机构地区:[1]郑州大学第一附属医院儿内科,河南郑州450052 [2]濮阳市人民医院儿内科,河南濮阳457000
出 处:《河南医学研究》2020年第23期4231-4235,共5页Henan Medical Research
基 金:河南省高等学校重点科研项目计划(18A320074)。
摘 要:目的探讨丹参酮ⅡA(TanⅡA)联合三氧化二砷(ATO)对人急性早幼粒细胞白血病细胞株NB4增殖、凋亡及自噬的影响及其可能的分子机制,为临床采用TanⅡA治疗或辅助治疗白血病策略提供实验和理论依据。方法取对数生长期细胞进行实验,共分6组:4、8、16 mg·L^-1 TanⅡA组,0.5 mg·L^-1 ATO组,8 mg·L^-1 TanⅡA+0.5 mg·L^-1 ATO组,对照组加入9 g·L^-1生理盐水。采用MTT法检测TanⅡA与ATO单药及两药联合对NB4细胞增殖的抑制作用。采用流式细胞术(FCM)检测各组NB4细胞的凋亡率和自噬率。采用蛋白质印迹法检测凋亡特异蛋白半胱天冬酶-3(Caspase-3)及自噬特异蛋白Beclin-1、微管相关蛋白1轻链3(LC3)-Ⅱ的表达水平。结果TanⅡA对NB4细胞增殖的抑制作用呈时间和剂量依赖性。两药联合组对NB4细胞增殖的抑制率大于8 mg·L^-1 TanⅡA组和0.5 mg·L^-1 ATO组(均P<0.05)。8 mg·L^-1 TanⅡA作用于NB4细胞48 h后的凋亡率和自噬率分别为(11.28±3.32)%、(18.42±7.63)%,0.5 mg·L^-1 ATO作用于NB4细胞48 h后凋亡率和自噬率分别为(32.69±5.60)%、(34.36±7.60)%,两药联合作用后细胞凋亡率和自噬率分别为(44.02±6.47)%、(53.26±7.39)%,高于单药组(均P<0.05)。蛋白质印迹实验显示,TanⅡA联合ATO组NB4细胞中Caspase-3、Beclin-1、LC3-Ⅱ的相对表达量高于单药组和对照组(均P<0.05)。结论TanⅡA联合ATO有协同抑制NB4细胞增殖、诱导凋亡及促进自噬的作用,上调Caspase-3、Beclin-1和LC3-Ⅱ蛋白的表达水平是其可能的分子机制。Objective To investigate the effects of tanshinoneⅡA(TanⅡA)combined with arsenic trioxide(ATO)on proliferation,apoptosis and autophagy of human acute promyelocytic leukemia cell line NB4 and its possible molecular mechanism,and to provide experimental and theoretical basis for clinical treatment of leukemia with TanⅡA.Methods Logarithmic growth phase cells were took for experiment and divided into 6 groups:4,8,16 mg·L^-1 TanⅡA group,0.5 mg·L^-1 ATO group,8 mg·L^-1 TanⅡA+0.5 mg·L^-1 ATO group and control group(9 g·L^-1 saline).MTT method was used to detect the inhibitory effect of TanⅡA and ATO on proliferation of NB4 cells.Flow cytometry(FCM)was used to detect apoptosis rate and autophagy rate of NB4 cells in each group.Western blotting was used to detect the expression levels of apoptosis-specific protein Caspase-3(Caspase-3),autophagy-specific protein Beclin-1,and microtubule-associated protein 1 light chain 3(LC3)-Ⅱ.Results The inhibitory effect of TanⅡA on proliferation of NB4 cells was time-and dose-dependent.The inhibition rate of the two-drug combination group on proliferation of NB4 cells was greater than that of 8 mg·L^-1 TanⅡA group and 0.5 mg·L^-1 ATO group(both P<0.05).After 8 mg·L^-1 TanⅡA was applied to NB4 cells for 48 hours,apoptosis rate and autophagy rate were(11.28±3.32)%and(18.42±7.63)%,respectively.With 0.5 mg·L^-1 ATO acting on NB4 cells for 48 hours,apoptosis rate and autophagy rate were(32.69±5.60)%and(34.36±7.60)%,respectively.A poptosis rate and autophagy rate after the combined action of the two drugs were(44.02±6.47)%and(53.26±7.39)%,respectively,which were higher than those in the single-agent group(both P<0.05).Western bloting showed that relative expression levels of Caspase-3,Beclin-1,and LC3-Ⅱin NB4 cells in combination group were higher than those in single-drug group and control group(all P<0.05).Conclusion TanⅡA combined with ATO could synergistically inhibit proliferation of NB4 cells,induce apoptosis and promote autophagy.Up-regulating t
关 键 词:急性早幼粒细胞白血病 丹参酮ⅡA 三氧化二砷 自噬 凋亡
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