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利用糖原合成酶激酶3的抑制剂氯化锂作用于A549细胞,观察细胞形态与增殖的改变及其对Polo样激酶1(Plk1)转录活性的影响.采用细胞计数检测细胞增殖,流式细胞术分析细胞周期变化;Western印迹检测磷酸化GSK3以及细胞周期相关蛋白p53、细胞周期蛋白B1和Plk1的表达变化;RT-PCR检测Plk1mRNA的表达;荧光素酶报告基因分析氯化锂对Plk1启动子活性的影响.结果显示,5mmol/L氯化锂作用48h后,A549细胞即发生明显的形态学改变,细胞增殖减慢并发生G2/M期阻滞;Plk1mRNA和蛋白表达均升高,p53蛋白表达增强,而细胞周期蛋白B1的蛋白表达无明显变化.氯化锂作用24h后,可见pGL2-Plk1转染组中荧光素酶活性增高(与对照质粒相比,P<0.05),48h后更明显.以上结果表明,氯化锂减慢A549细胞增殖,导致G2/M期阻滞,并能增强Plk1启动子活性,促进Plk1的表达.
The effect of lithium chloride, an inhibitor of glycogen synthase kinase 3, on A549 cells was observed in order to observe the changes of cell morphology and proliferation and its effect on Polk-like kinase 1 (Plk1) transcription.Cell proliferation was measured by flow cytometry The expression of phosphorylated GSK3 and p53, cyclin B1 and Plk1 were detected by Western blotting. The expression of Plk1 mRNA was detected by RT-PCR. The luciferase reporter gene was used to analyze the effect of lithium chloride on Plk1 promoter The results showed that A549 cells underwent morphological changes after 48h treatment with 5mmol / L lithium chloride, the cell proliferation slowed down and G2 / M phase arrest occurred, the expression of Plk1 mRNA and protein were increased, and p53 protein (P <0.05). After 48 hours, the luciferase activity of pGL2-Plk1 transfection group was increased (P <0.05 compared with the control plasmid), and after 48 hours, the expression of cyclin B1 was increased The above results show that lithium chloride slows the proliferation of A549 cells, leading to G2 / M arrest, Plk1 promoter activity and Plk1 expression.