外源性Wnt5a诱导K562细胞分化表型实验探究.docVIP

外源性Wnt5a诱导K562细胞分化表型实验探究 　作者：袁媛，司维柯，李招权，潘静，赵宸 【摘要】 本研究探讨外源性Wnt5a诱导K562细胞定向分化的作用。用重组腺病毒AdWnt5a、AdGFP感染CHO细胞，分别制备Wnt5a和GFP对照条件培养液，并处理K562细胞1-7天，用光学显微镜和电子显微镜观察细胞形态变化；采用过氧化物酶（POX）、糖原（PAS）和非特异性酯酶（αNAE）染色、细胞表面分化抗原CD13、CD14、CD68鉴定K562细胞的分化表型；用流式细胞术检测细胞分化周期的变化。结果表明： 与对照GFP条件培养液相比，Wnt5a条件培养液处理的K562细胞形态和超微结构均出现了分化成熟的特征；POX、PAS、αNAE细胞化学染色均为阳性，并且αNAE阳性70％可被氟化钠抑制；单核细胞系表面标志抗原CD14、CD68和粒细胞系表面标志抗原CD13经免疫细胞化学染色均呈阳性，但CD13与对照比较无显著性差异；Wnt5a处理K562细胞5天，细胞周期阻滞在G2期。结论： 外源性的Wnt5a可诱导K562细胞向成熟方向分化，并且有向单核细胞系分化的现象。 【关键词】 诱导分化 Differentiation Phenotypes of K562 Cells Induced by Exogenous Wnt5a Abstract This study was aimed to investigate the effect of exogenous Wnt5a on directional differentiation of K562 cells. Wnt5a and GFP condition mediums were prepared by recombinant adenoviral vector AdWnt5a and AdGFP transfecting CHO cells. K562 cells were treated with Wnt5a and the GFP condition mediums for 1-7 days as Wnt5a treated group and control group respectively. The morphological changes of K562 cells were observed by light microscope and electron microscope; the differentiation phenotypes of K562 cells were identified by the cytochemical staining of POX，PAS，αNAE and immunocytochemistry of CD13,CD14, CD68, and the effect of Wnt5a on cell cycle distribution of K562 cells was detected by flow cytometry. The results showed that the morphology and ultrastructure of K562 cells treated by Wnt5a displayed differentiation mature feature; both POX and PAS staining showed higher positive ratio in Wnt5a treated group than that in control group; the αNAE staining also was positive， but positive intensity in Wnt5a treated group could be inhibited up to 70％ by NaF. The expressions of monocytic differentiation antigens of CD14, CD68 in Wnt5a treated group were higher than those in control group， but the expression differences of granulocytic differentiation antigen CD13 between Wnt5a treated group and control group were not significant. The cell cycle in treated group was blocked at G2 phase as compared