吉西他滨对Beclinl基因沉默的人胰腺癌MiaPaCa-2细胞的周期及凋亡的影响

Effects of gemcitabine on the ceil cycle and apoptosis of pancreatic cancer cells MiaPaCa-2 with Beclinl gene silencing

目的 探讨吉西他滨对Beclinl基因沉默的人胰腺癌MiaPaCa-2细胞周期及凋亡的影响.方法 构建靶向Beclinl的siRNA,插入表达质粒,转染MiaPaCa-2细胞.采用RT-PCR法和蛋白质印迹法检测细胞Beclinl mRNA及蛋白的表达,应用吉西他滨处理Beclinl基因沉默的MiaPaCa-2细胞,应用流式细胞仪检测细胞周期及细胞凋亡状况.结果 成功获得Beclin1基因沉默的MiaPaCa-2细胞,转染后细胞的Beclin1 mRNA表达量从对照组的1.0下降到0.295;S、G2期细胞数减少,而G1期细胞增多;细胞凋亡未受影响.应用吉西他滨处理后,Beclin1基因沉默的MiaPaCa-2细胞的S期细胞数进一步减少,而G1、G2期细胞增多,细胞凋亡被抑制.结论 Beclinl表达沉默使人胰腺癌细胞系MiaPaCa-2细胞周期发生改变,并影响吉西他滨对细胞周期及凋亡的作用.

Objective To investigate the effect of gemcitabine on the cell cycle and apoptosis of the human pancreatic cancer cell line MiaPaCa-2 with Beclinl gene silencing. Methods siRNA targeting at Beclinl gene was constructed, then it was inserted into an expression vector and transfected into MiaPaCa-2 cells. The Beclinl mRNA and protein expression were detected by RT-PCR and Western blot. Gemcitabine was used to treat MiaPaCa-2 with Beclinl gene silencing, then the cell cycle and apoptosis were detected by flow cytometry. Results The MiaPaCa-2 cells with Beclinl gene silencing were successfully constructed, and the expression of Beclinl mRNA was decreased from 1.0 in control group to 0.295, and number of cells in S and G2 phase was decreased, but number of cells in Gl phase was increased, and there was no change in apoptosis. After gemcitabine treatment, number of cells in S phase was further decreased, but number of ceils in G1 , G2 phase was increased, and apoptosis was inhibited. Conclusions Beclinl gene silencing can change the cell cycle of pancreatic cancer cells MiaPaCa-2, and influence the effects of gemcitabine on cell cycle and apoptosis.