核仁磷酸蛋白基因突变对K562白血病细胞分化的影响

Effect of Nucleophosmin(NPM1) Mutations on the Differentiation of K562 Leukemic Cells

核仁磷酸蛋白基因(nucleophosmin,NPM1)突变在急性髓系白血病的发生发展中发挥着重要作用,而与白血病分化阻滞的关系尚未完全阐明。为探讨NPM1基因突变对白血病细胞体外分化的影响,将携带NPM1 A型突变(NPM1-mA)的表达质粒载体pEGFPC1-NPM1-mA转染白血病K562细胞系,构建稳定表达NPM1-mA蛋白的细胞株(K562 mA),同时设立野生型NPM1转染组(K562 wt)、空载体转染组(K562 C1)和未处理组(K562)为对照。利用豆蔻酰佛波醇乙酯(PMA)诱导各组细胞分化,瑞氏–吉姆萨染色观察细胞分化的形态改变,计算诱导分化率;相差显微镜计数贴壁细胞数量;流式细胞术分析细胞表面分化抗原CD41的表达。结果显示,PMA作用72 h后,与对照组相比,K562 mA组细胞的诱导分化率及贴壁细胞数明显降低(P<0.05);同时,CD41的表达受到显著抑制(P<0.01)。提示NPM1基因突变能够阻滞白血病细胞系K562的体外分化。

Nucleophosmin（NPM1） mutations play important roles in the leukemogenesis,whereas the relationship between NPM1 mutations and the differentiation block of leukemic cells have not yet been elucidated.To explore the effect of NPM1 mutations on the in vitro differentiation of leukemic cells,the plasmid pEGFPC1-NPM1-mA carrying NPM1 mutation A（NPM1-mA） was transfected into K562 cells,and the leukemic cells with stably expressed NPM1-mA protein（K562 mA） were established.The K562 cells transfected with pEGFPC1-NPM1-wt（K562 wt）,pEGFP-C1（K562 C1） and the untreated cells（K562） were used as control.After induction with phorbol-12-myristate-13-acetate（PMA）,the morphological changes of cells were observed under light microscope following Wright-Giemsa staining and the differentiation rate was estimated.Adherent cells were examined by phase contrast light microscopy.The expressions of surface marker CD41 were analyzed by flow cytometry.Compared with the control groups,the differentiation rates and the percentage of adherent cells in K562 mA group decreased obviously after PMA treatment for 72 h（P0.05）.Meanwhile,the expression of CD41 was inhibited significantly in K562 mA group（P0.01）.Our data indicate that NPM1 mutations block differentiation of K562 leukemic cells in vitro.