Abstract 13297: Rho-associated Kinase (ROCK) Mediates the Thrombin-Induced Production of Reactive Oxygen Species and Myosin Light Chain Phosphorylation-Independent Vasoconstriction in the Pulmonary Artery
Thrombotic pulmonary arteriopathy is a common pathological finding in primary pulmonary artery hypertension (PAH). Thrombin and its receptor PAR1 are thus suggested to play a critical role in the pathogenesis and pathophysiology of PAH. However, the contractile effect of thrombin in pulmonary artery and its mechanism still remains largely unknown. This study elucidated a novel role of ROCK in the thrombin-induced contraction by using the isolated porcine pulmonary artery. In the presence of extracellular Ca2+ ([Ca2+]o), thrombin induced a sustained contraction with an increase in [Ca2+]i and the phosphorylation of myosin light chain (MLC). The contractile effect of thrombin was abolished by PAR1 antagonist, while it was mimicked by PAR1 agonist peptide. In the absence of [Ca2+]o, thrombin induced a sustained contraction to a similar extent to that seen in the presence of [Ca2+]o, but with no increase in [Ca2+]i or MLC phosphorylation. A similar Ca2+-independent contraction was induced by H2O2, while N-acetyl cysteine substantially inhibited the Ca2+-independent contraction induced by thrombin and H2O2. The fluorescence imaging of reactive oxygen species (ROS) revealed that thrombin induced the production of ROS in the absence of [Ca2+]o. ROCK inhibitor inhibited not only the thrombin-induced contraction and MLC phosphorylation in the presence of [Ca2+]o, but also the Ca2+-independent contraction induced by thrombin and H2O2 in the absence of [Ca2+]o. ROCK inhibitor also inhibited the thrombin-induced ROS production. These suggest ROCK to function at both upstream and downstream of ROS. Thrombin increased the phosphorylation of MYPT1, a regulatory subunit of MLC phosphatase, at T853 and T696, in the presence of [Ca2+]o. In the absence of [Ca2+]o, thrombin and H2O2 preferentially increased the phosphorylation at T853. These observations indicate the activation of ROCK by thrombin. This study thus suggests that ROCK mediates not only the MLC phosphorylation-dependent contraction as previously reported, but also the production of ROS and the ROS-mediated, MLC phosphorylation-independent contraction. PAR1-ROCK axis is thus suggested to be useful as a novel therapeutic target for the treatment of PAH, especially when associated with thrombus.
- © 2010 by American Heart Association, Inc.