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提取用2μmol/LAs2O3对NB4细胞株和APL体外干预及用0.16mg·kg-1·天-1As2O3静脉输注治疗干预前后的白血病细胞线粒体基因,PCR扩增后直接测序,干预前后的序列对比分析,同时流式细胞仪对细胞凋亡率测定.分析了As2O3在体内和体外作用对原代急性早幼粒细胞白血病细胞(APL)和NB4细胞株的线粒体基因(mtDNA)非编码区D-loop的基因突变与细胞凋亡的关系.研究发现,As2O3处理后mtDNA的D-loop区基因突变位点数目较处理前显著增多,mtDNA的D-loop区基因突变率与细胞凋亡率呈正相关关系,相关系数rNB4-As2O3=0.973818,rAPL-As2O3=0.934703.突变类型包括转换、颠换、插入和缺失.本实验仅是初步研究,尚未发现As2O3对mtDNA的D-loop区稳定的突变位点和突变规律.结果显示,As2O3体内外干预均能加剧急性早幼粒细胞白血病细胞mtDNA的D-loop区基因突变,并与其促凋亡作用呈正相关,mtDNA的D-loop区可能是砷剂治疗白血病的一个靶点.
In vitro intervention of NB4 cell line and APL with 2μmol / L As2O3 and extraction of mitochondrial DNA of leukemia cells before and after intervention with 0.16mg · kg-1 · day-1As2O3 intravenous infusion of 2μmol / L As2O3 were performed, and the sequences were directly sequenced after PCR amplification , And the flow cytometry was used to measure the apoptosis rate.An in vitro and in vivo effects of As2O3 on mitochondrial DNA (mtDNA) D-loop of primary acute promyelocytic leukemia cells (APL) and NB4 cell lines Gene mutation and apoptosis.We found that the number of mtDNA D-loop gene mutation sites was significantly increased after treatment with As 2 O 3 than that before treatment, and the mutation rate of mtDNA D-loop region was positively correlated with the apoptosis rate , The correlation coefficient rNB4-As2O3 = 0.973818, rAPL-As2O3 = 0.934703. The types of mutations include translocation, transversion, insertion and deletion.This experiment is only a preliminary study, but we have not found a stable mutation site of As2O3 on mtDNA D-loop region and The results showed that as2O3 in vitro and in vivo can exacerbate the mutation of mtDNA D-loop gene in acute promyelocytic leukemia cells and positively correlated with its pro-apoptotic effect, and the D-loop region of mtDNA may be an arsenic agent in the treatment of leukemia A target of disease.