Abstract 1533: β-arrestins Regulate Proliferation and Migration of Smooth Muscle Cells in vitro & in vivo: Isoform-specific Roles
Seven-membrane-spanning receptors (7TMRs) contribute significantly to vascular smooth muscle cell (SMC) migration, proliferation, and survival. β-arrestin1 (βarr1) and -2 are adaptor proteins that desensitize 7TMR/G protein coupling while evoking distinct 7TMR signaling pathways, such as the extracellular signal-regulated kinase (ERK) cascade. We therefore tested the hypothesis that 7TMR-evoked, βarr-dependent ERK activation is important for SMC migration and DNA synthesis. Aortic primary SMC lines from congenic WT, βarr10 and βarr20 mice (3 lines/group) showed equivalent expression of 7TM lysophosphatidic acid (LPA), S1P, and PAR receptors, and similar LPA-mediated Ca+2 influx and PDGF-induced ERK activation. However, while the ERK response to LPA was 200±40% of WT in βarr10 SMCs, it was only 60±10% of WT in βarr20 SMCs (P < 0.05). Similarly, thrombin- and S1P-induced ERK activation were reduced by 45±9% and 36±8%, respectively, in βarr20 SMCs (P < 0.05). SMC migration (Transwell™) and [3H]thymidine incorporation in WT and βarr20 SMCs were equivalent in response to PDGF (2.6±0.2 and 2.2±0.2 fold/basal, resp.), but not to LPA: WT SMC responses were 1.7±0.2 and 1.9±0.3 fold/basal, respectively (P < 0.05), yet βarr20 SMC responses were undetectable. Thus, while βarr isoforms had no effect on PDGF-induced SMC activity, they had opposing effects on 7TMR-evoked signaling. To test SMC proliferation and migration in vivo, we provoked neointimal hyperplasia in our congenic mice by de-endothelializing the common carotid artery (0.35 mm wire; ≥4 mice/group, matched for age and gender). In WT mice, neointimal area (SMC actin-positive) was 16±2 and 28±2 μm2 (×103) at 2 and 4 wk post-injury, respectively; corresponding luminal stenosis was 35% and 65%. In contrast, neointimal hyperplasia was reduced in βarr20 mice (by 66% and 37% at 2 and 4 weeks, resp., P < 0.05), and increased in βarr10 mice (by 84% at 4 wk, P < 0.05). The ratio of proliferating cell nuclear antigen immunofluorescence/DNA fluorescence in 4-wk-injured carotid arteries was βarr10 > WT > βarr20, consistent with SMC DNA synthetic responses to 7TMR mitogens. We conclude that 7TMRs contribute to neointimal hyperplasia in a manner reciprocally regulated by βarr2 (enhancement) and βarr1 (inhibition).