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The establishment of humanmarrow stromal cell lines and their effects on the proliferation and differentiation of hematopoietic stem cells.

人骨髓基质细胞系的建立及其对造血干细胞增殖和分化的影响。

基本信息

批准号：
59480153
负责人：
HARIGAYA Kenichi
金额：
$ 4.1万
依托单位：
Keio University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1984
资助国家：
日本
起止时间：
1984 至 1986
项目状态：
已结题

项目摘要

Five functional clonal cell lines were established from the adherent cell populations in long-term liquid cultures after transfection with the recombinant plasmid pSV3gpt. The radioimmunoassay and northern blot analysis revealed that these cell lines produced granulopoietic colony-stimulating factor (G-CSF) and macrophage colony-stimulating factor (M-CSF). The conditioned medium from the cell line KM-102 contained a factor which stimulated the proliferation and differentiation of multipotent stem cells. This factor was purified 340000 folds by five sequential fractionations and gave a single broad band of protein with a molecular weight of approximately 18000 dalton. This purified factor was associated with burst promoting activity (BPA) in vitro and showed is a highest specific activity of BPA among the substances which have been reported so far. The N-terminal amino acid sequence of this factor was analogous to the one deduced from cDNA of human GM-CSF. A part of human hemic cell lin … More es, HL-60 and KG-1, attached to the marrow stromal cell lines. Furthermore, the proliferation and differentiation of the attached hemic cells were strongly inhibited even when the cocultures were supplemented with differentiation inducing agent, 1 <alpha> , <25(OH)_2> vitamin <D_3> . These attachment is specific between myeloid cells and marrow stromal cells since there was no binding between myeloid cell lines and stromal cells from human thymus and lymph nodes, and between T or B lymphocytic cell lines and the marrow stromal cells. These indicate that direct cell to cell interaction is present between myeloid hemic cells and marrow stromal cells. Gap junctional communication was frequently observed in primary marrow stromal cells and stromal cell lines, and sometimes between stromal cells and the attached round hemic cells. This suggests that direct communication of low molecular weight substances is present between these cells. Nevertheless, the nature of the communication has not been clarified. Less

用重组质粒pSV 3gpt转染长期液体培养的贴壁细胞群，建立了5个功能性克隆细胞系。放射免疫分析和北方印迹分析表明，这些细胞株产生粒细胞集落刺激因子（G-CSF）和巨噬细胞集落刺激因子（M-CSF）。KM-102细胞系的条件培养基含有刺激多能干细胞增殖和分化的因子。该因子通过5次连续分级纯化340000倍，得到分子量约为18000道尔顿的单一宽带蛋白。这种纯化的因子与体外爆发促进活性（BPA）有关，并且在迄今为止已报道的物质中显示出BPA的最高比活性。该因子的N-末端氨基酸序列与人GM-CSF的cDNA序列相似。人血细胞系的一部分 ...更多信息 es，HL-60和KG-1，附着于骨髓基质细胞系。此外，即使在共培养物中添加分化诱导剂1，<25（OH）_2>维生素，贴壁血细胞的增殖和分化也受到强烈抑制<alpha><D_3>。这种附着在骨髓细胞和骨髓基质细胞之间是特异性的，因为骨髓细胞系和来自人胸腺和淋巴结的基质细胞之间以及T或B淋巴细胞系和骨髓基质细胞之间没有结合。这表明髓系造血细胞和骨髓基质细胞之间存在直接的细胞间相互作用。在原代骨髓基质细胞和基质细胞系中经常观察到缝隙连接通讯，有时基质细胞和附着的圆形血细胞之间也观察到缝隙连接通讯。这表明这些细胞之间存在低分子量物质的直接交流。然而，来文的性质尚未得到澄清。少