核干细胞因子基因在急性白血病细胞表达的检测和意义

Detection and significance of expression of nucleostemin gene in human acute leukemia cells

目的检测核干细胞因子(Nucleostemin,NS)基因在急性白血病细胞的表达情况,探讨NS基因与急性白血病发病机制之间的关系。方法从NS基因的3个变异体中筛选出共同序列设计特异性引物,应用逆转录-聚合酶链式反应技术(RT-PCR)对白血病细胞系K562、HL60和急性粒细胞白血病(M1、M2a和M3),急性单核细胞白血病(M5a和M5b),急性淋巴细胞白血病(ALL)进行NS基因表达水平的检测,以健康人和良性贫血患者骨髓单个核细胞作为对照。PCR产物片段为418bp。结果K562、HL60明显高表达NS基因,它与内参相比灰度值分别为0·735±0·260、0·449±0·190;急性白血病病例有相似的结果。M1+M2a、M3、M5a、M5b、ALL的灰度值分别为0·687±0·210、0·408±0·160、0·866±0·270、0·448±0·190、0·403±0·190;对照组健康人和良性贫血不表达或极微弱表达NS基因;同一细胞类型白血病,细胞分化停滞在早期阶段的表达NS基因的水平明显高于细胞分化停滞在后期的白血病,如K562高于HL60;M1、M2a高于M3;M5a高于M5b(P<0·01)。结论NS基因在急性白血病细胞中呈高表达状态,这与白血病的发生、发展有一定关联;不同分化阶段的白血病细胞表达NS基因水平存在着差异。NS基因在急性白血病细胞的表达情况也显示了癌细胞和干细胞在某些方面的相似性,可能是一个潜在的基因治疗靶位。

Objective To determine the expression of Nucleostemin （NS） gene in acute leukemic cells, and to probe into the correlation between NS gene and pathogenesis of acute leukemia. Methods The common sequence was screened out from three variants of NS gene and the primers was designed. The NS gene expression was detected in K562 and HL60 leukemic cells by RT-PCR as well as in acute myeloblastic leukemia （ AML： M1, M2a, M3）, acute monocytic leukemia （AMoL ： M5a, M5b）, and acute lymphoblastic leukemia （ALL） respectively. The PCR product was a 418 bp fragment. The same detection was performed in bone marrow mononuclear cells of control groups including the healthy people and the patients with the benign anemia. Results The high level of NS gene expression was found in K562, HL60, and three groups of leukemic cells while no expression or only the extremely weak expression was detected in the healthy people group and the group of the patients with benign anemia. Compared with the internal standard of β- actin, the NS gene expression density scores of K562, HL60, M1 ＋ M2a, M3, M5a, M5b, and ALL were 0. 735 ±0. 260, 0. 449 ±0. 190, 0. 687±0. 210, 0.408 ±0. 160, 0. 866 ±0. 270, 0. 448 ±0. 190,0. 403 ± 0. 190, respectively. For the same cell type leukemia, the NS gene level expressed by the cells in the early stage of differentiation was significantly higher than that of the cells in later stage of differentiation. For example, the K562 expressed was higher than HL60, M1 ＋ M2a higher than M3, and M5a higher than M5b （P 〈 0. 01 ）. Conclusion The NS gene is overexpressed in acute leukemic cells, which is closely related to the origination and development of leukemia. There is difference among the expression level of NS gene in the leukemic cells of different differentiation stage. These results suggest that the cancer cells and the stem cells have similarity in some aspects. Maybe NS gene is a potential target in treatment of leukemia.