内质网应激在卵巢癌顺铂耐药中的作用及其机制

Effect and mechanism of endoplasmic reticulum stress on cisplatin resistance in ovarian carcinoma

目的探讨内质网应激（ERS）在卵巢癌顺铂耐药中的作用及其机制。方法沙奎那韦诱导肿瘤细胞SKOV3后，采用逆转录聚合酶链反应（RT-PCR）法和Westernblot法检测SKOV3细胞中哺乳动物雷帕霉素靶蛋白（mTOR）、Beclin1 mRNA及其蛋白的表达，四甲基偶氮唑蓝（MTT）法检测沙奎那韦对卵巢癌细胞顺铂敏感性的影响。结果顺铂对SKOV3细胞的半数抑制浓度（IC50）为（5．490±1．148）μg/ml。当10、20μmol／L沙奎那韦作用SKOV3细胞后，顺铂的IC50分别为（11．199±0．984）μg／ml和（14．906±2．015）μg/ml，提示卵巢癌细胞对顺铂敏感性下降，差异有统计学意义（P=0．001）。对照组、顺铂组、沙奎那韦＋顺铂组、LY294002组和沙奎那韦组中SKOV3细胞的mTOR、Beclin1 mRNA及其蛋白表达水平不同，差异有统计学意义（均P〈0．001）。mTOR、Beclin1 mRNA相对表达水平在沙奎那韦＋顺铂组中最高，分别为0．684±0．072和0．647±0．047；其次为沙奎那韦组，分别为0．577±0．016和0．565±0．037。mTOR、Beclin1蛋白相对表达水平在沙奎那韦＋顺铂组中最高，分别为0．624±0．058和0．924±0．033；其次为沙奎那韦组，分别为0．544±0．019和0．712±0．024。以3-甲基腺嘌呤抑制肿瘤细胞自噬，可阻断沙奎那韦诱导SKOV3细胞顺铂敏感性下降（P〈0．001）。结论沙奎那韦可有效诱导SKOV3细胞ERS，ERS可导致卵巢癌细胞顺铂敏感性下降，其机制可能与ERS通过调节mTOR和Beclin1表达而改变肿瘤细胞自噬水平有关。肿瘤细胞ERS与细胞自噬可能成为提高肿瘤化疗疗效及逆转耐药的新靶点。

Objective The study intended to investigate the effect and mechanism of endoplasmic reticulum stress on eisplatin resistance in ovarian carcinoma. Methods RT-PCR and Western blot were used to test the expression of roTOR and Beclinl mRNA and protein in ovarian cancer SKOV3 cells after saquinavir induction. MTT assay was used to analyze the influence of saquinavir on cisplatin sensitivity in SKOV3 cells. Results The IC50 of SKOV3 cells was （5.490± 1. 148 ） μg/ml. After induced by Saquinavair 10 μ mol/L and 20 μ mol/L, the IC50 of SKOV3 cells was increased to （ 11. 199 ± 0. 984） μg/ml and （ 14. 906 ± 2. 015 ） μg/ml, respectively. It suggested that the sensitivity of ovarian cancer cells to eisplatin was decreased significantly（ P = 0.001 ）. The expression of mTOR and Beclinl mRNA and protein was significantly different among the five groups： the （Saquinavair ＋ DDP） group of, Saquinavair group, LY294002 group, DDP group and control group（ P 〈 0.001 ）. The expressions of mTOR and Beclinl mRNA were highest in the （ Saquinavair ＋ DDP ） group, 0. 684 ± 0. 072 and 0. 647 ± 0. 047, respectively; Secondly, the Saquinavair group, 0. 577 ±0. 016 and 0. 565 ±0. 037, respectively. The expressions of mTOR and Beclinl proteins were also highest in the （Saquinavair ＋ DDP） group, 0.624 ±0. 058 and 0.924± 0.033, respectively, followed by the Saquinavair group, 0. 544 ± 0. 019 and 0.712 ± 0.024. 3-MA inhibited the autophagy and restored cisplatin sensitivity in the SKOV3 cells after Saquinavir induced ER stress （ P 〈 0. 001 ）. Conclusions Saquinavir can effectively induce endoplasmic reticulum stress in SKOV3 ceils. Endoplasmie reticulum stress can decrease the sensitivity to cisplatin in SKOV3 cells. The mechanism of the decrease of sensitivity to cisplatin in SKOV3 cells may be that ERS regulates cell autophagy through themTOR and Beclinl pathways. ERS of tumor cells and autophagy may become a new target to improve the therapeutic effect of chemotherapy and to reverse the drug resistance in tumor treatment.