摘要
目的探讨雷帕霉素衍生物依维莫司（RAD001）或联合组蛋白去乙酰化酶抑制剂LBH589对耐药急性髓系白血病细胞株HL-60／ADM细胞增殖、凋亡和逆转耐药的影响。方法采用不同浓度RAD001或联合LBH589处理HL-60／ADM细胞，四甲基偶氮唑盐（MTT）法检测细胞增殖活力，Hoechst33342染色法和AnnexinV—FITC／PI染色流式细胞仪检测细胞凋亡、阿霉素摄取率和多药耐药相关蛋白1（MRP1）表达评估逆转耐药效应。进一步检测处理前后细胞信号通路蛋白变化，探讨其机制。结果10～50μmol／LRAD001均能够抑制HL-60／ADM细胞增殖、诱导凋亡，在作用48和72h时30μmol／L药物浓度达到最大抑制效果，进一步增加药物浓度细胞增殖抑制率并没有明显增加（P〈0．05）。不同浓度RAD001和LBH589联合处理HL-60／ADM细胞，没有明显的协同抑制增殖作用[协同指数（CI）≥1．0]。10μmol／LRAD001处理HL-60／ADM细胞可以明显下调细胞表面MRP1表达（93．90％±4．20％比79．10％±3．28％，P〈0．05），提高HL-60／ADM细胞阿霉素蓄积率（8．53％±0．68％比15．37％±1．46％，P〈0．01）。深入机制研究表明，RAD001可以阻断P13K／AKT／mTOR信号通路活性，通过上调1）53抑制MRPI蛋白的活性。结论RAD001与LBH589联合没有协同抑制HL-60／ADM细胞增殖作用。但RAD001单药能够有效抑制HL-60／ADM细胞增殖，诱导凋亡，通过阻断P13K／AKT／mTOR信号通路抑制细胞MRP1蛋白的表达，提高细胞阿霉素摄取率而逆转耐药。
Objective To investigate the effects of everolimus （RAD001） or plus panobinostat （LBH589） on the proliferation, apoptosis and drug resistance in chemoresistant acute myeloid leukemic cells. Methods HL-60/ADM cells were treated with RAD001 alone or with LBH589. Proliferation and apoptosis were evaluated by 3-（ 4,5 ） -dimethylthiahiazo （ -z-yl ） -3,5-di-phenytetrazoliumromide （ MT＇F ） assay, Hoechst33342 and AnnexinV-FITC/PI stain. The altered expressions of multidrug resistanceassociated protein 1 （ MRP1 ） and intercellular adfiamycin accumulation were analyzed by flow cytometry. The change in protein level was analyzed by Western blot. Results Effective proliferative inhibition and apoptotic induction in HL60/ADM cells were observed in the treatment of 10 -50 μmol/L RAD001. The maximal effect was shown for the concentration of 30 μmol/L RAD001 at 48 and 72 hours. The inhibition ratio remained unchanged with the adjustment of drug doses （ P 〈 0. 05 ）. Moreover, there was no synergistic effects in the treatment with different concentration of RAD001 and LBH589 （CI ≥ 1.0）. A down-regulation of MRP1 （93.9% ± 4. 2% vs 79. 10% ± 3.28% ） and an up-regulation of adriamycin （ 8.53 % ± 0. 68% vs 15.37%± 1.46% ） were induced by the treatment with 10 μmol/L RAIX）01 （ both P 〈 0. 01 ）. RAD001 inhibited the p53-dependent expression of MRP1 via an inhibition of phosphoinositide 3-kinase （PI3K）/ AKT/mTOR signaling pathway. Conclusion The combined treatment of RAD001 and LBH589 has no synergistically inhibitory effect on HL60/ADM cells. But the sole treatment of RADO01 may inhibit proliferation, induce apoptosis and accumulate intercellular adriamycin through a down-regulated expression of MRP1 in HL60/ADM cells via an inhibition of PI3K/AKT/mTOR signaling pathway.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2011年第32期2287-2292,共6页
National Medical Journal of China
