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目的探讨α-干扰素(interferon-α,IFN-α)和全反式维甲酸(all-trans retinoic acid,ATRA)联合作用对耐药白血病细胞K562/ADM耐药性逆转效应及其作用机制。方法采用CCK-8法检测逆转剂IFN-α和ATRA的细胞毒性及药物作用后的逆转倍数;流式细胞术检测细胞凋亡率及细胞周期;RT-PCR法检测细胞PI3K/Akt通路中PI3K、Akt、Bad基因的表达;Western blot法检测PI3K、Akt、磷酸化Akt(p-Akt)、Bad蛋白的表达。结果 K562/ADM细胞对多柔比星(adriamycin,ADM)的耐药率达54倍,ADM分别与IFN-α、ATRA或联合应用时可逆转K562/ADM细胞对ADM的耐药倍数分别为1.24、2.34、8.14;应用ADM 4 mg·L~(-1)单用或联合IFN-α2.5×106U·L~(-1)、ATRA 7.5μmol·L~(-1)(无毒性浓度)均可使K562/ADM细胞的凋亡率明显增加,细胞周期被阻滞在G0/G1期;PI3K基因及蛋白表达均明显下调,Akt基因及蛋白无明显变化,Bad基因及蛋白表达均上调,p-Akt蛋白表达下降,当两药联用时上述表达更明显。结论 IFN-α联合ATRA能逆转K562/ADM细胞多药耐药,其机制可能是抑制了PI3K/Akt通路的作用。
Objective To investigate the reversal effect of multidrug resistance of interferon-α (IFN-α) and all-trans retinoic acid (ATRA) on drug resistance of K562 / ADM cells and its mechanism. Methods CCK-8 method was used to test the cytotoxicity of reversal agents IFN-α and ATRA and the reversal times after drug treatment. Flow cytometry was used to detect the apoptosis rate and cell cycle. The PI3K / Akt pathway in PI3K / Akt pathway , Akt, Bad gene expression; Western blot was used to detect the expression of PI3K, Akt, phosphorylated Akt (P-Akt), Bad protein. Results The drug resistance rate of adriamycin (ADM) in K562 / ADM cells was 54-fold. Compared with IFN-α and ATRA, the drug resistance rates of K562 / ADM cells reversible to ADM were 1.24 , 2.34,8.14, respectively. ADM 4 mg · L -1 alone or combined with IFN-α 2.5 × 106 U · L -1, ATRA 7.5 μmol·L -1 (non-toxic) The apoptosis rate of K562 / ADM cells was significantly increased and the cell cycle was arrested at G0 / G1 phase. The expression of PI3K gene and protein were significantly down-regulated while the expression of Akt gene and protein was unchanged. The expression of Bad gene and protein were up- p-Akt protein expression decreased, when the combination of the two drugs when the above expression is more obvious. Conclusions IFN-α combined with ATRA can reverse the multidrug resistance of K562 / ADM cells. The mechanism may be that the PI3K / Akt pathway is inhibited.