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S-9-7 Regulation of proteoglycan synthesis mediated by thrombin receptor in human coronary endothelial cells in culture
1T. Kaji, 1C. Yamamoto, 2M.G. Kinsella and 2T. N. Wight 1Department of Environmental Health, Faculty of Pharmaceutical Sciences,@Hokuriku University, Kanazawa, Japan, 2Department of Pathology, School of Medicine, University of Washington, Seattle, USA

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@Thrombin is not only a key enzyme of the blood coagulation-fibrinolytic system but also influences endothelial cell functions. To address whether thrombin receptor mediates the regulation of proteoglycan synthesis by vascular endothelial cells or not human coronary endothelial cells were treated with thrombin receptor agonist peptide (SFLLRN; TRAP) in the presence of [35S]sulfate or 35S-labeled amino acids. TRAP significantly increased the incorporation of [35S]sulfate into proteoglycans in the medium but not in the cell layer regardless of the cell density. DEAE-Sephacel ion exchange chromatography separated the radiolabeled proteoglycans synthesized in the presence of TRAP into a single peak of high charge density. Characterization of GAGs present in this peak revealed the presence of chondroitin/dermatan sulfate proteoglycans (CS/DSPGs). TRAP treatment altered the GAG length of these proteoglycans from M r 48,000 to M r 51,000 and 33,000. SDS-polyacrylamide gel electrophoresis of core proteins labeled with 35S-labeled amino acids indicated that TRAP increased CS/DSPG core proteins with a M r of approximately 50 kDa in the medium. Western blot analysis revealed the presence of biglycan core protein in the controls cultures whereas both biglycan and decorin core proteins were found in the TRAP treated cultures. Quantitative RT-PCR showed that TRAP treatment significantly increased biglycan and decorin mRNAs. The present data suggest that thrombin receptor activation results in elevated synthesis of biglycan that bears CS/DS chains of approximately M r 50,000 and in induction of the synthesis of decorin that bears CS/DS chains of approximately M r 30,000 in vascular endothelial cells.

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