Ventura G J, Hester J P, Buescher E S, Vadhan-Raj S, Durrett A, Reading C L
Department of Hematology, University of Texas M.D. Anderson Cancer Center, Houston.
Exp Hematol. 1990 Sep;18(8):878-82.
We have modified a limiting dilution liquid culture assay, used to quantify hematopoietic progenitor frequency, to simultaneously assess cellular proliferation and differentiation. The frequency of colonies from cord blood obtained with recombinant human interleukin 3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) in combination was comparable to cultures with IL-3 alone. However, IL-3 and GM-CSF in combination were synergistic for higher granulocyte proliferation than IL-3 alone, indicating that enhanced granulocyte production occurred via action of GM-CSF on progenitor cell populations already stimulated into proliferation by IL-3. Peak proliferation was evident at 4 weeks in culture, with metamyelocytes predominating; at 6 weeks, mostly neutrophils and eosinophils were present, and eosinophils were more numerous in cultures with IL-3. Increasing concentrations of erythropoietin (epo) in liquid culture with IL-3 or GM-CSF decreased absolute granulocyte yield while stimulating erythroid proliferation. The influence of epo on lineage morphology for cells plated in methylcellulose was markedly less evident by comparison, arguing against an inductive effect of epo to shift progenitor cell lineage. This liquid culture methodology may be a useful tool for preclinical screening of cytokines on human hematopoietic progenitor cells.
我们改良了一种用于定量造血祖细胞频率的有限稀释液体培养测定法，以同时评估细胞增殖和分化。用重组人白细胞介素3（IL-3）和粒细胞-巨噬细胞集落刺激因子（GM-CSF）联合培养获得的脐血集落频率与单独使用IL-3培养的结果相当。然而，IL-3和GM-CSF联合使用对粒细胞增殖的促进作用比单独使用IL-3更强，这表明粒细胞生成增加是通过GM-CSF作用于已被IL-3刺激进入增殖状态的祖细胞群体实现的。培养4周时增殖达到峰值，此时晚幼粒细胞占主导；6周时，大多是中性粒细胞和嗜酸性粒细胞，且在含有IL-3的培养物中嗜酸性粒细胞数量更多。在含有IL-3或GM-CSF的液体培养中，增加促红细胞生成素（epo）的浓度会降低粒细胞绝对产量，同时刺激红系增殖。相比之下，epo对接种在甲基纤维素中的细胞谱系形态的影响明显较小，这表明epo不存在诱导祖细胞谱系转变的作用。这种液体培养方法可能是在临床前筛选细胞因子对人造血祖细胞作用的有用工具。
