CULTURED HUMAN-FIBROBLASTS SYNTHESIZE AND SECRETE THROMBOSPONDIN AND INCORPORATE IT INTO EXTRACELLULAR-MATRIX
CULTURED HUMAN-FIBROBLASTS SYNTHESIZE AND SECRETE THROMBOSPONDIN AND INCORPORATE IT INTO EXTRACELLULAR-MATRIX
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DOI：
10.1073/pnas.80.4.998
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发表时间：
1983-01-01
期刊：
影响因子：
--
通讯作者：
MOSHER, DF
中科院分区：
文献类型：
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作者：
JAFFE, EA;RUGGIERO, JT;MOSHER, DF
血小板反应蛋白，一种主要的从α释放的糖蛋白。凝血酶刺激的血小板颗粒，是160千道尔顿亚基的二硫键三聚体。培养的人包皮和胎儿肺成纤维细胞以时间依赖性方式（分别为15.7和5.8 μ g/106个细胞/24小时）将血小板反应蛋白（通过酶联免疫吸附测定法测定）分泌到培养基中;分泌被放线菌酮阻断。从[3 H]亮氨酸标记的成纤维细胞培养后培养基中分离[3 H]血小板反应蛋白，并从具有与葡萄球菌蛋白A-Sepharose偶联的兔多克隆或小鼠单克隆抗血小板反应蛋白的细胞层中分离[3 H]血小板反应蛋白。免疫分离的[3 H]血小板反应蛋白迁移在十二烷基硫酸钠/聚丙烯酰胺凝胶与纯化的标记血小板血小板反应蛋白，无论有没有减少。使用兔多克隆和小鼠单克隆抗血小板反应蛋白抗体的免疫荧光显微镜将血小板反应蛋白定位于细胞周围的纤维状细胞外基质。因此，培养的人成纤维细胞分泌血小板反应蛋白并将其掺入细胞外基质中。
Thrombospondin, a major glycoprotein released from .alpha. granules of thrombin-stimulated platelets, is a disulfide-bonded trimer of 160-kilodalton subunits. Cultured human foreskin and fetal lung fibroblasts secreted thrombospondin (determined by enzyme-linked immunosorbent assay) into the culture medium in a time-dependent manner (15.7 and 5.8 .mu.g/106 cells per 24 h, respectively); secretion was blocked by cycloheximide. [3H]Thrombospondin was isolated from [3H]leucine-labeled fibroblast postculture medium and from cell layers with rabbit polyclonal or mouse monoclonal anti-thrombospondin coupled to staphylococcal protein A-Sepharose. The immunologically isolated [3H]thrombospondin migrated in sodium dodecyl sulfate/polyacrylamide gels with purified marker platelet thrombospondin both with and without reduction. Immunofluorescence microscopy using rabbit polyclonal and mouse monoclonal anti-thrombospondin antibodies localized thrombospondin to the fibrillar extracellular matrix surrounding the cells. Thus, cultured human fibroblasts secrete thrombospondin and incorporate it into the extracellular matrix.