Abstract 17947: Regulator of G-Protein Signaling 5 Prevents Smooth Muscle Cell Proliferation and Attenuates Neointima Formation

Abstract

Introduction: Regulator of G-protein signaling 5 (RGS5) is abundantly and specifically expressed in vascular smooth muscle cells (SMCs) and inhibits G-protein signaling by enhancing the GTP-hydrolyzing activity of Gα-subunits. Hypothesis: In the present study, we aimed to investigate the effects of RGS5 on vascular SMC function in vitro and neointima formation in vivo and to determine the underlying molecular mechanisms. Methods: Expression of RGS5 was assessed in vitro and in vivo by qPCR, immunoblotting and immunohistochemistry. Proliferation and migration of human coronary artery SMC were assessed in vitro following the adenoviral-mediated over expression of wild type (WT) RGS5, of a non-degradable mutant (C2A-RGS5) or siRNA-mediated knock down. In vivo, the proliferation and neointima formation were assessed in a wire-mediated injury model of the femoral artery in C57BL/6J mice. Results: We found a robust expression of RGS5 in native arteries of C57BL/6 mice and a significant down-regulation within neointimal lesions 10 and 21 days after vascular injury as assessed by qPCR, immunohistochemistry and immunoblotting. In vitro, RGS5 was found significantly down-regulated after mitogenic stimulation of SMCs. Overexpression of RGS5 or, even more prominently, the C2A-RGS5 mutant prevented SMC proliferation and migration. In contrast, the siRNA-mediated knockdown of RGS5 significantly augmented SMC proliferation. Following overexpression of RGS5, FACS analysis of propidium iodide-stained cells indicated cell cycle arrest in G0/G1 phase. Mechanistically, inhibition of ERK1/2 phosphorylation and MAPK downstream signaling was found to be responsible for the anti-proliferative effect since over expression of a constitutive active form of MEK reversed the growth-inhibitory effect of of RGS5. Following wire-induced injury, adenoviral-mediated overexpression of RGS5 or C2A-RGS5 significantly decreased ERK phosphorylation, SMC proliferation and neointima formation in vivo (n=8; p<0.001). Conclusions: Down-regulation of RGS5 is an important prerequisite for SMC proliferation and neointima Formation. Therefore, reconstitution of RGS5 levels might represent a promising therapeutic option to prevent vascular remodeling processes.