沉默胰腺癌上调因子对胰腺癌细胞株吉西他滨敏感性的影响

摘要

目的 观察沉默胰腺癌上调因子(PAUF)对胰腺癌BxPC-3细胞吉西他滨敏感性的影响.方法 BxPC-3细胞中PAUF基因呈高表达,分别应用PAUF-shCtrl、PAUF-shPAUF转染BxPC-3细胞得到PAUF高低表达不同的对照组和实验组.应用实时聚合酶链式反应(RT-PCR)及蛋白印迹(Western blot)检测进行沉默后验证.MTT比色法检测两株细胞在不同浓度吉西他滨(0、3.1、6.25、12.5、25、50、100、200 nmol/L)作用下对细胞增殖抑制率的影响.流式细胞术检测不同浓度吉西他滨(0、75、100 nmol/L)作用下对细胞凋亡率的影响.结果 BxPC-3_shCtrl细胞及亲本细胞PAUF mRNA的相对表达量为1.00±0.06、0.83 ±0.07,均显著高于BxPC-3_shPAUF细胞的0.25±0.02(均P＜0.05).BxPC-3 shCtrl细胞及亲本细胞PAUF蛋白相对表达量为0.89±0.07、0.95 ±0.04,显著高于BxPC-3_shPAUF细胞的0.31 ±0.03(均P＜0.05).吉西他滨对BxPC-3_shCtrl细胞的半数抑制浓度IC50为(22.88±2.43) nmol/L,显著高于BxPC-3_shPAUF细胞的(1.06 ±0.02) nmol/L(P ＜0.05);BxPC-3_shPAUF细胞的凋亡随着吉西他滨浓度的增加速度明显大于BxPC-3_shCtrl细胞.结论 PAUF沉默后明显增加胰腺癌BxPC-3细胞对吉西他滨的敏感性.%Objective To observe the influence on the sensitivity of pancreatic cancer cell line BxPC-3 to gemcitabine of silencing PAUF gene.Methods BxPC-3 cells,which overexpress PAUF,was stably transfected with PAUF-shCtrl and PAUF-shRNA to establish BxPC-3_shCtrl and BxPC-3_shPAUF cells as control and experiment group.Then the mRNA and protein expression level of PAUF in these two cell lines were detected by RT-PCR and western blot,respectively.The growth inhibition rates of these two cell lines treated with different concentrations of gemcitabine (0,3.1,6.25,12.5,25,50,100,200 nmol/L) were detected by MTT.Apoptosis rates in the cells treated with different concentrations of gemcitabine (0,75,100 nmol/L) were then observed by flow cytometry.Results The relative PAUF mRNA expression level in BxPC-3_shCtrl and BxPC-3 cells were 1.00 ± 0.06 and 0.83 ± 0.07,which were significantly high er than that in BxPC-3_shPAUF cells (0.25 ± 0.02;both P ＜ 0.05).The relative PAUF protein expression level in BxPC-3_shCtrl and BxPC-3 cells were 0.89 ± 0.07 and 0.95 ± 0.04,which were significantly high er than that in BxPC-3_shPAUF cells (0.31 ± 0.03;both P ＜ 0.05).The IC50 value of gemcitabine to BxPC-3_shCtrl cell was (22.88 ± 2.43) nmol/L,which was significantly higher than that of BxPC-3_shPAUF cells [(1.06 ± 0.02) nmol/L;P ＜ 0.05];apoptosis rate of BxPC-3_shPAUF cells treated by gemcitabine increased faster than that of BxPC-3_shCtrl cells.Conclusion PAUF silencing could greatly enhance the sensitivity of BxPC-3 cells to gemcitabine.