摘要
目的:探讨survivin基因对人胆管癌细胞凋亡信号通路的调节机制。方法:构建针对survivin基因的siRNA和对照siRNA,将QBC939人胆管癌细胞分为3组,分别为siRNA-survivin转染组,对照siRNA转染组和未转染组。转染后,首先用Western blot检测未转染QBC939细胞和QBC939/siRNA(-)细胞及QBC939/siRNA(+)细胞中survivin的表达,验证干扰效果,继而分别用流式细胞仪,激酶活性测定和Western blot检测以上3种状态的QBC939细胞的凋亡情况,caspase-3的活性和caspase-3,caspase-9及procaspase-9凋亡信号分子的表达。结果:QBC939/siRNA(+)细胞survivin蛋白表达量明显低于未转染的QBC939细胞(P<0.05),而QBC939/siRNA(-)细胞survivin蛋白表达量无明显改变(P>0.05)。与未转染的QBC939细胞比较,QBC939/siRNA(+)细胞凋亡明显增加,caspase-3活性明显升高,caspase-3和caspase-9表达明显上调,而procaspase-9表达降低(均P<0.05);上述指标在QBC939/siRNA(-)细胞与未转染的QBC939细胞间的差异无统计学意义(均P>0.05)。结论:survivin基因可能通过促进procaspase-9的活化阻止caspase-3和caspase-9的激活,从而抑制胆管癌细胞的凋亡。
Objective： To investigate the molecular mechanism of survivin regulaion of the apoptosis-related signaling in human cholangiocarcinoma cells.
Methods： The siRNA targeting survivin gene and nontargeting control siRNA were constructed, and humancholangiocarcinoma Q.BC939 cells were divided into three groups that were survivin targeting siRNA transfection group, control siRNA transfection group and untransfection group, respectively. After transfection, the surviving expression in the QBC939 cells, Q.BC939/siRNA （-） and Q BC939/siRNA （＋） cells wasdetermined by Western blot to identify the interference effect, and then, the apoptosis rate, capase-3 activity, and expression of caspase-3, caspase-9 and procaspase-9 in the QBC939 cells of above three statuses were detected by flow-cytometry, capase activity detection kit and Western blot, respectively.
Results： The protein expression level of survivin in the QBC939/siRNA （＋） cells was significantly decreased （P〈0.05）, and there was no obvious alteration in the QBC939/siRNA （-） cells compared with the untransfected Q.BC939 cells （P〉0.05）. Compared with the untransfected QBC939 cells, the QBC939/siRNA （＋） cells presented enhanced apoptosis, increased caspase-3 activity, and upregulated expression level of caspase-3 and caspase-9, but downregulted expression level of procaspase-9 （all P〈0.05）. In each of the above indexes, the differences between the Q.BC939/siRNA （-） cells and untransfected QBC939 cells had no statistical significance （allp 〉 0.05）.
Conclusion： In cholangiocarcinoma cells, apoptosis inhibition by survivin is possibly due to the inactivation of caspase-3 and caspase-9 via procaspase-9 activation.
出处
《中国普通外科杂志》
CAS
CSCD
北大核心
2012年第2期164-168,共5页
China Journal of General Surgery
基金
湖北省武汉市2008年度临床医学科研项目(WX08D04)
