抑制Survivin缓解小细胞肺癌依托泊苷耐药的功能及机制研究

Inhibition of Survivin Alleviates Etoposide Resistance in Small Cell Lung Cancer and Its Mechanism

目的探究Survivin对小细胞肺癌依托泊苷耐药细胞株增殖和凋亡的影响及机制。方法构建小细胞肺癌细胞NCI-H82和NCI-H446依托泊苷耐药细胞株(NCI-H82/ER和NCI-H446/ER),检测NCI-H82/ER和NCI-H446/ER的IC50及耐药指数,qRT-PCR和蛋白质印迹检测细胞株中Survivin表达。NCI-H82/ER和NCI-H446/ER细胞中转染Survivin siRNA(si-Survivin组)或Survivin siRNA Negative Control(si-NC组),CCK8检测细胞增殖,流式细胞术检测细胞凋亡,蛋白质印迹检测PI3K、AKT、mTOR磷酸化水平。结果NCI-H82/ER和NCI-H446/ER的IC50分别为154.5μmol/L和130.5μmol/L,耐药指数分别为5.40和6.21。相比于NCI-H82和NCI-H446细胞,NCI-H82/ER和NCI-H446/ER细胞中Survivin mRNA和蛋白表达均显著增加(P<0.001),相比于si-NC组,si-Survivin组NCI-H82/ER和NCI-H446/ER细胞增殖能力显著降低(P<0.05),凋亡水平显著增加(P<0.001),PI3K、AKT、mTOR磷酸化均显著降低(P<0.001)。结论抑制Survivin可抑制PI3K/AKT/mTOR信号通路影响依托泊苷耐药小细胞肺癌细胞的增殖和凋亡。

Objective To investigate the effect and mechanism of Survivin on the proliferation and apoptosis of etoposide resistant small cell lung cancer cell lines.Methods Etoposide resistance small cell lung cancer cell lines were constructed in NCI-H82 and NCI-H446 cell lines(NCI-H82/ER and NCI-H446/ER),the IC50 and resistance index of NCI-H82/ER and NCI-H446/ER were determined.qRT PCR and Western blotting assay were used to detect the expression level of Survivin in the cell lines.Survivin siRNA(si-Survivin group)and Survivin siRNA Negative Control(si-NC group)were transfected into NCI-H82/ER and NCI-H446/ER cells.Cell proliferation was determined by CCK8,apoptosis was determined by flow cytometry,and phosphorylation levels of PI3K,AKT,and mTOR were detected by Western blotting.Results The IC50 of NCI-H82/ER and NCIH446/ER were 154.5μmol/L and 130.5μmol/L,respectively,with resistance indices of 5.40 and 6.21.Compared with the NCI-H82 and NCI-H446 cells,the NCI-H82/ER and NCI-H446/ER cells had significantly increased expression levels of Survivin mRNA and protein(P<0.001).Compared with the cells in the si-NC group,NCI-H82/ER and NCI-H446/ER cells in the si-Survivin group had significantly reduced proliferation abilities(P<0.05),and significantly increased level of apoptosis(P<0.001),and significantly reduced phosphorylation levels of PI3K,AKT,and mTOR(P<0.001).Conclusion Inhibiting Survivin can inhibit the PI3K/AKT/mTOR signaling and affect the proliferation and apoptosis of etoposide resistant small cell lung cancer cells.