Veiby O P, Lyman S D, Jacobsen S E
Blood Cell Growth Factors Laboratory, Hipple Cancer Research Center, Dayton, OH 45439, USA.
Blood. 1996 Aug 15;88(4):1256-65.
Multiple cytokines can synergize to stimulate the in vitro proliferation and exclusive myeloid differentiation of multipotent bone marrow progenitor cells. The ligand for c-kit (stem cell factor [SCF]) plays a key role in stimulating myeloid and erythroid cell production of primitive hematopoietic progenitors. SCF in combination with interleukin-7 (IL-7) can also stimulate the combined myeloid and B-cell differentiation of uncommitted hematopoietic progenitor cells as well as the growth of early B-cell progenitor cells, although the involvement of c-kit in early B lymphopoiesis remains controversial. In the present study, the flt3-ligand (FL), which, in combination with other cytokines, has overlapping activities with SCF on myeloid cell production from uncommitted progenitors, was investigated for its ability to induce selective stroma-independent B-cell commitment from uncommitted Lin-Sca-1+ bone marrow progenitor cells. IL-7 alone did not induce any clonal growth and FL alone gave rise to a few clusters (< 50 cells) but no colonies (> 50 cells), whereas the combined stimulation with FL and IL-7 resulted in clonal growth of 10% of Lin-Sca-1+ bone marrow cells. After 12 days of incubation of Lin-Sca-1+ cells in FL + IL-7, an almost 400-fold increase in cell production was observed. Phenotyping showed that greater than 99% of the cells produced were of the B-cell lineage, in that they expressed B220, but not cell surface markers specific for myeloid, erythroid, or T-cell lineages. Furthermore, the cells did not express cytoplasmic mu-heavy chain (cmu) or surface IgM, but were positive for CD24 (heat stable antigen [HSA]) and CD43 (leukosialin), suggesting that the cells produced were blocked at a late pro-B-cell stage. Interestingly, although all FL + IL-7-responsive Lin-Sca-1+ progenitor cells and the resulting pro-B cells expressed c-kit, FL + IL-7 was much more potent (62-fold) than SCF + IL-7 in stimulating production of cells of the B-cell lineage. In addition, whereas FL + IL-7 selectively stimulated the production of pro-B cells, SCF + IL-7 predominantly stimulated the production of mature granulocytes. Replating studies showed that FL + IL-7-responsive Lin-Sca-1+ progenitors were not committed to the B-cell lineage, because after 2 days of incubation in FL + IL-7, 80% of the progenitors retained a myeloid potential. As much as 27% of the FL + IL-7-responsive progenitors remained uncommitted after 7 days of incubation, but all had committed to the B-cell lineage after 10 days of incubation in FL + IL-7. These results show that FL much more potently and selectively than SCF synergizes with IL-7 to enhance B-cell commitment and development from uncommitted progenitor cells.
多种细胞因子可协同刺激多能骨髓祖细胞的体外增殖及特异性髓系分化。c-kit的配体（干细胞因子[SCF]）在刺激原始造血祖细胞的髓系和红系细胞生成中起关键作用。SCF与白细胞介素-7（IL-7）联合使用时，也可刺激未定向造血祖细胞的髓系和B细胞联合分化以及早期B细胞祖细胞的生长，尽管c-kit在早期B淋巴细胞生成中的作用仍存在争议。在本研究中，研究了fms样酪氨酸激酶3配体（FL），它与其他细胞因子联合使用时，在未定向祖细胞的髓系细胞生成方面与SCF具有重叠活性，探讨其能否诱导未定向的Lin-Sca-1⁺骨髓祖细胞发生选择性的不依赖基质的B细胞定向分化。单独使用IL-7不会诱导任何克隆生长，单独使用FL可形成少数细胞簇（<50个细胞）但无集落（>50个细胞），而FL与IL-7联合刺激可导致10%的Lin-Sca-1⁺骨髓细胞发生克隆生长。将Lin-Sca-1⁺细胞在FL + IL-7中孵育12天后，细胞产量增加了近400倍。表型分析显示，所产生的细胞中超过99%为B细胞系，因为它们表达B220，但不表达髓系、红系或T细胞系特异性的细胞表面标志物。此外，这些细胞不表达细胞质μ重链（cmu）或表面IgM，但CD24（热稳定抗原[HSA]）和CD43（白细胞唾液酸蛋白）呈阳性，表明所产生的细胞在晚期前B细胞阶段受阻。有趣的是，尽管所有对FL + IL-7有反应的Lin-Sca-1⁺祖细胞及产生的前B细胞均表达c-kit，但在刺激B细胞系细胞生成方面，FL + IL-7比SCF + IL-7的效力要强得多（62倍）。此外，虽然FL + IL-7选择性地刺激前B细胞的生成，但SCF + IL-7主要刺激成熟粒细胞的生成。传代培养研究表明，对FL + IL-7有反应的Lin-Sca-1⁺祖细胞并未定向分化为B细胞系，因为在FL + IL-7中孵育2天后，80%的祖细胞仍保留髓系潜能。在FL + IL-7中孵育7天后，多达27%对FL + IL-7有反应的祖细胞仍未定向分化，但在FL + IL-7中孵育10天后均已定向分化为B细胞系。这些结果表明，与SCF相比，FL与IL-7协同作用时能更有效且更有选择性地增强未定向祖细胞向B细胞的定向分化和发育。
