Vischer P, Völker W, Schmidt A, Sinclair N
Institut für Arterioskleroseforschung, Universität, Münster/Bundesrepublik Deutschland.
Eur J Cell Biol. 1988 Oct;47(1):36-46.
Different biochemical and cytochemical techniques were applied to characterize the sites of localization of thrombospondin in cultured endothelial cells. The results obtained by [35S]methionine labeling, immunoblotting, immunoprecipitation, fluorescence microscopy, ultracytochemistry, immunogold labeling, and silver enhancement experiments revealed that thrombospondin secreted by endothelial cells is structurally organized together with proteoheparan sulfate in spherical granules at the cell surface. These granules are about 100 to 300 nm in size. Heparin or enzymatic degradation with heparitinase, but not with ABC lyase, release thrombospondin from the cell surface. Fibronectin is expressed in the extracellular matrix of endothelial cells in a fibrillar organization, clearly distinct from the punctate pattern of thrombospondin on the cell surface. Furthermore, secreted thrombospondin is highly enriched together with fibronectin and proteoheparan sulfate in cell attachment sites and in cell migration tracks. In cell migration tracks proteoheparan sulfate more clearly resembles the fibrillar distribution pattern of fibronectin, whereas thrombospondin reveals a rather monodisperse pattern. The obtained data suggest preferential sites of interaction between thrombospondin and heparan sulfate proteoglycans on the cell surface and a participation of thrombospondin in cell adhesion and cell migration.
应用不同的生化和细胞化学技术来表征血小板反应蛋白在培养的内皮细胞中的定位位点。通过[35S]甲硫氨酸标记、免疫印迹、免疫沉淀、荧光显微镜、超微细胞化学、免疫金标记和银增强实验获得的结果表明，内皮细胞分泌的血小板反应蛋白与硫酸乙酰肝素蛋白聚糖一起在细胞表面的球形颗粒中进行结构组装。这些颗粒大小约为100至300纳米。肝素或用乙酰肝素酶而非ABC裂解酶进行酶促降解可从细胞表面释放血小板反应蛋白。纤连蛋白以纤维状结构在内皮细胞的细胞外基质中表达，与细胞表面血小板反应蛋白的点状模式明显不同。此外，分泌的血小板反应蛋白与纤连蛋白和硫酸乙酰肝素蛋白聚糖在细胞附着位点和细胞迁移轨迹中高度富集。在细胞迁移轨迹中，硫酸乙酰肝素蛋白聚糖更明显地类似于纤连蛋白的纤维状分布模式，而血小板反应蛋白则呈现出相当分散的模式。所获得的数据表明血小板反应蛋白与细胞表面硫酸乙酰肝素蛋白聚糖之间优先的相互作用位点，以及血小板反应蛋白参与细胞黏附和细胞迁移。
