沉默SIRT1基因表达对胰腺癌细胞PANC1增殖和凋亡的影响  

Effects of silencing SIRT1 gene expression on proliferation and apoptosis of pancreatic cancer cell line PANC1

作　　者：赵刚[1] 崔静[1] 王春友[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院胰腺外科,湖北武汉430022

出　　处：《中华胰腺病杂志》2010年第2期102-105,共4页Chinese Journal of Pancreatology

基　　金：国家自然基金资助项目（30600594）

摘　　要：目的 应用RNA干扰技术沉默胰腺癌PANC1细胞的SIRT1基因表达,观察其对细胞增殖和凋亡的影响.方法 构建靶向SIRT1基因表达的短发夹RNA（shRNA）真核表达质粒pGC-shRNA,转染胰腺癌细胞PANC1.设对照shRNA（shRNA-C）转染组和未转染对照组.实时定量PCR和免疫细胞化学法检测转染后细胞SIRT1 mRNA及蛋白的表达;MTT法检测细胞的增殖率;ELASA法检测细胞caspase-3和caspase-9活性;Western bloting检测细胞Bax、Bcl-2蛋白表达.结果 与未转染组相比,shRNA组转染后48 h,PANC1细胞SIRT1 mRNA及蛋白表达的抑制率分别为（76.2%±10.4）%和（80.1±11.6）%;细胞增殖抑制率为（45.1±6.5）%;caspase-3和caspase-9酶活性显著增高;Bax蛋白表达上调,Bcl-2蛋白表达下调.结论 应用RNA干扰技术能有效沉默PANC1细胞SIRT1基因的表达,其机制可能与caspasa酶活性升高及Bax表达上调、Bcl-2表达下调有关.Objective To investigate the effects of SIRT1 gene expression inhibited by shRNA on proliferation and apoptosis of pancreatic carcinoma PANC1 cells. Methods The eukaryotic expression plasmid of short hairpin RNA （shRNA） targeting SIRT1 gene （SIRT1-shRNA） was constructed as pGC-shRNA and transfected into PANC1 cells. There were shRNA-control transfection group and untransfectian control group. The expressions of SIRT1 mRNA and protein were detected with real-time PCR and immunocytochemistry assay, respectively. The growth rate of PANC1 cells was detected by MTT. Activity of caspase-3 and caspase-9 were detected by ELASA. Expressions of Bax, Bcl-2 proteins were determined by Western blotting. ResultsCompared with untransfected group, the inhibition rate of expressions of SIRT1 mRNA and protein were （76.2 ± 10.4） % and （80.1 ± 11.6） %, cytostasis rate was （45.1 ± 6.5） %, caspase-3 and caspase-9 activity was significantly increased, Bax protein was up-regulated, while Bcl-2 protein was down-regulated 48h after transfection. Conclusions Recombinant expression plasmid SIRT1 shRNA could significantly inhibit the expression of SIRT1 gene, and the mechanism may include increased caspase-3 and caspase-9 activity and Bax protein up-regulation, as well as Bcl-2 down-regulation.

关 键 词：胰腺肿瘤 SIRT1 RNA 小分子干扰 细胞增殖 凋亡 

分 类 号：R735.9[医药卫生—肿瘤]