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目的探讨抑制NANOG表达对肝癌细胞HepG2中多药耐药基因1(multidrug resistance gene 1,MDR1)表达及阿霉素敏感性的影响。方法将靶向NANOG基因的特异性siRNA转染肝癌细胞HepG2,Real-time PCR和Western blot检测siRNA沉默NANOG基因后NANOG、MDR1 mRNA和蛋白的表达,平板克隆形成实验检测沉默NANOG基因后对细胞增殖能力的影响,流式细胞术检测沉默NANOG基因后对细胞周期的影响,CCK-8法检测沉默NANOG基因后HepG2细胞对阿霉素的敏感性情况。结果靶向NANOG基因特异性siRNA转染肝癌细胞HepG2后,能有效抑制NANOG mRNA和蛋白表达,与Mock组比值分别为(0.32±0.05)和(0.38±0.08);与Mock组比较,沉默NANOG基因后,细胞克隆形成率下降[(8.51±3.63)%vs(17.13±2.24)%,P<0.05],进入G0/G1期的细胞比例增多[(75.33±8.21)%vs(57.81±5.05)%,P<0.05],HepG2细胞对阿霉素的敏感性增强(P<0.05),HepG2细胞内MDR1 mRNA和蛋白表达下降,与Mock组比值分别为(0.35±0.06)和(0.41±0.08)。结论抑制NANOG表达可引起肝癌细胞HepG2中MDR1的表达下调并增强细胞对阿霉素的敏感性。
Objective To investigate the effect of inhibiting NANOG expression on the expression of multidrug resistance gene 1 (MDR1) and adriamycin sensitivity in HepG2 cells. Methods Specific siRNA targeting NANOG gene was transfected into HepG2 hepatoma cells. The expression of NANOG, MDR1 mRNA and protein was detected by Real-time PCR and Western blot after NANOG gene was silenced. The expression of NANOG, MDR1 mRNA and protein were detected by real-time PCR and Western blot. The effect of silencing NANOG gene on cell cycle was detected by flow cytometry. The susceptibility of HepG2 cells to doxorubicin after silencing NANOG gene was detected by CCK-8 assay. Results The siRNA targeting NANOG gene could effectively inhibit the expression of NANOG mRNA and protein in HepG2 cells after transfection with HepG2, with a ratio of (0.32 ± 0.05) and (0.38 ± 0.08), respectively. Compared with Mock group, NANOG gene silencing (P <0.05). The percentage of cells entering the G0 / G1 phase increased ([(75.33 ± 8.21)% vs (57.81 ± 5.05)%] , P <0.05]. The sensitivity of HepG2 cells to doxorubicin was enhanced (P <0.05). The mRNA and protein expressions of MDR1 in HepG2 cells were decreased compared with Mock group (0.35 ± 0.06) and (0.41 ± 0.08), respectively. Conclusion Inhibition of NANOG expression can down-regulate the expression of MDR1 in HepG2 cells and enhance the sensitivity of cells to doxorubicin.