siRNA沉默livin基因对结肠癌细胞5-氟尿嘧啶和奥沙利铂药物敏感性的影响

Effect of siRNA-Mediated Silencing of Livin Gene on Sensitivity to 5-Fluorouracil and Oxaliplatin in Colon Cancer Cells

目的 探讨siRNA沉默livin基因对结肠癌细胞5-氟尿嘧啶(5-fluorouracil,5-Fu)和奥沙利铂(oxaliplatin,L-OHP)敏感性的影响。方法 构建含livin-siRNA序列的稳定表达质粒,通过Lipofectamine TM 2000转染至结肠癌CT-26细胞株,转染后筛选出稳定和低表达livin的结肠癌细胞(livin-siRNA组),同时设空白对照组(mock组)和转染阴性对照组(control siRNA组)。应用反转录-聚合酶链反应(RT-PCR)和Western blot检测CT-26细胞中livin的mRNA和蛋白表达;流式细胞术检测各组CT-26细胞凋亡情况;MTT法检测CT-26细胞对5-Fu和 L-OHP 的敏感性。结果 转染livin-siRNA后,CT-26细胞中的livin mRNA和蛋白量较mock组和control siRNA组明显降低(P<0.05);转染48 h后,livin-siRNA组CT-26细胞凋亡率明显高于mock组和control siRNA组(P<0.05)。转染后livin-siRNA组的CT-26细胞较mock组和control siRNA组对5-Fu和L-OHP的敏感性显著增加(P<0.05)。结论 通过干扰RNA沉默livin基因可促进肿瘤细胞凋亡,增加结肠癌CT-26细胞对5-Fu和 L-OHP 药物的敏感性。

Objective To investigate the effect of siRNA-mediated silencing of livin gene on the sensitivity of 5-fluorouracil(5-Fu) and oxaliplatin(L-OHP) in colon cancer cells.Methods The plasmids carrying livin-siRNA sequence were constructed and transfected into colon cancer CT-26 cells by Lipofectamine TM 2000.The colon cancer cells stably and lowly expressing livin were selected as the livin-siRNA group.In addition,mock-and negative control siRNA-transfected cells were served as mock group and control siRNA group,respectively.The mRNA and protein expression of livin in CT-26 cells was detected by RT-PCR and Western blot,respectively.The apoptosis of cells was measured by flow cytometry.The sensitivity of CT-26 cells to 5-Fu and L-OHP was determined by MTT assay.Results The expression of livin mRNA and protein in livin-siRNA group was lower than that in mock or control siRNA group(P<0.05).At 48 hours after transfection,the apoptosis rate in livin-siRNA group was higher than that in mock or control siRNA group(P<0.05).Furthermore,the sensitivity to 5-Fu and L-OHP in livin-siRNA group was higher than that in mock or control siRNA group(P<0.05).Conclusion The siRNA-mediated silencing of livin gene can promote the apoptosis and increase the sensitivity of 5-Fu and L-OHP in colon cancer CT-26 cells.