Takagi J, Sudo Y, Saito T, Saito Y
Department of Biological Sciences, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
Eur J Biochem. 1994 Jun 15;222(3):861-7. doi: 10.1111/j.1432-1033.1994.tb18933.x.
Cell-adhesion activity of the bovine propolypeptide of von Willebrand factor (pp-vWF) was assessed by means of an in vitro assay with several cell lines of both normal and tumor-cell origin. pp-vWF promoted adhesion and spreading of B16 mouse melanoma cells and G-361 human melanoma cells. However, it could not induce adhesion of any other cell lines tested including endothelial cells, normal fibroblasts, and tumor cells of sarcoma, carcinoma, neuroblastoma and leukemia origin. A monospecific polyclonal antibody against pp-vWF, but not against fibronectin, laminin, and von Willebrand factor (vWF), completely blocked the pp-vWF-mediated adhesion, indicating that the cell adhesion was due to the pp-vWF molecule and not due to possible contamination of these three well-known adhesive proteins. The cell-adhesion activity was also observed with human pp-vWF and, furthermore, the adhesion to both bovine and human pp-vWF was not affected by a peptide containing the Arg-Gly-Asp sequence while the peptide abolished the cell adhesion to vWF. The adhesion was completely dependent on Mg2+ and inhibited by Ca2+. Inhibition by an anti-(beta 1 integrin) mAb (4B4) indicates that the receptor for this protein belongs to the beta 1-integrin family. A monoclonal antibody (TC4) among several antibodies directed against bovine pp-vWF inhibited the B16 adhesion to immobilized pp-vWF. The epitope for this monoclonal antibody lies in a central 8-kDa portion of pp-vWF, suggesting that this region is important for the cell-adhesion activity. This idea was supported by the finding that purified 8-kDa fragment promoted adhesion of B16 cells in a concentration-dependent manner. As pp-vWF shows unique cell-type specificity in its adhesion activity, which is completely different from that of fibronectin, laminin, vWF and collagen, it may be a novel type of adhesive glycoprotein that utilizes a beta 1-integrin receptor.
通过对正常细胞系和肿瘤细胞系的几种细胞系进行体外试验，评估了牛血管性血友病因子原多肽（pp-vWF）的细胞黏附活性。pp-vWF促进了B16小鼠黑色素瘤细胞和G-361人黑色素瘤细胞的黏附与铺展。然而，它不能诱导包括内皮细胞、正常成纤维细胞以及肉瘤、癌、神经母细胞瘤和白血病来源的肿瘤细胞在内的任何其他受试细胞系的黏附。一种针对pp-vWF而非纤连蛋白、层粘连蛋白和血管性血友病因子（vWF）的单特异性多克隆抗体完全阻断了pp-vWF介导的黏附，这表明细胞黏附是由于pp-vWF分子，而非这三种知名黏附蛋白可能的污染。人pp-vWF也观察到了细胞黏附活性，此外，对牛和人pp-vWF的黏附不受含精氨酸-甘氨酸-天冬氨酸序列的肽的影响，而该肽消除了细胞对vWF的黏附。黏附完全依赖于Mg2+并受到Ca2+的抑制。抗（β1整合素）单克隆抗体（4B4）的抑制表明该蛋白的受体属于β1整合素家族。在几种针对牛pp-vWF的抗体中，一种单克隆抗体（TC4）抑制了B16细胞对固定化pp-vWF的黏附。该单克隆抗体的表位位于pp-vWF的中央8 kDa部分，这表明该区域对细胞黏附活性很重要。纯化的8 kDa片段以浓度依赖的方式促进B16细胞黏附这一发现支持了这一观点。由于pp-vWF在其黏附活性方面表现出独特的细胞类型特异性，这与纤连蛋白、层粘连蛋白、vWF和胶原蛋白完全不同，它可能是一种利用β1整合素受体的新型黏附糖蛋白。
