偏钒酸钠对白血病细胞株HL-60细胞增殖和凋亡的影响及其与N-乙酰氨基半乳糖转移酶2的相关性

Effect of Sodium Metavanadate on Leukemia Cell Line HL-60 and Its Relation with ppGalNacT2

目的探讨偏钒酸钠(NaVO3)对HL-60细胞增殖及凋亡的影响,初步判断该影响与ppGalNac-T2的相关性。方法用不同浓度NaVO3作用HL-60细胞后,采用MTT法观察HL-60细胞生长抑制情况,琼脂糖凝胶电泳检测DNA条带,Hoechst 33258染色后荧光显微镜观察细胞核的变化,流式细胞术(FCM)检测细胞凋亡率,分子对接与分子动力学模拟初步判断钒对糖基转移酶ppGalNac-T2活性的影响。结果 MTT检测结果显示,低浓度的Na-VO3促进细胞增殖,高浓度的NaVO3抑制细胞的生长;高浓度NaVO3作用HL-60细胞24 h可见到DNA出现梯形条带,且细胞核呈现核浓缩现象;FCM测定不同浓度NaVO3(5×10-7、1×10-7、1×10-8、1×10-9、1×10-11mol/L)处理HL-60细胞24 h后细胞凋亡率分别是(69.1±2.3)%、(56.3±4.9)%、(39.6±6.5)%、(31.4±1.6)%、(12.2±3.4)%,明显高于HL-60细胞自然凋亡率(均P<0.05);VO3可以与N-乙酰氨基半乳糖转酶2(ppGalNac-T2)的活性中心相关结构形成氢键,并影响ppGalNAc-T2催化相关柔性结构的构象变化,进而影响底物进入和离开酶的活性中心,从而抑制酶的催化功能。结论 NaVO3可以诱导HL-60细胞凋亡,该作用可能与ppGalNAc-T2活性改变相关。

Objective To explore the effect of sodium metavanadate on cell apoptosis in HL-60 cells,and preliminarily judge whether the impact associate with ppGalNacT2.Methods MTT was used to observe cell proliferation.DNA（deoxyribonucleic acid） line was detected by agarose gel electrophresis.Changes of cell nucleus stained by Hoechst 33258 in HL-60 cells treated by sodium metavanadate were studied by fluorescence microscope.Flow cytometry（FCM） was used to detect cell apoptosis.Molecular docking and molecular dynamics simulations were used to judge the relationship between vanadium and the activity of ppGalNac-T2.Results Different concentrations of sodium metavanadate were used to treat HL-60 cells;24,48,72 h later MTT results showed that low doses of sodium metavanadated promoted cell proliferation,high doses of sodium metavanadate inhibited cell growth.DNA fragment appeared after HL-60 cells were exposed to high concentrations of sodium metavanadate,and cell nucleus were concentrated and fragmented.Treated with different concentrations of sodium metavanadate in concentration of 5×10-7,1×10-7,1×10-8,1×10-9,1×10-11mol/L,the apoptotic rate was（69.1±2.3）%,（56.3±4.9）%,（39.6±6.5）%,（31.4±1.6）%,（12.2±3.4）%.The apoptotic rate in the negative control group was only（1.2±0.5）%（P0.05）.VO3 might affect the activity of the enzyme by stemming conformational changes of a key loop of ppGalNAcT2.Conclusion Sodium metavanadate can induce apoptosis in HL-60 cells,which may associate with the activity of ppGalNac-T2.