中华临床医师杂志（电子版） 2010年8月,4卷8期

论　著

siRNA靶向沉默hTERT基因对人胰腺癌的凋亡抑制基因bcl-2和环氧合酶-2基因表达的影响

钟英强，黄花荣，付玉如，朱兆华

钟英强、黄花荣、朱兆华，广州 中山大学附属第二医院消化内科，510120；付玉如，医学研究中心，510120

摘要:目的　应用siRNA靶向沉默人胰腺癌Capan-2细胞的hTERT基因表达，观察其对凋亡抑制基因(bcl-2)和环氧合酶-2(COX-2)基因表达的影响。方法　应用脂质体Lipofectamine 2000作为基因转染的载体，实时PCR进行扩增检测转染细胞的hTERT、bcl-2、COX-2基因的mRNA表达水平，应用Western blotting检测hTERT、bcl-2、COX-2蛋白表达水平。结果　在hTERT-siRNA完全沉默hTERT表达的基础上，bcl-2、COX-2基因mRNA表达的量在hTERT-siRNA转染后48 h均明显受到抑制(P<0.001)，抑制率分别为87.86%、100.00%，但在72 h后均恢复至与对照组相似的水平，而阴性对照组、Lipo对照组与空白对照组的各基因表达量差异均无统计学意义(P>0.05)。bcl-2、COX-2蛋白水平在转染48 h和72 h后被分别下调了58.54%和63.44%、50.06%和82.77%(P<0.01)，Lipo对照组和阴性对照组与空白对照组差异无统计学意义(P>0.05)。结论　siRNA靶向沉默hTERT基因表达，可在一定程度上抑制人胰腺癌Capan-2细胞的bcl-2、COX-2基因的表达。

关键词:胰腺肿瘤；　基因，bcl-2；　环氧化酶2；　端粒，末端转移酶；　RNA干扰；　脂质体

广东省自然科学基金(7001607)；广东省科技计划项目(2008B030301115)

Effects of silencing expression of hTERT gene by siRNA on the expression of bcl-2 and COX-2 genes of human pancreatic cancer cells in vitro

ZHONG Ying-qiang，HUANG Hua-rong，FU Yu-ru，ZHU Zhao-hua.

Department of Gastroenterology，Second Affiliated Hospital，Sun Yat-sen University，Guangzhou 510120，China

Abstract:Objective　To investiate effects of silencing expression of human telomerase reverse transcriptase (hTERT) gene by siRNA targeting to hTERT gene on the expressions of bcl-2，COX-2 genes of Capan-2 cells of human pancreatic cancer in vitro.Methods　The media of transfection gene was Lipofectamine 2000.The expressions of hTERT mRNA，COX-2 mRNA and bcl-2 mRNA were tested by realtime PCR，and the control was GAPDH.The expressions of hTERT，COX-2 and bcl-2 proteins were tested by Western blotting，and the control was actin.Results　Under the same experiment condition，the inhibited rate of expression of hTERT-mRNA by siRNA was 100%，and the expressions of COX-2，bcl-2 mRNA were inhibited remarkably by transfectation of hTERT-siRNA after 48 hours，and their inhibited rates were 100%,87.86% resprectively(P<0.001)，but they recovered the same level as the control after 72 hours.Meanwhile，the expressions of COX-2，bcl-2 proteins were downregulated 50.06% and 82.77%,58.54% and 63.44% after 48 hours and 72 hours resprectively (P<0.01).But no difference among the controls (P>0.05).Conclusions　Targing silence to hTERT gene by siRNA，it can inhibit the expressions of COX-2，bcl-2 genes of Capan-2 cells of human pancreatic cancer in vitro in some degree.

Keywords:Pancreatic neoplasms；　Genes，bcl-2；　Cyclooxygenase 2；　Telomerase；　RNA interference；　Liposomes

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(编辑：巨娟梅 收稿日期：2010-03-01)