Collagen synthesis by bovine aortic endothelial cells in culture

Publisher Website
Google Scholar
Abstract
Endothelial cells isolated from bovine aorta synthesize and secrete type III procollagen in culture. The procollagen, which represents the major collagenous protein in culture medium, was specifically precipitated by antibodies to bovine type III procollagen and was purified by DEAE cellulose chromatography. Unequivocal identification of the pepsin-treated collagen was made by direct comparison with type III collagen isolated by pepsin digestion of bovine, skin, utilizing peptide cleavage patterns generated by vertebrate collagenase, CNBr and mast cell protease. The type III collagen was hydroxylated to a high degree, having a hydroxyproline/proline ratio of 1.5:1.0. Pulse-chase studies indicated that the procollagen was not processed to procollagen intermediates or to collagen. Pepsin treatment of cell layers, followed by salt fractionation at acidic and neutral pH, produced several components which were sensitive to bacterial collagenase and which comigrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with .alpha.A, .alpha.B and type IV collagen chains purified from human placenta by similar techniques. Bovine aortic endothelial cells also secreted fibronectin and a bacterial collagenase-insensitive glycoprotein which, after reduction, had a MW of 135,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (using procollagen MW standards) and which was not precipitable by antibodies to cold-insoluble globulin or to .alpha.2-macroglobulin. Collagen biosynthesis by these cells provides and interesting model system for studying the polarity of protein secretion and the attachment of cells to an extracellular matrix. The presence of type III collagen in the subendothelium and the specific interaction of this protein with fibronectin and platelets suggest the involvement of this collagen in thrombus formation following endothelial cell injury.