丙戊酸钠诱导髓系白血病细胞分化相关的分子标志物研究

Molecular marker associated with valproate induced differentiation in myeloid leukemia cells

目的:研究组蛋白去乙酰化酶(HDAC)抑制剂丙戊酸钠(VPA)诱导髓系白血病细胞分化相关的分子标志物。方法:采用流式细胞术检测细胞表面分化抗原的表达,采用实时定量PCR和蛋白印迹法检测细胞周期相关蛋白、凋亡相关蛋白、活性氧簇(ROS)相关蛋白及组蛋白甲基化调控蛋白的表达。结果:VPA较低剂量(1 mmol/L)即诱导HL-60细胞分化,K562细胞对VPA诱导分化不敏感。VPA诱导细胞分化作用与组蛋白乙酰化基础水平及药物处理后调变间无显著相关性,VPA诱导分化与细胞周期相关蛋白、凋亡相关蛋白、ROS相关调控蛋白的表达无影响。VPA诱导分化效应与细胞不同HDAC表达水平相关,K562细胞高表达HDAC6,对VPA诱导分化不敏感,且经VPA处理后细胞组蛋白K9甲基化水平较HL-60细胞明显提高。结论:VPA诱导白血病细胞分化的作用可能与细胞HDAC6的低水平表达及组蛋白K9甲基化水平调控有关,而与组蛋白乙酰化基础水平及细胞周期相关蛋白、ROS相关调控蛋白等无关。

Objective To investigate the molecular marker associated with valproate（VPA）,a histone deacetylase（HDAC） inhibitor,induced differentiation in myeloid cells.Methods Expression of cell surface differentiation antigen was analyzed by flow cytometry,and real-time quantitative PCR and Western blot were used to detect cell cycle,apoptosis,ROS related proteins and histone methylation regulatory protein.Results VPA at low concentration（1 mmol/L）can induce differentiation in HL-60 cells,while K562 cells were not sensitive to VPA induced differentiation.No significant relevance was detected between VPA induced differentiation and histone acetylation level before and after the treatment.The expressions of cell cycle,apoptosis and ROS related proteins were similar in both HL-60 and K562 cells.Expression of HDAC was different in HL60 and K562 cells.The expression of HDAC6 was significantly higher in K562 cells than in HL-60 cells.Upregulation of histone K9 methylation was significantly higher in K562 than in HL-60 cells after treatment with VPA.Conclusions VPA can induce differentiation in HL-60 cells,which might be associated with the low level of HDAC6 and modulation of histone K9 methylation,but not the level of acetylated histone protein and expressions of cell cycle,apoptosis and ROS related proteins.