高三尖杉酯碱对白血病K562细胞增殖和DNA损伤相关蛋白表达的影响

Effects of homoharringtonine on proliferation and DNA damage related protein expression in leukemia K562 cells

目的研究高三尖杉酯碱(HHT)对人白血病细胞K562细胞增殖和DNA损伤及相关蛋白表达的影响。方法体外培养人白血病K562细胞,将其分为HHT组和对照组,HHT组又分为5×10-9、1×10-8、5×10-8、1×10-7 mol/L不同浓度的HHT亚组。CCK-8方法检测不同浓度HHT对K562细胞增殖的影响;Western blot检测不同浓度的HHT对K562细胞增殖[兔抗人细胞周期蛋白D1(Cyclin D1)和兔抗人核蛋白MKI67(Ki-67)]和DNA损伤[兔抗人组蛋白H2AX的磷酸化(γ-H2AX)和磷酸化兔抗人毛细血管扩张性共济失调症突变蛋白激酶(p-ATM)]相关蛋白表达的影响。结果HHT对人白血病K562细胞有显著抑制作用,不同浓度(5×10-9、1×10-8、5×10-8、1×10-7 mol/L)HHT抑制率分别为(4.70±0.45)%、(9.80±0.35)%、(34.00±0.29)%和(41.00±0.64)%;与对照组比较,5×10-9、1×10-8、5×10-8、1×10-7 mol/L HHT组均可下调Cyclin D1蛋白表达,差异均有统计学意义(均P<0.05)。与对照组比较,5×10-8、1×10-7 mol/L HHT组Ki-67蛋白表达水平明显下调,差异均有统计学意义(均P<0.05)。与对照组比较,5×10-8、1×10-7 mol/L HHT组可诱导γ-H2AX蛋白的表达上调,差异均有统计学意义(均P<0.05)。与对照组比较,1×10-8、5×10-8、1×10-7 mol/L HHT组可上调p-ATM水平,差异均有统计学意义(均P<0.05)。结论HHT可通过下调增殖相关蛋白的表达抑制白血病K562细胞的增殖,并可能对细胞DNA损伤具有诱导作用。

Objective To study the effects of homoharringtonine(HHT)on proliferation and DNA damage related protein expression in human leukemic K562 cells.Methods Human leukemia K562 cells were cultured in vitro,and divided into HHT group and control group.HHT group was further divided into HHT subgroups with different concentrations of 5×10-9,1×10-8,5×10-8 and 1×10-7 mol/L.The effects of HHT at different concentrations on the proliferation of K562 cells were detected by CCK-8 method.The effects of different HHT concentrations on K562 cells proliferation(rabbit anti human cyclin D1[Cyclin D1]and rabbit anti human nuclear protein MKI67[KI-67])and the expression of DNA damage(rabbit anti human histone H2AX phosphorylation[γ-H2AX]and phosphorylated rabbit anti human ataxia telangiectasia mutated protein kinase[p-ATM])relative protein were analyzed by Western blot.Results HHT significantly inhibited human leukemic K562 cells,with HHT inhibition rates(4.70±0.45)%,(9.80±0.35)%,(34.00±0.29)%,and(41.00±0.64)%at different concentrations(5×10-9,1×10-8,5×10-8 and 1×10-7 mol/L),respectively.Compared with the control group,Cyclin D1 protein expression could be down-regulated in 5×10-9,1×10-8,5×10-8 and 1×10-7 mol/L HHT groups,and the differences were statistically significant(all P<0.05).Compared with the control group,Ki-67 expression levels in 5×10-8 and 1×10-7 mol/L HHT group were significantly down-regulated,with statistically significant differences(all P<0.05).Compared with the control group,the upregulation of the expression ofγ-H2AX proteins were induced in 5×10-8 and 1×10-7 mol/L HHT groups,with statistically significant differences(all P<0.05).Compared with the control group,p-ATM levels were increased in the 1×10-8,5×10-8 and 1×10-7 mol/L HHT groups,with statistically significant differences(all P<0.05).Conclusion HHT can inhibit the proliferation of leukemic K562 cells by down-regulating the expression of proliferation-related proteins,and may induce cell DNA damage.