大黄素体外逆转口腔鳞癌耐药细胞株KBV200的机制初步探讨

摘要

目的 初步探讨大黄素 (EM) 体外逆转口腔鳞癌耐药细胞株KBV200的机制.方法 用MTT法测定EM对KBV200细胞的逆转作用及量效关系.Western blot法、免疫细胞化学法分别检测单用长春新碱(VCR)和EM联合VCR处理KBV200后,细胞中耐药相关蛋白P-gp、肺耐药相关蛋白 (LRP) 的量,GST-π、TOPO-Ⅱ的活性,以及肿瘤细胞凋亡蛋白P53、Bcl-2/Bax比值的变化.结果 0.7、1.0和1.4 μg/ml 的EM及5.0 μg/ml维拉帕米(VRP)分别联合VCR对KBV200的逆转倍数依次为1.44、2.87、1.76和1.83倍.与单用VCR组相比,EM联合VCR组上调P53的表达 (P<0.01),增强TOPO-Ⅱ的活性 (P<0.01),下调P-gp的表达 (P<0.01);下调LRP的表达量及Bcl-2/Bax比值 (P<0.01).结论 EM有逆转人口腔鳞癌细胞KBV200多药耐药性的作用,其机制可能与提高肿瘤细胞内药物的累积量,促进肿瘤细胞的凋亡有关.%Objective To investigate the reversal mechanism of multidrug resistance (MDR) of human oral squamous carcinoma cells KBV200 by emodin (EM) in vitro. Methods The cytotoxicity of EM in KBV200 cells was detected by MTT assay. The expression of resistance-associated proteins P-gp and LRP, the activity of GST-π, TOPO- Ⅱ, and the tumor cell apoptosis proteins P53 and the ratio value of Bcl-2/Bax were determined by Western blot and immunocytochemistry. Results 0.7, 1.0 and 1.4 μg/ml EM combined with vincristine (VCR) reversed the MDR of KBV200 cells by 1.44, 2.87 and 1.76-fold respectively, while 5.0 μg'ml verapamil combined with VCR only had reversal folds (RF) value by 1.83-fold. EM combined with VCR up-regulated the expression of P53 (P ＜0.01 ) and down-regulated the expression of P-gp ( P ＜0.01 ). The expression of LRP and the ratio of Bcl-2/Bax also decreased ( P ＜ 0.01 ) after treatment with EM combined with VCR. The activity of TOPO- Ⅱ in KBV200 cells obviously increased after treatment with EM combined with VCR (P ＜ 0.01 ). Conclusions EM is potent MDR-reverse agent in vitro. The mechanism is probably associated with increasing intracellular concentration of anti-tumor drugs and promoting apoptosis of KBV200 cells.