Protein Kinase C Delta Promotes Adventitial Cell Migration to Neointima by Upregulation of Monocyte Chemoattractant Protein-1 in Smooth Muscle Cells

Abstract

Introduction: Accumulating evidence suggests that adventitial cells directly contribute to neointima formation by migrating into the intima. We have previously reported that gene transfer of Protein kinase C delta(PKC δ) attenuates intimal hyperplasia by inducing apoptosis of smooth muscle cells (SMCs). More recently, we showed that PKCδ mediates expression of monocyte chemoattractant protein-1(MCP-1). in the current study, we tested the hypothesis that PKCδ promotes adventitial cell migration by stimulating SMCs to produce chemokines such as MCP-1. Methods: In vitro migration of isolated adventitial cells was evaluated by chemotaxis assay. Gene transfer to SMCs was achieved by intraluminal perfusion with adenoviruses expressing PKCδ(AdPKCδ) or empty vector (AdNull) following rat carotid angioplasty. Adventitia cells were labeled by administration of AdLacZ via pluronic gel on the adventitial surface prior to wound closure. Results: Media conditioned by PKCδ-overexpressing SMCs stimulated migration of adventitial cells, which was blocked by quenching MCP-1 with a neutralizing antibody or by knocking down the MCP-1 receptor, CCR2. Similarly, gene transfer of PKCδ to injury arteries stimulated adventitial cell migration in vivo. Fourteen days following injury, accumulation of adventitial cells in the neointima was close to 10 times higher in AdPKCδ-infected arteries than in AdNull-infected arteries(LacZ positive area in pixel:769±67*103 vs 84±11*103, p<0.01). In addition, PKCδ gene transfer increased MCP-1 expression in the media by ∼4 folds (MCP-1 positive areas in pixel:397±20*103 vs 99±18*103, p<0.05). to further prove that PKCδ mediates migration of adventitia cells through MCP-1, we randomly divided AdPKCδ-treated rats into two groups, one injected with a neutralization antibody to MCP-1 and the other with nonimmunized IgG. Neutralizing MCP-1 significantly reduced the number of adventitial cells accumulated in the neointima (LacZ positive area in pixel:357±37*103 vs 894±42*103, p<0.01). Furthermore, combining PKCδ gene transfer and anti-MCP-1 treatment inhibited I/M ratio by 62% as compared to a 36.3% and 30.8% reduction produced by a single treatment of PKCδ and anti-MCP-1, respectively. None of the treatments altered macrophage infiltration, at least at the time points (7 and 14 days) we assessed. Conclusions: PKCδ appears to play dual functions during arterial injury response. in addition to inducing apoptosis, PKCδ promotes migration of adventitial cells by stimulating MCP-1 expression in SMCs. A combined strategy targeting both SMC apoptosis and chemokine production may be an efficient approach to inhibit intimal hyperplasia.