γ分泌酶抑制剂阻断Notch1信号通路对人结肠癌耐药细胞化疗敏感性的影响  

Effect of blocking Notch1 signaling pathway by gamma-secretase inhibitor on sensitivity of drug-resistant human colon carcinoma cells to chemotherapy

作　　者：胡敏[1] 李少林[1] 袁犁[1] 王勃[1] 

机构地区：[1]重庆医科大学放射医学教研室,重庆400016

出　　处：《中国生物制品学杂志》2012年第1期23-28,共6页Chinese Journal of Biologicals

摘　　要：目的探讨应用γ分泌酶抑制剂(Gamma-secretase inhibitor,GSI)阻断Notch1信号通路后对人结肠癌耐药细胞株SW480/L-OHP化疗敏感性的影响。方法应用人结肠癌细胞系SW480,以反复诱导、逐渐递增奥沙利铂(L-OHP)浓度的方法,体外构建耐药细胞株SW480/L-OHP。采用CCK-8法检测细胞株对L-OHP的耐药指数,RT-PCR法检测survivin、notch1、hes1、cyclinD1基因mRNA的转录,Western blot检测Notch1和Hes1蛋白的表达。将耐药细胞分成对照组、L-OHP组(3.0μmol/L)、GSI组(10μmol/L)和GSI+L-OHP组,分别处理后,检测各组细胞的抑制率、凋亡率及survivin、notch1、hes1、cyclinD1基因mRNA的转录水平和Notch1、Nicd、Hes1蛋白的表达水平。结果与SW480亲本细胞相比,SW480/L-OHP细胞中survivin、notch1、hes1和cyclinD1基因mRNA转录水平及Notch1和Hes1蛋白表达水平均上调。GSI组耐药细胞的增殖受到抑制,GSI+L-OHP组抑制作用更明显;GSI组耐药细胞survivin、hes1、cyclinD1基因mRNA转录水平下调,Nicd和Hes1蛋白表达水平下调,GSI+L-OHP组下调更明显。结论 GSI通过阻断Notch1胞内段Nicd的释放,影响下游Hes1基因和蛋白的表达,从而抑制SW480/L-OHP细胞增殖,提高其化疗敏感性;且与L-OHP联合使用具有协同作用。Objective To investigate the effect of blocking Notchl signaling pathway by gamma-secretase inhibitor （GSI） on sensitivity of drug-resistant human colon carcinoma SW480/L-OHP cells to chemotherapy. Methods Drug-resistant SW480/L-OHP cell strain was established in vitro by continuous exposure of human colon carcinoma SW480 cells to oxaliplatin （L-OHP） at gradually increasing concentrations, then determined for resistance index to L-OHP by CCK-8 method, for transcr/ptions of survivin, notch1, hesl, cyclinD1 mRNAs by RT-PCR, and for expressions of Notchl and Hesl proteins by Western blot. SW480 / L-OHP cells were divided into four groups and treated with medium containing 1% DMSO （control）, 3. 0 I^mol/L L-OHP, 10 p, mol/L GSI and 10 p.mol/L GSI ＋ 3. 0 ~xmol/L L-OHP respectively, then determined for inhibition rate, apoptosis rate, the transcription levels of survivin, notch1, hesl and cyclinD1 mRNAs and the expression levels of Notchl, Nicd and Hesl proteins. Results Compared with those in SW480 cells, the transcription levels of survivin, notchl, hesl and cyclinD1 mRNAs as well as the expression levels of Notchl and Hesl proteins in SW480/L-OHP cells increased. The proliferation of cells in GSI group was inhibited, while that in GSI ＋ L-OHP group was inhibited ob^i0usly, The transcription levels of survivin, hesl and cyclinD1 mRNAs as well as expression levels of Nicd and Hesl proteins in SW480/L-OHP ceils in GS! group decreased, while those in GSI ＋ L-OHP group decreased obviously. Conclusion GSI inhibited the proliferation level and increased the sensitivity of SW480/L-OHP cells to chemotherapy by blocking the release of Nicd in intracellular domain of Notchl and influencing the expressions of Hesl gene and protein downstream, which showed synergistic action with L-OHP.

关 键 词：结肠肿瘤 抗药性 肿瘤 NOTCH1 γ分泌酶抑制剂 

分 类 号：R735.35[医药卫生—肿瘤]