These results indicate that Rho-kinase is upregulated at the spastic site and plays a key role in inducing vascular smooth muscle hypercontraction by inhibiting myosin phosphatase through the phosphorylation of MBS in our porcine model.

Abstract-Intracellular signaling pathway mediated by small GTPase Rho and its effector Rho-kinase plays an important role in regulation of vascular smooth muscle contraction and other cellular functions. We have recently demonstrated that Rho-kinase is substantially involved in angiotensin II-induced gene expressions and various cellular responses in vitro. However, it remains to be examined whether Rho-kinase is involved in the angiotensin II-induced cardiovascular hypertrophy in vivo and, if so, what mechanisms are involved. Long-term infusion of angiotensin II for 4 weeks caused hypertrophic changes of vascular smooth muscle and cardiomyocytes in rats. Both changes were significantly suppressed by concomitant oral treatment with fasudil, which is metabolized to a specific Rho-kinase inhibitor, hydroxyfasudil, after oral administration. Angiotensin II caused a perivascular accumulation of macrophages and Rho-kinase activation, both of which were also significantly suppressed by fasudil. Vascular NAD(P)H oxidase expression (nox1, nox4, gp91phox, and p22phox) and endothelial production of superoxide anions were markedly increased by angiotensin II, both of which were also significantly suppressed by fasudil. Thus, fasudil ameliorated the impaired endothelium-dependent relaxations caused by angiotensin II without affecting vasodilator function of vascular smooth muscle. These results provide evidence that Rho-kinase is substantially involved in the angiotensin II-induced cardiovascular hypertrophy in rats in vivo. The suppression of endothelial NAD(P)H oxidase upregulation and resultant superoxide production and the amelioration of endothelial vasodilator function may be involved in this process. 4 It has been demonstrated that Rho-kinase/ROK␣/ROCK II (an isoform to p160ROCK/ROK␤/ROCK I), 5 which is an effector of the small GTPase Rho, inhibits MLC phosphatase activity by phosphorylating its myosin-binding subunit 6,7 and thus plays a central role in agonist-induced Ca 2ϩ sensitization and hypercontraction of VSMCs. 4,5,7 We have demonstrated that Rho-kinase plays an important role in angiotensin II-induced mRNA expression of monocyte chemoattractant protein-1 8 and of plasminogen activator inhibitor-1 9 in cultured rat aortic VSMCs. We have recently demonstrated that in cultured human coronary VSMCs, the expression of Rho-kinase is enhanced by inflammatory stimuli, such as angiotensin II and interleukin-1␤ (IL-1␤). 10 However, it remains to be determined whether Rho-kinase is involved in angiotensin II-induced cardiovascular hypertrophy in vivo and, if so, what mechanisms are involved. The present study was thus designed to examine these points in rats in vivo. Materials and Methods AnimalsThis experiment was reviewed and approved by the Committee on Ethics of Animal Experiments of the Kyushu University. A total of 111 adult male WKY rats (14 to 17 weeks old, weighing 300 to 350 g) obtained from the colony at the Kyushu University were used. Animals were anesthetized with intraperitoneal pentobarbital

The endothelium plays an important role in maintaining vascular homeostasis by synthesizing and releasing several vasodilating factors, including prostacyclin, NO, and endothelium-derived hyperpolarizing factor (EDHF). We have recently identified that endothelium-derived H2O2 is an EDHF in mesenteric arteries of mice and humans and in porcine coronary microvessels. However, the mechanism for the endothelial production of H2O2 as an EDHF remains to be elucidated. In this study, we tested our hypothesis that Cu,Zn-superoxide dismutase (Cu,Zn-SOD) plays a pivotal role in endothelium-dependent hyperpolarization, using control and Cu,Zn-SOD–/– mice. In mesenteric arteries, EDHF-mediated relaxations and hyperpolarizations were significantly reduced in Cu,Zn-SOD–/– mice with no inhibitory effect of catalase, while endothelium-independent relaxations and hyperpolarizations were preserved. Endothelial H2O2 production also was significantly reduced in Cu,Zn-SOD–/– mice. In Langendorff isolated heart, bradykinin-induced increase in coronary flow was significantly reduced in Cu,Zn-SOD–/– mice, again with no inhibitory effect of catalase. The exogenous SOD mimetic tempol significantly improved EDHF-mediated relaxations and hyperpolarizations and coronary flow response in Cu,Zn-SOD–/– mice. These results prove the novel concept that endothelial Cu,Zn-SOD plays an important role as an “EDHF synthase” in mice, in addition to its classical role to scavenge superoxide anions

The rates in Japan of cardiac arrest and death during anesthesia and surgery due to all etiologies as well as those totally attributable to anesthesia are comparable to those of other developed countries.