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

重组质粒pSV3gpt转染后，从长期液体培养的贴壁细胞群中建立了5株功能克隆细胞系。放射免疫分析和northern blot分析显示，这些细胞系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）。从KM-102细胞系提取的条件培养基中含有刺激多能干细胞增殖和分化的因子。该因子经5次连续分离纯化340000倍，得到分子量约为18000道尔顿的单一宽带蛋白。该纯化因子与体外促爆活性（BPA）相关，是目前报道的物质中BPA比活性最高的。该因子的n端氨基酸序列与人GM-CSF的cDNA序列相似。人造血细胞系的一部分，如HL-60和KG-1，附着于骨髓基质细胞系。此外，在共培养中添加1 < α - >, <25(OH)_2>，维生素<D_3>的诱导分化剂，对附着的血细胞的增殖和分化也有明显的抑制作用。骨髓细胞和骨髓基质细胞之间的这种附着是特异性的，因为骨髓细胞系和来自人类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。这表明骨髓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间隙连接通讯，有时在基质细胞和附着的圆形血细胞之间也观察到间隙连接通讯。这表明在这些细胞之间存在低分子量物质的直接通讯。然而，来文的性质尚未得到澄清。少

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