Abstract 12619: EBP50 is Necessary for Intimal Hyperplasia following Angioplasty: Regulation of EGF Receptor Signaling and p21 Expression
Background: The Ezrin Binding Protein 50 (EBP50), a scaffolding protein regulates expression and activity of various membrane receptors. The role of EBP50 in the vasculature is poorly understood. Previous work showed that EBP50 expression increases after balloon injury in rat carotids, suggesting that it may contribute to neointima formation. The current study was designed to determine the role of EBP50 on vascular smooth muscle cells (VSMC) proliferation, an important contributing factor to arterial restenosis
Methods and Results: To determine the role of EBP50 on neointima formation, wire injury was performed in wild type (WT) and EBP50 knockout (KO) mice. As expected, two weeks after injury marked neointimal hyperplasia was observed in WT mice (neointima/media ratio: 3.11 ± 0.7, n=4). In striking contrast, virtually no arterial restenosis occurred in KO mice (ratio: 0.36 ± 0.16, n=3, p<0.05 vs. WT). Proliferation of KO VSMC in response to serum and EGF was significantly lower than WT cells, measured by cell number, 3H-Thyimidine and BrDU incorporation assays. In addition, overexpression of EBP50 increased cell proliferation whereas knockdown of EBP50 using siRNA attenuated VSMC proliferation. Cell cycle distribution, determined by flow cytometry, revealed cell cycle arrest in KO cells, which correlated with markedly increased expression and nuclear localization of the cyclin-dependent kinase inhibitor, p21. In addition, p21 knockdown by siRNA was sufficient to increase proliferation of KO VSMC. The regulation of p21 in VSMC was partly dependent on EGFR signaling. In KO VSMC, EGF receptor expression, determined by Western Blot, RT-PCR and immunostaining was dramatically reduced. EGF receptor signaling was also significantly reduced in KO, as indicated by decreased phosphorylation of AKT and ERK compared to WT VSMC. Interestingly, treatment of WT VSMC with the EGF receptor inhibitor AG1478 induced p21 accumulation indicating that EGF receptor signaling contributes to the regulation of p21 expression.
Conclusions: EBP50 is necessary for neointima formation. EBP50 regulates VSMC proliferation by controlling EGF receptor signaling and consequently p21 expression.
- © 2010 by American Heart Association, Inc.