目的：探讨小片段干扰 RNA（siRNA）干扰 NF － E2相关因子2（NF － E2－ related factor 2，Nrf2）对人喉癌 Hep2细胞化疗敏感性的影响及凋亡的差异。方法体外培养人喉癌细胞系 Hep2，设立实验组和对照组，实验组（Hep2／siRNA 组）采用脂质体转染法，转染 siRNA 至 Hep2细胞系，对照组（Hep2／siRNA － control 组）转染空质粒 siRNA － control 。经 Western Blot 验证其转染效果后，采用 CCK －8法检测计算 Hep2／siRNA 与 Hep2／siRNA － control 经不同浓度梯度顺铂（分别为1、2、4、8、16μg／ml）处理后的细胞增殖抑制率及 IC50值。通过凋亡试剂盒分别染色 Hep2／siRNA 与 Hep2／siRNA － control 细胞系，采用流式细胞仪检测两组细胞的凋亡率。结果实验组的 Nrf2蛋白表达比对照组发生下调，经不同浓度梯度顺铂处理24 h 后，Hep2／siRNA 细胞系增殖抑制率相对 Hep2／siRNA － control 逐渐升高，顺铂浓度为4μg／ml 时，对照组细胞增殖率为35．55％±6．14％，而实验组细胞增殖率为46．07％±5．21％，IC50值下调，细胞凋亡率由17．1％（对照组）升高至26．6％（实验组）。结论 siR‐NA 干扰 Nrf2基因可增强 Hep2细胞系对顺铂的敏感性。%Objective To determine the inhibitory effect of the synthetic Nrf 2 siRNA on the expression of Nrf2 gene in human laryngeal cancer cell lines Hep2 and to investigate the effects of Nrf2 siRNA on chemosensitivity of laryngeal carcinoma to cisplatin by detection growth and apoptosis in Hep2 cells .Methods The recombinant plas‐mid control siRNA and Nrf2 siRNA were transfected into Hep2 cells ,and western blot analysis of Nrf2 expression in Hep2 cells was performed 48 h after transfection .In order to determine whether Nrf2 siRNA can enhance the sensi‐tivity of Hep2 laryngeal cells to cisplatin ,we treated Hep2 cells with different concentrations of cisplatin after 24 h , and evaluated these cells for proliferation ,and apoptosis .CCK - 8 and flow cytometry assay were used for determi‐nation of cells proliferation and apoptosis in Hep2 cells .We calculated the inhibition rate and IC50 of the cell after treating with different concentrations of ciplatin .Results The laryngeal carcinoma cell stain Hep2 was transfected by Nrf2 siRNA and control siRNA respectively .The result of western blot showed the Nrf 2 expression was signifi‐cantly impeded at protein levels .CCK - 8 assay showed the proliferation of Hep2/Nrf2 siRNA and Hep2/ control siRNA was inhibited to 35 .55% to 46 .07% at 24 h respectively after treating with 4 μg/ml cisplatin .The chemo‐sensitivity to cisplatin in Hep2/Nrf2 siRNA was markly increased compared with Hep2/control siRNA .The IC50 in Hep2/Nrf2 siRNA was 5 .27 μg/ml contrast to 8 .107 μg /ml compared in Hep2/control siRNA .The result of flow cytometry analysis showed the apoptosis rate after Nrf 2 depletion was increased from 17 .1% to 26 .6% .Conclusion This study demonstrates that Nrf2 siRNA effectively inhibits Nrf2 gene expression in Hep2 cells leading to growth suppression and induction of apoptosis in Hep2 cells under cisplatin .The use of siRNA technique may pro‐vide a novel therapeutic approach to treat laryngeal cancer for enhance chemosensitivity .
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