摘要
本研究旨在探讨SHP-1及C-kit基因在急性白血病细胞中的表达水平及三氧化二砷(As2O3)去甲基化作用对SHP-1及C-kit表达的影响。用RT-PCR方法检测药物处理的HL-60细胞组与对照组SHP-1、C-kit基因的mRNA的表达水平。用甲基化特异性聚合酶链反应(MSP)检测HL-60细胞中SHP-1基因的甲基化状态的变化。结果表明,在原本不表达SHP-1 mRNA的HL-60细胞中,经过As2O3的去甲基化作用后,SHP-1得到表达,而使原来高表达的C-kit mRNA的表达水平随之下降。当1.0μmol/L、2.5μmol/L、5.0μmol/L As2O3分别作用于白血病细胞株时,去甲基化作用增强,SHP-1 mRNA的表达水平呈上升趋势,C-kit mRNA的表达水平呈下降趋势,经两两比较,二者间有统计学意义(P<0.05)。结论:HL-60细胞株中SHP-1 mRNA表达缺如,经去甲基化作用后表达恢复,说明SHP-1基因高度甲基化与白血病发生有关;在白血病形成中可能存在C-kit mRNA表达的异常增高。As2O3对SHP-1与C-kit的表达水平的影响呈剂量依赖性,浓度越高,SHP-1平均表达水平越高,C-kit mRNA表达水平越低,在特定的浓度下,呈现时间依赖性;SHP-1 mRNA与C-kit mRNA表达呈负相关,提示白血病细胞中SHP-1表达降低或缺如削弱了对C-kit信号通路的负调控而在白血病形成中发挥作用。
This study was aimed to investigate the expression level of SHP-1 and C-kit genes in acute leukemia HL- 60 cells and effect of inhibitor As2 03 demethylation on SHP-1 and C-kit genes expression. RT-PCR was used to detect the expression level of SHP-1 and C-kit mRNA in drug-treated cell group and control group. The methylation specific PCR（MSP） was applied to measure the methylation status of SHP-1 gene in HL-60 cells. The results showed that after being treated with As203 the recovery of SHP-1 gene expression was observed in HL-60 cells in which SHP-1 mRNA originally did not expressed, meanwhile the expression level of C-kit mRNA in HL-60 cells with high expression de creased. When HL-60 cells were treated with As203 of 1.0,2.5,5.0 μmol/L, the demethylation effects was enhanced, the expression of SHP-1 mRNA displayed an ascending tendency, and expression of C-kit mRNA showed an descending tendency in dose-dependent manner （P 〈 0.05）. It is concluded that the absence of SHP-1 mRNA expression in HL-60 cells and recovery of expression after treatment with As203 suggest the hypermethylation of SHP-1 gene related with pathogenesis of leukemia, and the abnormal increase of C-kit mRNA expression maybe exist in formation of leukemia. The effect of AS203 on expression of SHP-1 and C-kit shows dose-dependency, the higher the As203 cocentration, the higher the SHP-1 expression and the lower the C-kit expression, moreover, the effect of As203 shows time-dependency in specific concentration. The SHP-1 mRNA expression negatively relates with C-kit mRNA expression, suggesting that the decrease or absence of SHP-1 expression in leukemia cells weakens the negative regulation on C-kit signathing pathway, thus plays a role in the formation of leukemia.
出处
《中国实验血液学杂志》
CAS
CSCD
北大核心
2013年第3期613-616,共4页
Journal of Experimental Hematology
