Abstract 12299: ROCK2 in Vascular Smooth Muscle Cells Plays a Crucial Role for Hypoxia-Induced Pulmonary Hypertension in Mice
Background: We have previously demonstrated that the Rho-kinase (ROCK) pathway in vascular smooth muscle cells (VSMC) plays an important role in the pathogenesis of cardiovascular diseases by enhancing VSMC migration and proliferation. Rho-kinase has 2 isoforms, ROCK1 and ROCK2, with different functions in different cells; ROCK1 for circulating inflammatory cells and ROCK2 for the vasculature. In this study, we thus examined whether ROCK2 in VSMC is involved in the pathogenesis of PH.
Methods and Results: In patients with PH as compared with controls, the immunoreactivity of ROCK2 and phosphorylated ERK1/2 co-expressed with αSMA in pulmonary arterial media were markedly increased. Thus, we developed VSMC-specific ROCK2-deficient (ROCK2+/-) and VSMC-specific ROCK2-overexpressing (ROCK2-Tg) mice. The extent of hypoxia-induced PH (10% O2 for 4 weeks), as evaluated by pulmonary vascular remodeling, right ventricular (RV) systolic pressure and RV hypertrophy, was significantly reduced in ROCK2+/- mice and was enhanced in ROCK2-Tg mice compared with the littermates (n=10 each, all P<0.05). The expression of ROCK activity and phosphorylated ERK1/2 in lung tissues analyzed by Western blotting and the numbers of Ki67+ proliferating VSMC and CD45+ inflammatory cells in pulmonary arterioles revealed by immunostaining were significantly reduced in the ROCK2+/- mice and were increased in the ROCK2-Tg mice compared with the littermates (n=4 each, all P<0.05). In cultured murine aortic VSMC, both scratch assay and Boyden chamber assay showed that migration activity was significantly reduced in the ROCK2+/- VSMC and was increased in the ROCK2-Tg VSMC compared with littermates (n=5 each, both P<0.01). In addition, in primary pulmonary arterial VSMC (PAVSMC) cultured from patients with PH, siRNA knockdown (-81%) of ROCK2 expression (ROCK2-siRNA) significantly suppressed the expression of ROCK activity and PDGF-induced phosphorylated ERK1/2 (n=4 each, both P<0.05). The migration activity evaluated by scratch assay was significantly reduced in the ROCK2-siRNA PAVSMC compared with the control-siRNA PAVSMC (P<0.01).
Conclusions: ROCK2 in VSMC plays a crucial role in the pathogenesis of PH through enhanced VSMC migration and proliferation in mice in vivo.
- © 2012 by American Heart Association, Inc.