Shin HK, Salomone S, Potts EM, et al. Rho-kinase inhibition acutely augments blood flow in focal cerebral ischemia via endothelial mechanisms. J Cereb Blood Flow Metab 2007;27:998-1009

Department of Radiology, Stroke and Neurovascular Regulation Laboratory, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts 02129, USA.
Journal of Cerebral Blood Flow & Metabolism (Impact Factor: 5.41). 06/2007; 27(5):998-1009. DOI: 10.1038/sj.jcbfm.9600406
Source: PubMed


Rho-kinase is a serine threonine kinase that increases vasomotor tone via its effects on both endothelium and smooth muscle. Rho-kinase inhibition reduces cerebral infarct size in wild type, but not endothelial nitric oxide synthase deficient (eNOS-/-) mice. The mechanism may be related to Rho-kinase activation under hypoxic/ischemic conditions and impaired vasodilation because of downregulation of eNOS activity. To further implicate Rho-kinase in impaired vascular relaxation during hypoxia/ischemia, we exposed isolated vessels from rat and mouse to 60 mins of hypoxia, and showed that hypoxia reversibly abolished acetylcholine-induced eNOS-dependent relaxation, and that Rho-kinase inhibitor hydroxyfasudil partially preserved this relaxation during hypoxia. We, therefore, hypothesized that if hypoxia-induced Rho-kinase activation acutely impairs vasodilation in ischemic cortex, in vivo, then Rho-kinase inhibitors would acutely augment cerebral blood flow (CBF) as a mechanism by which they reduce infarct size. To test this, we studied the acute cerebral hemodynamic effects of Rho-kinase inhibitors in ischemic core and penumbra during distal middle cerebral artery occlusion (dMCAO) in wild-type and eNOS-/- mice using laser speckle flowmetry. When administered 60 mins before or immediately after dMCAO, Rho-kinase inhibitors hydroxyfasudil and Y-27632 reduced the area of severely ischemic cortex. However, hydroxyfasudil did not reduce the area of CBF deficit in eNOS-/- mice, suggesting that its effect on CBF within the ischemic cortex is primarily endothelium-dependent, and not mediated by its direct vasodilator effect on vascular smooth muscle. Our results suggest that Rho-kinase negatively regulates eNOS activity in acutely ischemic brain, thereby worsening the CBF deficit. Therefore, rapid nontranscriptional upregulation of eNOS activity by small molecule inhibitors of Rho-kinase may be a viable therapeutic approach in acute stroke.

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    • "Further, apocynin has antioxidant properties [93] and also inhibits rho kinase [94,95]. Rho kinase-inhibition could be the actual neuroprotective mechanism of apocynin as Rho kinase is implicated in stroke [96]. Therefore, any results on the role of NADPH oxidase obtained by using apocyin should be questioned. "
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    ABSTRACT: As recently reviewed, 1026 neuroprotective drug candidates in stroke research have all failed on their road towards validation and clinical translation, reasons being quality issues in preclinical research and publication bias. Quality control guidelines for preclinical stroke studies have now been established. However, sufficient understanding of the underlying mechanisms of neuronal death after stroke that could be possibly translated into new therapies is lacking. One exception is the hypothesis that cellular death is mediated by oxidative stress. Oxidative stress is defined as an excess of reactive oxygen species (ROS) derived from different possible enzymatic sources. Among these, NADPH oxidases (NOX1-5) stand out as they represent the only known enzyme family that has no other function than to produce ROS. Based on data from different NOX knockout mouse models in ischemic stroke, the most relevant isoform appears to be NOX4. Here we discuss the state-of-the-art of this target with respect to stroke and open questions that need to be addressed on the path towards clinical translation.
    Experimental and Translational Stroke Medicine 05/2012; 4(1):11. DOI:10.1186/2040-7378-4-11
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    • "c o m / l o c a t e / e j p h a r guanosine triphosphatases of the Ras-homologus (Rho) family and one of their effectors, Rho-kinase, are known to act as molecular switches controlling several critical functions, including, cell adhesion and contraction, migration, reactive oxygen species formation and oncogenic transformation (Alblas et al., 2001; Itoh et al., 1999; Slotta et al., 2006). Interestingly, Rho-kinase inhibitors have been reported to attenuate ischemia/reperfusion and endotoxemic injury in the liver (Slotta et al., 2008) as well as protecting against tissue fibrosis (Kitamura et al., 2007), cholestasis (Laschke et al., 2010), cerebral and intestinal ischemia (Santen et al., 2010; Shin et al., 2007). However, the role of the Rho-kinase signaling in regulating pulmonary recruitment of neutrophils and tissue damage in abdominal sepsis is not known. "
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    ABSTRACT: We hypothesized that Rho-kinase signaling plays a role in mechanical and adhesive mechanisms of neutrophil accumulation in lung. Male C57BL/6 mice were treated with the Rho-kinase inhibitor Y-27632 prior to cecal ligation and puncture (CLP). Lung levels of myeloperoxidase (MPO) and histological tissue damage were determined 6h and 24h after CLP. Expression of Mac-1 and F-actin formation in neutrophils were quantified by using flow cytometry 6h after CLP. Mac-1 expression and F-actin formation were also determined in isolated neutrophils up to 3h after stimulation with CXCL2. Labeled and activated neutrophils co-incubated with Y-27632, an anti-Mac-1 antibody and cytochalasin B were adoptively transferred to CLP mice. Y-27632 reduced the CLP-induced pulmonary injury and MPO activity as well as Mac-1 on neutrophils. Neutrophil F-actin formation peaked at 6h and returned to baseline levels 24h after CLP induction. Rho-kinase inhibition decreased CLP-provoked F-actin formation in neutrophils. CXCL2 rapidly increased Mac-1 expression and F-actin formation in neutrophils. Co-incubation with Y-27632 abolished CXCL2-induced Mac-1 up-regulation and formation of F-actin in neutrophils. Notably, co-incubation with cytochalasin B inhibited formation of F-actin but did not reduce Mac-1 expression on activated neutrophils. Adoptive transfer experiments revealed that co-incubation of neutrophils with the anti-Mac-1 antibody or cytochalasin B significantly decreased pulmonary accumulation of neutrophils in septic mice. Our data show that targeting Rho-kinase effectively reduces neutrophil recruitment and tissue damage in abdominal sepsis. Moreover, these findings demonstrate that Rho-kinase-dependent neutrophil accumulation in septic lung injury is regulated by both adhesive and mechanical mechanisms.
    European journal of pharmacology 02/2012; 682(1-3):181-7. DOI:10.1016/j.ejphar.2012.02.022 · 2.53 Impact Factor
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    • "Additionally, fasudil protects neurological function (Li et al. 2009; Satoh et al. 1996, 2008; Toshima et al. 2000; Yamashita et al. 2007). The tissue sparing was attributable to enhanced bloodflow related to increased endothelial nitric oxide synthase activity (Li et al. 2009; Rikitake et al. 2005; Satoh et al. 2008; Shin et al. 2007). Fasudil has also been reported to protect neurons from cell death after hypoxia (Ding et al. 2010) and acute (Yamashita et al. 2007) and chronic (Huang et al. 2008) ischemia. "
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    Cell and Tissue Research 02/2012; 349(1):119-32. DOI:10.1007/s00441-012-1334-7 · 3.57 Impact Factor
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