调控血红素加氧酶-1诱导K562A02细胞增殖、凋亡及耐药机制研究

Study on regulating heme oxygenase-1 for inducing proliferation,apoptosis and drug resistance mechanism of K562A02 cell

目的通过血红素加氧酶-1(HO-1)诱导剂Hemin及抑制剂ZNPP IX调控HO-1,并联合阿霉素逆转K562A02细胞化疗耐药机制的研究,为慢性髓系白血病(CML)的逆转耐药提供新的策略。方法培养K562及K562A02细胞,采用荧光原位杂交(FISH)法检测K562A02细胞中bcr-abl融合基因表达。分别用HO-1诱导剂Hemin及抑制剂ZNPP IX调控HO-1基因表达联合阿霉素处理K562A02细胞后;流式细胞术检测药物诱导细胞凋亡情况。Western blot检测耐药相关基因及凋亡基因蛋白表达水平。结果 K562A02细胞中bcr-abl融合基因阳性细胞占94%。阿霉素处理细胞后,随着阿霉素浓度的增加,HO-1表达下降,耐药相关基因MDR1、NF-κB(P65)、MRP1、TopoⅡα、ABCD2表达亦降低;用HO-1诱导剂Hemin、抑制剂ZNPP IX、阿霉素单药分别及联合处理K562A02细胞后,显示HO-1高表达后耐药相关基因表达升高,细胞凋亡率下降。而降低HO-1表达,耐药相关基因表达下降,细胞凋亡率增加。结论 HO-1可作为逆转耐药的靶基因,可以使K562A02对阿霉素重新敏感,起到增敏效应。

Objective To regulate heme oxygenase-1（HO-1）by the inducer Hemin of HO-1and inhibitor ZNPP.IX and combined with adriamycin for reversing the chemotherapeutic drug-resistance mechanism of K562A02 cells so as to provide a new strategy for chemoresistance reversion of chronic myeloid leukemia（CML）.Methods K562 and K562A02cells were cultured and the expression of bcr-abl fusion gene in K562A02 cells was detected by fluorescence in situ hybridization（FISH）.Then the HO-1gene expression was respectively regulated by Hemin as the HO-1inducer and ZNPP as the inhibitor and adriamycin was combined for treating K562A02 cells with different concentrations.The apoptosis induced by medication was detected by flow cytometry.The expression levels of drug resistance related gene and apoptosis gene protein were detected by Western blot.Results The positive cells of bcr-abl fusion gene in the K562A02 cells accounted for 94%.The HO-1expression was decreased with the adriamycin concentration increase after treating cells by adriamycin,the expression levels of drug resistance related genes MDR1,NF-κB（P65）,MRP1,TopoⅡαand ABCD2 were also decreased;after treating K562A02 cells by single drug and combination of Hemin,ZNPP.IX and adriamycin,increasing HO-1expression elevated the expression of drug resistance related genes and decreased the cellular apoptosis,while reducing HO-1expression could decrease the expression of drug resistance related genes and increased cellular apoptosis.Conclusion HO-1can act as a target gene for drug resistance reversion and can make K562A02 cells to regain sensitivity to adriamycin,thus plays a sensitivity-increasing effect.