.43±1.13） h vs （2.46±0.09） h,P 0.05／].There was no significant difference in half life of M 2 receptor mRNA of CHOm2 cells between ZMS 3 group and its corresponding control ／[（3.06±0.23） h vs （3.00±0.20） h,P 0.05／]. Conclusion De novo protein synthesis is required for the enhancement of M 2 receptor mRNA stability regulated by ZMS.

关 键 词：知母活性成分 CHOm2细胞 毒蕈碱样乙酰胆碱2型受体mRNA REAL-TIMEPCR 蛋白质合成抑制剂 

分 类 号：Q75[生物学—分子生物学]