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

将重组表达载体pSV3gpt导入长期液体培养的贴壁细胞，建立了5个功能克隆细胞系。放射免疫分析和Northern印迹分析表明，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)。KM-102细胞的条件培养液中含有一种刺激多能干细胞增殖和分化的因子。该因子经5次连续分级提纯340000倍，得到一条分子量约为18000道尔顿的单一宽带。该纯化因子在体外具有促爆裂活性(BPA)，是迄今报道的BPA中比活性最高的物质。该因子的N端氨基酸序列与人GM-CSF基因的N端氨基酸序列相似。人白血病细胞LIN-…的一部分更多的ES，HL-60和KG-1附着在骨髓基质细胞系上。联合培养中加入分化诱导剂1-lt；α-gt；、-lt；25(OH)_2&gt；维生素&lt；D_3&gt；对贴壁细胞的增殖和分化也有明显抑制作用。由于髓系细胞系与人胸腺和淋巴结的基质细胞之间、T或B淋巴细胞系与骨髓基质细胞系之间没有结合，因此髓系细胞与骨髓基质细胞之间的这种附着是特异的。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。缝隙连接通讯常见于原代骨髓基质细胞和基质细胞系，有时在基质细胞和附着的圆形血球细胞之间。这表明这些细胞之间存在着低分子物质的直接联系。然而，来文的性质还没有得到澄清。较少

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