组蛋白去乙酰化酶抑制剂SAHA阻断MAPK信号通路并诱导HL-60细胞凋亡  

Histone Deacetylase Inhibitor SAHA Induces Inactivation of MAPK Signaling and Apoptosis in HL-60 Cells

作　　者：王颖[1] 王生余[1] 侯春梅[1] 徐元基[1] 杜芝燕[1] 于晓妉[1] 

机构地区：[1]军事医学科学院基础医学研究所病理生物学研究室,北京100850

出　　处：《中国实验血液学杂志》2007年第2期267-271,共5页Journal of Experimental Hematology

基　　金：国家自然科学基金重点项目(编号300330620);面上项目(编号30470898)资助

摘　　要：本研究旨在探讨组蛋白去乙酰化酶抑制剂-SAHA(suberoylanilide hydroxamicacid)诱导人急性髓性白血病细胞株HL-60凋亡的分子机制。采用MTT法,瑞氏-姬姆萨染色和Hoechst33342/PI荧光染色方法,检测SAHA对HL-60细胞生长的抑制作用及细胞的形态变化;通过流式细胞术、Western-blot方法测定HL-60细胞的周期变化以及多种信号蛋白的表达。结果表明:SAHA可以呈剂量时间依赖性抑制HL-60细胞的生长;经2μmol/L SAHA处理12-48小时,HL-60细胞显示细胞周期被阻滞于G0/G1期;经Hoechst33342荧光染色后细胞核出现明显的凋亡小体结构;Western blot检测证实,SAHA作用HL-60细胞后caspase3被激活,其底物蛋白聚ADP核糖聚合酶[po-ly(ADP-ribose)polymerase,PARP]发生剪切,细胞发生凋亡;对细胞凋亡相关信号转导通路P44/42 MAPK及PI3K/Akt检测结果表明,涉及MAPK信号通路的Raf及ERK激酶磷酸化受到明显抑制,而Akt及其下游mTOR激酶的活性没有明显改变。结论:SAHA对HL-60细胞的杀伤作用主要是通过诱导细胞凋亡实现,与细胞增殖相关的P44/42 MAPK信号通路被阻断是细胞凋亡发生的机制之一。The study was aimed to investigate the molecular mechanisms of histone deacetylase inhibitor SAHA -induced apoptosis of acute myeloid leukemia cell line HL-60. The effect of SAHA on HL-60 cell proliferation was detected by MTT assay and the cell morphological changes were observed with Wright-Giemsa and Hoechst33342 staining. The cell cycle distribution was determined by flow cytometry and the expression of cell signaling proteins were detected by Western-blot analysis. The results showed that SAHA inhibited the proliferation of HL-60 cells in dose- and time-dependent manners,after 2 μmol/L SAHA exposure for 12 -48 hours, the cell cycle was arrested at G0/G1 phase and apoptotic cell death was confirmed by either defined apoptotic bodies stained by Hoechst33342, Western blot showed cleaved-PARP, which represents the activation of caspase 3. The Western blot analysis indicated the activation of two important survival signal pathways after SAHA treatment, the phosphorylation of Raf and its downstream ERK kinases were remarkable downregulated, whereas the phosphorylation of AKT and its downstream molecular mTOR were not changed. It is concluded that SAHA-induced apoptosis of HL-60 cells is mediated by inactivation of p44/42 MAPK signaling pathway.

关 键 词：组蛋白去乙酰化酶抑制剂 SAHA MAPK 信号转导 细胞凋亡 HL-60细胞 

分 类 号：R733.7[医药卫生—肿瘤] R329.28[医药卫生—临床医学]