Petursson S R, Chervenick P A
Department of Medicine, University of Pittsburgh School of Medicine, Pennsylvania 15261.
Exp Hematol. 1988 Sep;16(8):660-6.
The effect of partially purified thrombopoietic stimulatory factor (TSF) on megakaryocytopoiesis was studied using the soft-gel colony-forming assay and a short-term marrow liquid culture system (STLC) and compared to the effects of megakaryocyte colony-stimulating activity present in pokeweed mitogen-stimulated spleen cell-conditioned medium (PWCM). Nonadherent cells from STLC were sampled daily for acetylcholinesterase-positive cells and megakaryocyte progenitor cells (CFU-M). CFU-M were assayed in the soft-gel colony-forming system using PWCM as a source of colony-stimulating activity. Proliferative capacity of CFU-M obtained from liquid culture was determined from megakaryocyte colony size (number of megakaryocytes per colony) following plating of cells in a secondary colony-forming assay. Megakaryocytes were grouped into four maturation classes and megakaryocyte diameter was determined on acetylcholinesterase-stained cytocentrifuged cells using an eye-piece micrometer. TSF produced no CFU-M-derived colonies in the soft-gel colony-forming assay. Addition of TSF to STLC had no effect on the total number of CFU-M, megakaryocyte colony size, or total number of megakaryocytes compared to unstimulated STLC. However, on days 4-9 there was a significant increase in megakaryocyte diameter and the proportion of mature (stage III, IV) megakaryocytes obtained from TSF containing STLC compared to unstimulated STLC. In contrast, 5 days after addition of PWCM to STLC a sixfold increase in the total number of CFU-M per flask and a threefold increase in megakaryocytes was observed compared to unstimulated STLC. However, megakaryocyte colony size and megakaryocyte size were significantly reduced and a greater number of immature (stage I, II) megakaryocytes were present in STLC containing PWCM compared to unstimulated STLC. These results indicate that TSF accelerates the maturation of megakaryocytes in vitro and that a factor or factors present in spleen cell-conditioned medium, in addition to influencing megakaryocyte progenitor cell proliferation, also affect(s) megakaryocyte size.
采用软凝胶集落形成试验和短期骨髓液体培养系统（STLC），研究了部分纯化的血小板生成刺激因子（TSF）对巨核细胞生成的影响，并与商陆有丝分裂原刺激的脾细胞条件培养基（PWCM）中存在的巨核细胞集落刺激活性的作用进行了比较。每天从STLC的非贴壁细胞中取样，检测乙酰胆碱酯酶阳性细胞和巨核细胞祖细胞（CFU-M）。在软凝胶集落形成系统中，以PWCM作为集落刺激活性来源，对CFU-M进行检测。从液体培养中获得的CFU-M的增殖能力，通过在二次集落形成试验中接种细胞后巨核细胞集落大小（每个集落中的巨核细胞数量）来确定。将巨核细胞分为四个成熟类别，并使用目镜测微计在乙酰胆碱酯酶染色的细胞离心涂片上测定巨核细胞直径。在软凝胶集落形成试验中，TSF未产生CFU-M来源的集落。与未刺激的STLC相比，向STLC中添加TSF对CFU-M的总数、巨核细胞集落大小或巨核细胞总数没有影响。然而，在第4至9天，与未刺激的STLC相比，从含TSF的STLC中获得的巨核细胞直径和成熟（III期、IV期）巨核细胞的比例显著增加。相比之下，与未刺激的STLC相比，向STLC中添加PWCM 5天后，每个培养瓶中CFU-M的总数增加了六倍，巨核细胞增加了三倍。然而，与未刺激的STLC相比，含PWCM的STLC中巨核细胞集落大小和巨核细胞大小显著减小，并且存在更多未成熟（I期、II期）巨核细胞。这些结果表明，TSF在体外可加速巨核细胞的成熟，并且脾细胞条件培养基中存在的一种或多种因子，除了影响巨核细胞祖细胞增殖外，还会影响巨核细胞大小。
