Thrombin causes vascular endothelial growth factor expression in vascular smooth muscle cells: role of reactive oxygen species.

Vascular endothelial growth factor (VEGF) has been implicated in the reendothelialization of the vascular wall after balloon injury. This study investigated whether thrombin, which is formed during activation of the coagulation cascade at sites of vascular injury, upregulates VEGF expression in vascular smooth muscle cells (VSMCs). VEGF expression was assessed in native and cultured VSMCs by Northern blot analysis and reverse transcription-polymerase chain reaction and the release of VEGF protein by immunoassay. alpha-Thrombin time- and concentration-dependently increased VEGF mRNA levels, mainly that mRNA coding for the soluble splice variant VEGF(164/165), and stimulated the release of VEGF protein. These effects required the proteolytic activity of thrombin and were mimicked by a thrombin receptor activating-peptide. Upregulation of VEGF expression was also induced by conditioned medium from alpha-thrombin-stimulated VSMCs. Both the early and the delayed alpha-thrombin-induced VEGF expressions were attenuated by antioxidants and by diphenyleneiodonium. alpha-Thrombin-induced VEGF release was significantly reduced by a platelet-derived growth factor (PDGF)-, a transforming growth factor (TGF)-beta-, and a basic fibroblast growth factor (bFGF)-neutralizing antibody. Thrombin caused a redox-sensitive upregulation of expression of VEGF in VSMCs through a direct and an indirect effect, which was dependent on the endogenous formation of PDGF, TGF-beta, and bFGF. Upregulation of VEGF expression may represent an important mechanism by which the coagulation cascade contributes to the regeneration of the endothelial lining at sites of balloon injury.