目的 观察三氧化二砷（arsenic trioxide,ATO）在体外影响反转录病毒整合位点1基因（ecotropic viral integration site-1,EVI1）基因对造血转录因子的调控作用。方法 实验选取EVI1高表达的急性髓系白血病细胞株THP-1,利用健康成人外周血单个核细胞作为对照,通过反转录实时荧光定量聚合酶链反应（real-time fluorescent quantitative polymerase chain reaction,RFQ-PCR）,检测EVI1基因及造血转录因子GATA1、GATA2、RUNX1、MPO、LMO2、CMYB、PU.1和SCL的mRNA相对表达量。分别以1 μmol/L、3 μmol/L、5 μmol/L的ATO溶液处理THP-1细胞株后,再通过反转录RFQ-PCR检测EVI1基因及造血转录因子的mRNA相对表达量。结果 通过反转录RFQ-PCR检测证实,存在EVI1基因的THP-1细胞中GATA2和CMYB基因的表达上调,而GATA1、RUNX1、MPO、LMO2、PU.1及SCL基因的表达水平下调。ATO对EVI1基因的下调作用具有浓度与时间依赖性,并对其他造血转录因子进行调控。结论 通过体外研究发现,高表达EVI1基因的THP-1细胞有GATA2基因的表达上调,同时存在其他造血转录因子的表达异常,与促进原始细胞增殖及髓系、红系的分化成熟受抑密切相关。ATO可特异性地下调EVI1基因的表达,并抑制GATA2转录因子的激活。
Objective: To investigate the effect of arsenic trioxide (ATO) on EVI1 gene in regulating hematopoietic transcription factors in vitro. Methods: Acute myeloid leukemia cell line with high EVI1 gene expression-THP-1 cell line was chosen to study the relative mRNA expressions of EVI1 gene and hematopoietic transcription factors GATA1, GATA2, RUNX1, MPO, LMO2, CMYB, PU.1 and SCL by reverse transcription-real-time fluorescent quantitative polymerase chain reaction (RFQ-PCR) in vitro. Healthy adult peripheral blood mononuclear cells were used as controls. THP-1 cell line was then treated with ATO solution with concentrations of 1 μmol/L, 3 μmol/L, 5 μmol/L. Then the relative mRNA expressions of EVI1 gene and the above-mentioned hematopoietic transcription factors were re-determined by RFQ-PCR. Results: The expressions of GATA2 and CMYB increased in THP-1 cell line expressing high EVI1 gene, and the levels of GATA1, RUNX1, MPO, LMO2, PU.1 and SCL decreased in vitro. ATO had the ability to down-regulate EVI1 gene in a dose-dependent and time-dependent manner, and the in turn regulatory effect on other hematopoietic transcription factors was observed. Conclusions: THP-1 cells have an increased expression of GATA2 and then indirectly influencing other transcription factors to promote the proliferation of immature cells and inhibit the differentiation and maturation of myeloid and erythroid cells. ATO could specifically down-regulate the expression of EVI1 gene and decrease the activation of GATA2.
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