ABSTRACT
We investigated the neuroprotective effects of fasudil's active metabolite, hydroxyfasudil, a Rho-kinase inhibitor, in a rat stroke model in which endothelial damage and subsequent thrombotic occlusion were selectively induced in perforating arteries. By examining the effects on the endothelial damage/dysfunction, we thought to explore the mechanism of Rho-kinase inhibitors. Hydroxyfasudil (10mg/kg, i.p., once daily for 3 days) significantly improved neurological functions and reduced the size of the infarct area produced by internal carotid artery injection of sodium laurate in a rat cerebral microthrombosis model. Treatment with fasudil or hydroxyfasudil concentration-dependently inhibited tumor necrosis factor alpha-induced tissue factor expression on the surface of cultured human umbilical vein endothelial cells. They also inhibited thrombin-induced endothelial hyperpermeability. The present findings suggest that hydroxyfasudil is efficacious in preventing brain damage associated with cerebral ischemia, and is partially responsible for fasudil's cytoprotective potential. The results also suggest that the therapeutic benefits against ischemic injury of Rho-kinase inhibitors are attributed, at least in part, to activity upon endothelial damage/dysfunction.
Subject(s)
1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/analogs & derivatives , Brain Ischemia/drug therapy , Endothelium/drug effects , Enzyme Inhibitors/pharmacology , Neuroprotective Agents/pharmacology , rho-Associated Kinases/antagonists & inhibitors , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/administration & dosage , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/pharmacokinetics , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/pharmacology , Animals , Brain/blood supply , Brain/drug effects , Brain/pathology , Brain Ischemia/pathology , Capillary Permeability/drug effects , Cells, Cultured , Disease Models, Animal , Endothelium/metabolism , Endothelium/pathology , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/pharmacokinetics , Humans , In Vitro Techniques , Male , Neuroprotective Agents/administration & dosage , Neuroprotective Agents/pharmacokinetics , Rats , Rats, Sprague-Dawley , Stroke/drug therapy , Stroke/pathology , Thromboplastin/metabolism , Tumor Necrosis Factor-alpha/metabolism , Umbilical Veins/drug effects , Umbilical Veins/metabolismABSTRACT
We studied whether there was a relationship between the anticoagulant effects of recombinant human soluble thrombomodulin (rhsTM) and activation of protein C in a primate model of acute vascular graft thrombosis in 11 baboons (Papio species). Baboons were pretreated with 0.1, 1 and 5 mg/kg of rhsTM, with or without co-injection of a neutralising monoclonal antibody to protein C (HPC4) in the 1 mg/kg rhsTM group. Subsequently, thrombogenic polyester grafts were deployed for 3 h into chronic exteriorised arteriovenous shunts. Thrombus growth in the graft, plasma-activated protein C (APC) levels, coagulation and thrombosis markers were determined. In untreated baboons, baseline circulating APC levels more than doubled and graft thrombi propagated until reaching equilibrium in about 1 h. Treatment with rhsTM reduced thrombus propagation rates, prolonged the clotting and bleeding times, decreased thrombin-antithrombin complex, beta-thromboglobulin and fibrinopeptide A levels, and, surprisingly, also decreased systemic APC levels, in a dose-dependent manner. In the presence of HPC4 antibody to inhibit APC generation, the acute antithrombotic activity of rhsTM on graft thromboses was not attenuated for up to 80 min, but sustained thrombus accumulation was observed over a 180-min period. These findings suggest that, in contrast to the prevailing hypotheses, the primary antithrombotic activity of rhsTM is independent of protein C, at least in this primate model. Direct inhibition of thrombin's prothrombotic activity upon complex formation with rhsTM might explain the molecular mechanism of the observed antithrombotic effect.