雷帕霉素联合顺铂对宫颈癌Hela细胞生长和对HIF-1α与VEGF表达影响的观察

Effect of rapamycin combined with cisplatin on the growth of Hela cells in vitro and the expression of HIF-1α and VEGF

目的:探讨mTOR抑制剂雷帕霉素与顺铂联合应用对宫颈癌Hela细胞生长的抑制作用及对HIF-1α和VEGF表达的影响。方法:分别以雷帕霉素、顺铂及雷帕霉素联合顺铂处理体外培养人宫颈癌Hela细胞,采用MTT比色法检测人宫颈癌Hela细胞的抑制率,金氏公式评价两药联合用药的效果,采用RT-PCR法、蛋白质印迹法检测各组细胞HIF-1α和VEGF mRNA和蛋白的表达。结果:雷帕霉素和顺铂对宫颈癌Hela细胞的生长有明显的抑制作用,并呈剂量依赖性,分别联合应用2种浓度雷帕霉素(10和20nmol/mL)与顺铂(0.25和0.5mg/mL)协同治疗指数q值均>1.15,两者有协同作用。雷帕霉素与顺铂联合用药HIF-1αmRNA的表达为0.242±0.048,单独用药雷帕霉素组为0.428±0.068,顺铂组为0.357±0.051;雷帕霉素与顺铂联合用药组VEGF mRNA的表达为0.498±0.093,单独用药雷帕霉素组为0.651±0.112,顺铂组为0.623±0.125,联合用药组与单独用药组比较表达量明显降低,P<0.05;雷帕霉素与顺铂联合用药组HIF-1α蛋白的表达为0.514±0.092,单独用药雷帕霉素组为0.625±0.132,顺铂组为0.635±0.120;雷帕霉素与顺铂联合用药组VEGF蛋白的表达为0.409±0.082,单独用药雷帕霉素组为0.650±0.114,顺铂组为0.623±0.102,联合用药组与单独用药组比较表达量明显降低,P<0.05。结论:单独及联合应用雷帕霉素与顺铂对宫颈癌Hela细胞的生长均有抑制作用,两药联合应用有明显的协同作用,雷帕霉素联合顺铂具有明显下调HIF-1α、VEGF基因和蛋白表达的作用。

OBJECTIVE： To eximine the effect of rapamycin combined with cisplatin on the inhibition of Hela cells in vitro and the expression of HIF-la and VEGF. METHODS：Cervical carcinoma Hela cell were treated by rapamycin,cispla- tin,rapamycin and cisplatin respectively. The inhibitory effects of chemotherapy on the proliferation of Hela cell were measured with MTT assay,and the combined effect of rapamycin and cisplatin were evaluated using King＇s formula. The expressions of mRNA and the protein of the HIF-la and VEGF were detected using transcription-polymerase chain reaction（RT-PCR）and Western Blot. RESULTS： Both rapamycin and cisplatin showed inhibitory effect on proliferation of Hela cells and with dose-dependence in two concentrations of rapamycin （10 and 20 nmol/mL） and eisplatin （0. 25 and 0.5 mg/mL） respectively. The synergistic therapeutic index qvalues were both more than 1.15,which showed synergistic effect. The expressions mRNA of HIF-1 a and VEGF of each group respectively was rapamycin combination with cisplatin （0.242±0.048,0.498±0. 093） ,rapamycin （0. 428±0. 068,0. 651± 0. 112）, cisplatin（0. 3572=0. 051,0. 623±0. 125）. The expressions of mRNA of HIF-la and VEGF were significantly downregulated in the combined group （P〈0.05） ; The expression of protein of HIF-1 a and VEGF mRNA of each group respectively was Rapamycin and cisplatin combination（0. 514±0. 092,0. 409±0. 082） ,rapamycin（0. 625±0. 132,0. 650±0. 114） ,cisplatin（0. 635±0. 120,0. 623±0. 102）. The expressions of protein of HIF-la and VEGF were significantly downregulated in the combined group （P〈0.05）. CONCLU- SION.. Both rapamycin alone or combined use of rapamycin and cisplatin showed inhibitory effect on the proliferation of Hela cells, the combined use of rapamyein and the cisplatin showed synergistic effects in the chemotherapy of Hela cell, The expressions of mRNA and protein HIF-la and VEGF were significantly downregulated by the combined group.