NAC-1特异的siRNA增强紫杉醇诱导的人卵巢癌细胞凋亡

目的: 探讨 NAC-1 (nucleus accumbens-1)基因特异的小干扰RNA(small interfering RNA,siRNA)联合紫杉醇诱导人卵巢癌HO8910细胞凋亡的协同作用及其可能机制。方法: 测定不同浓度 NAC-1 基因特异的siRNA与紫杉醇单用或者合用对HO8910细胞增殖与凋亡的作用,设立阴性siRNA对照和空白对照。以实时荧光定量RT-PCR 检测NAC-1 mRNA表达水平、Western印迹法检测NAC-1蛋白、表皮生长因子受体(EGFR)下游信号磷酸化Akt(p-Akt)和磷酸化细胞外信号调节激酶(p-ERK),以MTT法检测细胞增殖率,流式细胞术检测细胞周期分布和凋亡。结果: 单用 NAC-1 基因特异的siRNA作用48 h能抑制HO8910细胞NAC-1 mRNA和蛋白的表达,分别下调71.2%和80.5%,细胞周期被阻滞于G₁期,p-Akt和p-ERK蛋白水平分别下降43.7%和49.8%;作用72 h细胞增殖被抑制45.6%,与转染阴性siRNA 及未转染组比较,差异显著(P<0.05)。 NAC-1 siRNA(0.5 μmol/L)与紫杉醇(2 μmol/L)联合作用于卵巢癌HO8910细胞72 h细胞凋亡率为(30.93±4.57)%,显著高于单用紫杉醇时的细胞凋亡率(23.85±3.65)%],P<0.05。结论: 靶向抑制 NAC-1 基因表达能显著抑制卵巢癌HO8910细胞增殖,增强其对紫杉醇的敏感性,这与部分抑制EGFR下游信号途径有关。

NAC -1-specific siRNA enhances paelitaxel-induced apoptosis of ovarian cancer cell line HO8910

ABM:To observe the effect of NAC -1-specific siRNA alone,or in combination with paelitaxel on proliferation and apoptosis of human ovarian cancer cell line HO8910.METHODS:Ovarian cancer cells were treated with NAC -1 siRNA alone or in combination with paelitaxel.The level of NAC-1 mRNA was assessed by real-time quantitative PCR.Western blotting analysis was used to detect NAC-1 protein and the activation of epidermal growth factor receptor（EGFR） downstream signals,Akt and ERK.The cell proliferation rate was measured by MTT assay,and the cell cycle and apoptosis were determined by flow cytometry.RESULTS:After treated with NAC -1-specific siRNA for 48 h,the expression of NAC-1 at mRNA and protein levels in HO8910 cells decreased by 71.1%and 80.5%,respectively. The cells in G₁ phase increased.The protein levels of p-Akt and p-ERK were decreased by 43.7%and 49.8%, respectively.After treated with NAC-1-specific siRNA for 72 h,the proliferation inhibitory rate of the cells was increased to 45.6%as compared with the cells treated with negative siRNA.Apoptotic rate of the cells treated with NAC-1 siRNA（0.5μmol/L combined with 2μmol/L of paelitaxel） for 72 h was（30.93±4.57）%,higher than that of the cells treated with paelitaxel alone（23.85±3.65）%].CONCLUSION:NAC -1 siRNA suppresses NAC -1 gene expression and EGFR downstream signaling activation,inhibits cell proliferation and enhances the responsiveness of ovarian cancer cells to paelitaxel.The combination treatment produces synergistic inhibition.