Abstract 13444: G Protein-coupled Receptor Kinase-5 Attenuates Atherosclerosis by Desensitizing Monocyte/Macrophage Responses to Cytokines and by Enhancing Smooth Muscle Cell PDGF Receptor Ubiquitination
G protein-coupled receptor kinase-5 (GRK5) is a Ser/Thr kinase that up-regulates in atherosclerotic arteries; it phosphorylates and desensitizes many receptors important to atherogenesis. We showed previously that Apoe−/−/Grk5−/− mice develop worse atherosclerosis than congenic Apoe−/− mice. Since arterial wall and bone marrow-derived cells contribute to atherogenesis, we tested the hypothesis that GRK5 activity is anti-atherogenic in monocytes, macrophages (MPs), and arterial wall cells. To test this hypothesis in arterial wall cells, we transplanted carotid arteries from Grk5−/− and WT mice into congenic Apoe−/− mice as carotid interposition grafts. After 6 weeks, Grk5−/− grafts had 45% more neointimal area (100±2 vs 48±6 μm2 × 103, p<0.01), comprising MPs and smooth muscle cells (SMCs), and 175% higher neointimal SMC prevalence (p<0.05) than WT grafts. Two wk post-op, before intimal MP infiltration, quantitative immunofluorescence yielded the following ratios for Grk5−/−/WT carotid grafts: activated PDGF receptor-β (pY1021-PDGFRβ), 2.0±0.1; ICAM-1, 1.5±0.1; VCAM-1, 2.0±0.2; MCP-1, 1.8±0.1; PCNA, 1.2±0.1 (p<0.05 for each). Concordantly, PDGFRβ ubiquitination and degradation were 2.6±0.5-fold greater in WT than in Grk5−/− SMCs, assessed by IP/IB (p=0.01). To test anti-atherogenic GRK5 activity in monocytes/MPs, these cells were isolated from bone marrow by negative selection with magnetic beads (yielding 84±4% monocytes by flow cytometry, CD11bhi/lineagelow) or by culture with CSF-1 (yielding 100% Mac3+ cells), respectively. Monocyte migration (Neuroprobe plates) was greater in Grk5−/− than WT monocytes in response to MCP-1, CSF-1, and macrophage-conditioned medium (by 200±30%, 210±30% and 200±6%, respectively) (p<0.05), but not in response to phorbol ester. After stimulation with LPS or TNF, Grk5−/− and WT MPs demonstrated equivalent rates of IκBα phosphorylation and degradation; however, there was 50% greater ERK activation in Grk5−/− than in WT MPs (p<0.02). We conclude that GRK5 attenuates atherosclerosis, at least in part, by enhancing the ubiquitination and degradation of PDGFRβ in SMCs, decreasing TNFR1 and TLR4-promoted ERK activation in MPs, and by decreasing monocyte migration toward atherogenic chemokines.
- © 2010 by American Heart Association, Inc.