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阿霉素诱导下多发性骨髓瘤细胞中自噬和氧化应激的相互作用
引用本文:罗綦,陈建斌.阿霉素诱导下多发性骨髓瘤细胞中自噬和氧化应激的相互作用[J].中国病理生理杂志,2016,32(4):665-670.
作者姓名:罗綦  陈建斌
作者单位:重庆医科大学附属第一医院血液科, 重庆 400016
基金项目:重庆市卫生计生委医学科研计划项目(No.2011-1-032)
摘    要: 目的: 探讨阿霉素(doxorubicin,DOX)诱导对多发性骨髓瘤细胞系RPMI-8226细胞内自噬和活性氧簇(reactive oxidative species,ROS)生成的影响及其相互作用关系。方法: 不同浓度阿霉素诱导RPMI-8226细胞24 h,采用Western blot技术检测细胞内beclin 1、LC3等自噬相关蛋白的表达水平。采用DCFH-DA荧光染色法检测RPMI-8226细胞内ROS的水平,荧光显微镜采集图像。采用氧自由基清除剂N-乙酰半胱氨酸(N-acetyl-L-cysteine,NAC)及tempol处理RPMI-8226细胞后,通过Western blot技术检测阿霉素诱导下细胞内beclin 1、LC3等蛋白的表达水平。采用自噬抑制剂3-甲基腺嘌呤(3-methyladenine,3-MA)处理PRMI-8226细胞后,检测阿霉素诱导下细胞内ROS和凋亡的表达水平。结果: RPMI-8226细胞内beclin 1和LC3Ⅱ/LC3Ⅰ表达水平呈阿霉素剂量依赖性增加,当阿霉素诱导浓度达2 mg/L时,与对照组比较显著增加(P<0.05)。采用2 mg/L阿霉素处理RPMI-8226细胞,通过荧光显微镜观察,ROS水平较对照组明显增加。氧自由基清除剂NAC和tempol处理RPMI-8226细胞后,beclin 1和LC3Ⅱ/Ⅰ的表达水平较阿霉素处理组下降。采用自噬抑制剂3-MA处理细胞后,RPMI-8226细胞内ROS和凋亡的水平较阿霉素处理组及对照组增加。结论: 阿霉素能增加RPMI-8226细胞内自噬和ROS的生成,抑制自噬能增加阿霉素诱导下ROS和凋亡的水平,抑制ROS后能减少阿霉素诱导下多发性骨髓瘤细胞中的自噬。

关 键 词:多发性骨髓瘤  阿霉素  自噬  活性氧簇  
收稿时间:2015-11-20

Crosstalk of autophagy and ROS in multiple myeloma cells stimulated with doxorubicin
LUO Qi,CHEN Jian-bin.Crosstalk of autophagy and ROS in multiple myeloma cells stimulated with doxorubicin[J].Chinese Journal of Pathophysiology,2016,32(4):665-670.
Authors:LUO Qi  CHEN Jian-bin
Institution:Department of Hematology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China
Abstract:AIM: To investigate the relationship of autophagy and reactive oxygen species(ROS) in multiple myeloma cell line RPMI-8226 stimulated with doxorubicin. METHODS: The RPMI-8226 cells were stimulated with doxorubicin at different doses, and untreated cells were used as control. The protein expression of beclin 1 and LC3 was detected by Western blot. ROS production was analyzed by DCFH-DA fluorescence staining. After treated with or without 3-methyladenine(3-MA), the ROS production and apoptosis in RPMI-8226 cells were determined by DCFH-DA and flow cytometry, respectively. After treated with or without antioxidants tempol and N-acetyl-L-cysteine(NAC), the expression of beclin 1 and LC3 in RPMI-8226 cells was determined by Western blot. RESULTS: The protein levels of beclin 1 and LC3Ⅱ/LC3Ⅰ were increased in the RPMI-8226 cells stimulated with doxorubicin compared with untreated group. The ROS production was increased in the RPMI-8226 cells stimulated with 2 mg/L doxorubicin compared with untreated group. After treated with 3-MA, the ROS production and apoptosis in the RPMI-8226 cells stimulated with doxorubicin were increased compared with doxorubicin group. After treated with antioxidant NAC or tempol, the expression of beclin 1 and LC3 Ⅱ/I in the RPMI-8226 cells stimulated with doxorubicin was decreased compared with doxorubicin group.CONCLUSION: The autophagy and ROS levels are increased in RPMI-8226 cells stimulated with doxorubicin. Inhibition of autophagy increases the ROS production and apoptosis of RPMI-8226 cells stimulated with doxorubicin. Inhibition of ROS production reduces doxorubicin-induced autophagy in multiple myeloma cells.
Keywords:Multiple myeloma  Doxorubicin  Autophagy  Reactive oxygen species
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