四嗪二甲酰胺对人白血病细胞系SHI-1细胞体外作用的研究  

Effects of N,N-di-(m-methylphenyl)-3,6-dimethyl-1,4-dihydro-1,2,4,5-tetrazine-1,4-dicarboamide(ZGDHu-1)on SHI-1 leukemia cells in vitro

作　　者：周永列[1] 吕亚萍[2] 胡惟孝[2] 邱莲女[1] 王文松[1] 刘建栋[1] 

机构地区：[1]浙江省人民医院中心实验室,杭州310014 [2]浙江工业大学药学院

出　　处：《中华血液学杂志》2006年第6期361-365,共5页Chinese Journal of Hematology

基　　金：浙江省科技厅资助项目（2003c33019）

摘　　要：目的观察四嗪二甲酰胺(ZGDHu-1)体外抑制白血病细胞系 SHI-1细胞增殖,诱导细胞分化和凋亡作用,并初步探讨其作用机制。方法将不同浓度 ZGDHu-1与 SHI-1细胞在体外共培养,用细胞计数、锥虫蓝染色、MTT 法、5′-溴-2′脱氧尿苷(Brdu)-ELISA 法观察 ZGDHu-1对 SHI-1细胞增殖的抑制作用;用细胞形态学、DNA 含量测定及细胞周期分析、膜联蛋白Ⅴ/碘化丙锭(Annexin Ⅴ/P1)双标记和 Hoechst 33258荧光染色等分析细胞凋亡。通过 NBT 试验、细胞表面抗原 CD11b、CD14、CD64检测和细胞形态学观察分化情况。用流式细胞术检测 ZGDHu-1诱导 SHI-1细胞分化和凋亡过程中 P38MAPK 和 STAT3磷酸化表达的变化。结果 ZGDHu-1能抑制 SHI-1细胞增殖和活力,呈现作用时间和剂量的量效关系,48和72h/C_(50)值分别为250ng/ml,85 ng/ml。ZGDHu-1作用于 SHI-1细胞后大部分细胞阻滞于 G_2/M 期,200ng/ml ZGDHu-1作用48h,SH-1细胞 G_2/M 期比例为(48.4±2.1)%;出现典型的细胞凋亡形态改变,DNA 片断化,检出亚 G_1峰,AnnexinⅤ/PI 和 Hoechst 33258荧光染色显示凋亡细胞的特征性改变。SHI-1细胞经2～50ng/ml ZGDHu-1作用3 d 后,细胞发生部分分化,NBT 阳性率增加,细胞表面 CD11b、CD14、CD64表达上调。ZGDHu-1作用 SHI-1细胞后磷酸化P38MAPK 表达增强,而磷酸化 STAT3表达降低。结论 ZGDHu-1能抑制 SHI-1细胞增殖和细胞活力.诱导细胞分化,促进细胞凋亡,其机制可能与 P38MAPK 的活化增强和 STAT3的活化抑制有关。Objective To study the effect of ZGDHu-1 on proliferation, differentiation and apoptosis in SHI-1 human leukemia cell line and explore its possible mechanism. Methods SHI-1 ceils were cultured with different concentration of ZGDHu-1 and for different time. The cell proliferation was analysed by cell counting, alive cell count, MTT assay and Brdu-ELISA. Cell apoptosis was analysed by morphology, DNA content, Annexin-V/PI and Hoechst 33258 labeling method. Cell differentiation were assayed by morpholo- gy,expression of CD11 b,CDI4 and CD64 and NBT reduction. 3＇he expressions of phosphorylated p38MAPK or STA＇I3 were analysed by flow cytometry. Results ZGDHu-1 inhibited SHI-1 cell proliferation in a time and dose dependent manner, the IC50- 48 h and IC50- 72 h were 250 ng/ml and 85 ng/ml, respectively. The majority of SHI-1 cells were an＇ested in G2/M phase. 48h after treated with 200 ng/ml ZGDHu- 1, and those in G2/M phase accounted for （48.4 ± 2.1 ）%. The SHI-1 cells apeposis was increased with a time- and doesdependent manner. The morphology of SHI-1 cells cultured with 2 -50 ng/ml ZGDHu-1 for three days become more mature with higher NBT positivity and up-regulated expressions of CDllb, CDI4 and CD64. The expression of phosphor-p38MAPK was increased and phosphor-STAT3 down-regulated by the treatment of ZGDHu-1, Conclusion ZGI）Hu-1 can inhibit SHI-1 cell proliferation and induce the cell differentiation and apoptosis. The mechanism may associate with its up-regulation of phosphor-p38MAPK and down-regulation phosphor-STAT3.

关 键 词：甲酰胺类 细胞系 SHI-1 细胞凋亡 细胞分化 

分 类 号：R544.1[医药卫生—心血管疾病]