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Purification and Functional Analysis of Adfesion Related Molecules Between Hematopoietic Stem Cells and Marrow Stromal Cells.

造血干细胞与骨髓基质细胞粘附相关分子的纯化和功能分析。

基本信息

批准号：
63480141
负责人：
HARIGAYA Kenichi
金额：
$ 4.16万
依托单位：
Chiba 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：
1988
资助国家：
日本
起止时间：
1988 至 1990
项目状态：
已结题

项目摘要

In order to investigate novel adhesion molecule (s) in stromal hematopoiesis, we raised monoclonal antibodies (MAb) against human marrow stromal cell lines KM-101 and KM-102 and myeloid hematopoietic cell line (CMK). The antibodies were screened by the direct addition into coculture plates containing KM cell line and K562. One of the antibodies enhanced the attachment between K562 and stromal cell lines. This antibody recognized the 135-Kilodalton membrane molecules and its Fab fragment markedly inhibited the adhesion between stromal cells and K562 cells. Immunohistochemical staining of this Mab revealed positive reaction in stromal cells of wide variety of tissues and vascular endothelia, smooth muscle cells, and some of epithelial cells including renal collecting tubules, colon and pancreatic duct. No positive reaction was found in neural cells or skeletal muscles. The distribution of the molecules are similar to CD44 antigen with a few exception, but the size of the molecules are much different. Direct binding of ^<125>I on the surface membrane of various kind of blood cells showed that peripheral white blood cells had low radiolabelling while relatively immature hematopoieticcell lines incorporated twice or more radioactive antibodies. The amount of the direct binding was modulated when the hematopoietic cell lines were treated with 10ng/ml TPA 3 days in advance. These results indicates that some sosts of exogenous stimuli modulate the expression of 135 K dalton membrane molecule. Taking together with the distribution of the molecules among a wide varieties of stroma and parenchyma, the molecules recognized by Mab C-F-9 may be one of signaling molecules between parenchymal and stromal signal transduction, as well as an adhesion related protein.

为了研究基质造血中的新型粘附分子，我们制备了针对人骨髓基质细胞系 KM-101 和 KM-102 以及骨髓造血细胞系 (CMK) 的单克隆抗体 (MAb)。通过直接添加到含有KM细胞系和K562的共培养板中来筛选抗体。其中一种抗体增强了 K562 和基质细胞系之间的附着。该抗体可识别 135 千道尔顿的膜分子，其 Fab 片段显着抑制基质细胞与 K562 细胞之间的粘附。该单克隆抗体的免疫组织化学染色显示多种组织和血管内皮的基质细胞、平滑肌细胞以及包括肾集合管、结肠和胰管在内的一些上皮细胞呈阳性反应。神经细胞或骨骼肌中未发现阳性反应。除少数例外，分子分布与CD44抗原相似，但分子大小有很大不同。 125 I在各种血细胞表面膜上的直接结合表明外周白细胞具有低放射性标记，而相对不成熟的造血细胞系掺入了两次或更多次放射性抗体。当提前3天用10ng/ml TPA处理造血细胞系时，调节直接结合的量。这些结果表明一些外源刺激调节135K道尔顿膜分子的表达。结合该分子在多种间质和实质中的分布，Mab C-F-9识别的分子可能是实质和间质信号转导之间的信号分子之一，也是粘附相关蛋白。