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-α 2.5×106 U·L-1、ATRA 7.5 μmol·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。
Abstract
OBJECTIVE To investigate the effects of interferon-α(IFN-α) and all-trans retinoic acid(ATRA) on multidrug resistance reversal effect and mechanism of human leukemia K562/ADM cells. METHODS The cytotoxicity and reversal times of IFN-α and ATRA were detected by CCK-8 method. Apoptosis rate and cell cycle were detected by flow cytometry. PI3K, Akt and Bad mRNA were detected by RT-PCR method. Western blot method was used to detect the expression of PI3K, AKt, P-AKt and Bad protein.RESULTS The drug resistance of K562/ADM cells to adriamycin(ADM) was 54 times. ADM, respectively, with IFN-α, ATRA or combined application, the drug resistance of K562/ADM cells to ADM was 1.24, 2.34 and 8.14, respectively. The apoptosis rate of K562/ADM cells was significantly increased by using ADM 4 mg·L-1alone or in combination with IFN-α 2.5×106 U·L-1, ATRA 7.5 μmol·L-1, and the cell cycle was blocked in G0/G1 phase. PI3K mRNA and protein expression were significantly lowered, Akt mRNA and protein has no obvious change, Bad mRNA and protein expression are raised, phosphorylated Akt protein expression decreased, the expression is more obvious when the two drug combination. CONCLUSION IFN-α and ATRA can reverse the multidrug resistance of K562/ADM cells, its mechanism may be the inhibition of the PI3K/Akt pathway.
关键词
α-干扰素 /
全反式维甲酸 /
多药耐药 /
K562/ADM /
逆转
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Key words
interferon-α /
all-trans retinoic acid /
multidrug resistance /
K562/ADM /
reversal
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参考文献
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脚注
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基金
四川省科技厅应用基础研究(川科发计【14JC0193】)
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