西黄丸含药血清通过P糖蛋白途径逆转肝癌细胞多药耐药的体外研究

In vitro study on the reversal effect of serum containing Xihuang pill on the multidrug-resistance of hepatic cancer cells via P-glytoprotein pathway

[目的]探讨西黄丸含药血清对多药耐药人肝癌细胞株BEL-7404/Adr细胞和HepG2/Adr细胞耐阿霉素作用的影响及机理。[方法]制备西黄丸含药血清;BEL-7404/Adr细胞和HepG2/Adr细胞分别用西黄丸含药血清、阿霉素及二者联合处理;MTT法检测细胞增殖;Annexin V-FITC/PI双染色流式细胞术检测细胞凋亡;荧光酶标仪测定细胞内阿霉素的荧光强度;Western-blot法检测细胞P-糖蛋白(P-gp)的表达。[结果]西黄丸含药血清联合不同浓度阿霉素处理24h后,BEL-7404/Adr细胞和HepG2/Adr细胞增殖抑制率和凋亡率高于单用同浓度阿霉素组,比较差异有统计学意义(P<0.05);西黄丸含药血清联合不同浓度阿霉素处理12h后,BEL-7404/Adr细胞和HepG2/Adr细胞内阿霉素的荧光强度高于单用同浓度阿霉素组,比较差异有统计学意义(P<0.05);西黄丸含药血清处理12h后,BEL-7404/Adr细胞和HepG2/Adr细胞P-gp蛋白表达水平降低,较对照组差异有统计学意义(P<0.05)。[结论]西黄丸含药血清显著逆转BEL-7404/Adr细胞和HepG2/Adr细胞的耐阿霉素作用,其机制可能与西黄丸含药血清下调细胞多药耐药蛋白P-gp表达相关。

[Objective]To investigate the reversal effects and mechanisms of serum containing Xihuang pill on adriamycin resistance of the multidrug-resistant hepatic cancer cells including the human hepatocellular carcinoma BEL-7404/Adr cell and HepG2/Adr cell.[Methods]Serum containing Xihuang pill was prepared.BEL-7404/Adr cell and HepG2/Adr cell were treated with serum containing Xihuang pill,adriamycin,or their combinations.The proliferation and apoptosis of cells were analyzed by MTT assay and flow cytometry（AnnexinV-FITC/PI double staining assay）.Intracellular accumulation of adriamycin was measured by the detection of fluorescence intensity of cell lysates using microplate reader.The protein expres-sions of P-gp was detected by Western blot.[Results]The growth inhibition rate and apoptosis rate of the BEL-7404/Adr cell and HepG2/Adr cell treated with the combination of serum containing Xihuang pill and adriamycin were higher compared with the cells treated with adriamycin alone（P0.05）.Intracellular accumulation of adriamycin was significantly enhanced and the protein expression of P-gp decreased by the treatment of serum containing Xihuang pill（P0.05）.[Conclusion]Serum containing Xihuang pill reverses the multidrug-resistance of the human hepatocellular carcinoma BEL-7404/Adr cell and HepG2/Adr cell,and the P-glytoprotein pathway may be involved in the effect.