Abstract 1177: TGF-beta, Through Smad3 Signaling, Stimulates Bone Marrow Progenitor Cell Migration and Recruitment To Sites of Vascular Injury
Background: Transforming growth factor beta (TGF-β) is believed to play an essential role in the development of intimal hyperplasia after vascular injury. We have previously shown that overexpression of Smad3, a downstream signaling molecule for TGF-β, in medial smooth muscle cells (SMC) results in an enhanced neointimal lesion in the rat carotid balloon injury model. The purpose of this study was to examine the effect of TGF-β and Smad3 on bone marrow progenitor cell recruitment.
Methods/Results: Male Sprague-Dawley rats underwent carotid balloon angioplasty followed by adenovirus mediated gene transfer of either Smad3 (n = 3) or empty virus (n = 3). Using c-kit expression as a progenitor cell marker, we found that Smad3 overexpression resulted in an increased percentage of c-kit positive cells in the arterial wall 5 days after injury (36.1 ± 2.4% vs. 23.7 ± 2.6%, p < 0.05). This suggested that Smad3 and TGF-β may stimulate progenitor cell recruitment to the site of injury. Chemotaxis assays revealed that conditioned media from rat SMCs infected with adenovirus expressing Smad3 and stimulated with TGF-β significantly increased rat bone marrow cell and human CD34+ cell migration up to 6-fold compared to media from control cells. Notably, TGF-β itself had no direct effect on bone marrow or CD34+ cell migration, suggesting the presence of SMC secreted factors that were induced by the TGF-β/Smad3 pathway. A microarray analysis comparing gene expression in SMCs with or without activation of the TGF-β/Smad3 pathway revealed that monocyte-chemoattractant protein-1 (MCP-1) gene expression was induced over 80-fold over control. This was confirmed by reverse transcriptase PCR and MCP-1 ELISA. To further support the role of MCP-1 in progenitor cell recruitment, recombinant MCP-1 was found to induce rat bone marrow cell and human CD34+ cell migration. Conversely, blockade of MCP-1 expression or activity in SMC-conditioned media using an siRNA to Smad3 or MCP-1 or an MCP-1 blocking antibody inhibited bone marrow cell migration.
Conclusion: Our data suggest that TGF-β and Smad3 stimulate bone marrow progenitor cell migration and recruitment to sites of vascular injury and that this may be mediated by TGF-β and Smad3 induced MCP-1 expression in medial smooth muscle cells.