人参皂苷Rd通过下调Nrf2表达调控H460细胞的增殖和凋亡

Ginsenoside Rd Regulates Proliferation and Apoptosis of H460 Cells by Suppressing Nrf2

核转录因子红细胞系-2p45相关因子-2(nuclear factor erythroid-2p45-related factor 2,Nrf2)是细胞应对外界应激的主要调控因子,通过调控一系列细胞保护酶,维持细胞稳态。然而,在许多肿瘤细胞中Nrf2过度激活,导致肿瘤细胞获得增殖优势并产生化疗耐药,因此,靶向抑制Nrf2是肿瘤增敏治疗的一种新思路。人参皂苷Rd是人参皂苷中的重要活性成分,具有显著的抗肿瘤作用。本研究以不同浓度的人参皂苷Rd处理人非小细胞肺癌(non-small cell lung cancer,NSCLC)H460细胞,利用CCK-8法检测细胞活力;倒置显微镜观察H460细胞的形态变化;流式细胞术检测细胞周期及凋亡率;RT-qPCR和Western印迹检测的Nrf2及其下游调控基因的表达情况;此外通过转染Nrf2-siRNA下调H460细胞中Nrf2的表达,观察其对人参皂苷Rd增敏的影响。结果显示,与对照组相比,人参皂苷Rd能够抑制H460细胞增殖活力,诱导细胞G0/G1期阻滞,促进细胞凋亡,具有剂量依赖性(P<0.05);此外,人参皂苷Rd能够下调Nrf2,醌氧化还原酶1[NAD(P)H:quinoneoxidoreductase,NQO1],谷氨酰半胱氨酸连接酶催化亚基(glutamate--cysteine ligase catalytic subunit,GCLC)和调节亚基(glutamate--cysteine ligase regulatory subunit,GCLM)的mRNA和蛋白质水平,同时增强H460细胞对化疗药的敏感性(P<0.05),而转染Nrf2-siRNA后,人参皂苷Rd的增敏作用减弱。表明人参皂苷Rd可以抑制非小细胞肺癌H460细胞活性并增敏化疗,其机制可能是通过抑制Nrf2信号通路来实现。

Nuclear factor erythroid 2-related factor 2(Nrf2)is a critical intracellular regulator of the adaptive response to environmental stresses and activates an array of genes involved in cytoprotective enzymes and responsible for cellular homeostasis.Nevertheless,in many types of cancer,the high constitutive expression of Nrf2 leads to proliferation and chemoresistance.Therefore,inhibition of Nrf2 may be an effective strategy for overcoming drug resistance.Ginsenoside Rd is one of the principal active constituents of ginsenosides,and it has been proposed to be a potent anti-tumor agent.Here,we investigated the effects of ginsenoside Rd on the Nrf2 signaling pathway in non-small cell lung cancer(NSCLC)cells.H460 cells were treated with different concentrations of ginsenoside Rd,a cell counting kit-8 assay was used to determine relative cell viability,and flow cytometry was used to evaluate cell cycle and apoptosis;RT-qPCR and Western blotting analysis were performed to measure the expression of Nrf2 and its target genes.RNA interference was used to down-regulate the expression of Nrf2 and determine the role of Nrf2 signaling in H460 cells.We found that ginsenoside Rd inhibited the proliferation of H460 cells.It also induced G0/G1 phase arrest and apoptosis.Ginsenoside Rd also inhibited the mRNA and protein expression of nuinoneoxidoreductase-1(NQO1),glutamate-cysteine ligase catalytic subunit(GCLC)and glutamate--cysteine ligase regulatory subunit(GCLM)and significantly sensitized H460 cells to therapeutic drugs,whereas transfection of Nrf2-siRNA could attenuate the synergistic effects.We conclude that ginsenoside Rd inhibits the cell viability and promotes sensitization of H460 cells,which may take place through inhibition of the Nrf2 pathway.