NF-κB抑制剂PDTC逆转K562/AO_2细胞耐药性及其机制的研究

Reversal Effect of Nuclear Factor-κB Protease Inhibitor PDTC on Multidrug Resistance of K562/AO_2 Cells and Its Mechanism

为研究NF-κB的异常活化在K562/AO2细胞耐药机制中的作用,采用非细胞毒剂量抗氧化剂吡咯烷二硫代氨基甲酸盐(PDTC)抑制K562/AO2细胞NF-κB表达,用MTT法检测K562/AO2细胞对药物敏感性的变化,RT-PCR、流式细胞术分别检测K562、K562/AO2细胞mdr-1 mRNA、P-gp表达水平以及不同浓度PDTC作用一定时间对其影响。结果显示:①阿霉素(ADM)诱导耐药的K562/AO2细胞对ADM的耐药性是K562细胞的59倍;非细胞毒剂量PDTC预作用后,ADM对K562/AO2细胞的IC50显著降低,相对逆转耐药效率为93.03%,强于经典耐药逆转剂维拉帕米(Ver);②K562/AO2细胞NF-κB表达水平高于K562细胞(p<0.01);3组非细胞毒剂量PDTC作用后K562/AO2细胞NF-κB表达均受抑制;0.2μmol/LPDTC作用24小时抑制效应最强,K562/AO2细胞NF-κB表达降至接近K562细胞水平(p>0.05);③K562/AO2细胞mdr-1 mRNA、P-gp表达均高于K562细胞(p<0.01);3组非细胞毒剂量PDTC作用K562/AO2细胞48小时后,其mdr-1 mRNA、P-gp表达明显减少。结论:抑制NF-κB异常活化可部分逆转K562/AO2细胞耐药性,其机制与mdr-1 mRNA及其编码的P-gp表达减少有关。

This study was purposed to investigate the relationship between activation of nuclear factor-κB （NF-κB） and multidrug resistance in K562/AO2 cells and its mechanism. Human erythroleukemic cell line K562 and its adriamycin-resistant counterpart K562/AO2 cells were used in the study. After inhibiting the activation of NF-κB with noncytotoxic concentration of antioxidant pyrrolidine dithioearbama （PDTC） in vitro, the multiple of drug resistance of K562/ AO2 cells was assessed by MTT assay. RT-PCR and flow cytometry method were used to detect the relative expression of mdr-1 mRNA and P-gp, respectively. The results showed that （ 1 ） mulfidrug resistance of K562/AO2 cells to ADM was 59 times higher than that of K562 cells. When being pretreated with 0.2 μmol/L PDTC which is noncytotoxic to cells, the IC50 of ADM in K562/AO2 cells was sharply decreased with relative reverse efficiency of 93.03 % , which was more higher than that of classic modifying agents Verapamil （Vet） ; （2）NF-κB activity of K562/AO2 cells was significantly higher than that of K562 cells（p 〈0.01 ）. When being treated with PDTC, the activation of NF-κB was sharply decreased in K562/AO2 cells; with 0.2 μmol/L PDTC for 24 hours it decreased to the lowest, nearly to the K562 cell level（p 〉 0.05 ） ; （3）the relative expression of both mdr-1 mRNA and P-gp in K562/AO2 cells was more higher; the expressions of mdr-1 mRNA and P-gp both were inhibited in K562/AO2 cell group treated with PDTC for 48 hours. It is concluded that the PDTC used as an inhibitor of NF-κB activity can partially reverse the multidrug resistance of 1（562/ AO2 cells, which mechanism can be associated with the down-regulation of mdr-1 mRNA and P-gp.