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本文探讨了异汉防己碱对P-糖蛋白(P-gp)介导的人乳腺癌细胞多药耐药性的逆转作用。首先以RT-PCR和免疫组化方法分别从RNA和蛋白水平检测MCF-7/DOX细胞P-gp表达情况,以明确MCF-7/DOX细胞的耐药特征;然后采用MTT法检测异汉防己碱的内在细胞毒性及其对阿霉素(DOX)的增敏作用,并以RF(reversal fold)值评价其逆转效果;同时应用流式细胞仪(FCM)对细胞内DOX的蓄积量进行了分析;再以免疫组化方法检测异汉防己碱对MCF-7/DOX细胞P-gp表达水平的影响;最后采用罗丹明蓄积和外排试验检测了异汉防己碱对P-gp功能的影响。整个试验以维拉帕米作为阳性对照。实验结果表明:MCF-7/DOX细胞是具有多药耐药表型且P-gp表达阳性的细胞株;无毒剂量异汉防己碱可明显增强DOX对MCF-7/DOX细胞的细胞毒性(RF=3.89),明显高于维拉帕米(RF=2.54)的逆转活性(P<0.05),但其几乎不影响DOX对MCF-7细胞的抑制作用;异汉防己碱对MCF-7/DOX细胞P-gp表达水平无明显影响,但其可有效抑制P-gp的药物外排功能。因此,异汉防己碱可有效逆转P-gp介导的人乳腺癌细胞的多药耐药性,它可能成为有效多药耐药逆转剂的候选药物。
This article explores the reversal effect of is tetrandrine on P-gp-mediated multidrug resistance in human breast cancer cells. First, the expression of P-gp in MCF-7 / DOX cells was detected by RT-PCR and immunohistochemistry respectively to confirm the drug resistance of MCF-7 / DOX cells; Alkaline cytotoxicity and its sensitization to doxorubicin (DOX), and its reversal effect was evaluated by the value of reversal fold (RF). At the same time, the amount of intracellular DOX accumulation was analyzed by flow cytometry (FCM) The effect of isopycaline on the expression of P-gp in MCF-7 / DOX cells was detected by immunohistochemical method. Finally, the effects of isopyrine on P-gp function were tested by rhodamine accumulation and efflux assay . Verapamil was used as a positive control throughout the trial. The experimental results showed that MCF-7 / DOX cells were multidrug resistance phenotype and P-gp positive cell lines; non-toxic dose of isopyclidine could significantly increase the cytotoxicity of DOX on MCF-7 / DOX cells RF = 3.89) was significantly higher than that of verapamil (RF = 2.54) (P <0.05), but it had little effect on the inhibitory effect of DOX on MCF-7 cells. DOX cells P-gp expression level had no significant effect, but it can effectively inhibit the P-gp drug efflux function. Therefore, different levels of tetrandrine can effectively reverse the multi-drug resistance of P-gp-mediated human breast cancer cells, which may become a candidate drug for effective multidrug resistance reversal agents.