Role of cyclooxygenase and derived reactive oxygen species in rho-kinase-mediated impairment of endothelium-dependent vasodilation and blood flow after ischemia-reperfusion of the rat kidney.

BACKGROUND/AIMS: Decreased endothelium-dependent vasodilation and blood flow in renal ischemia-reperfusion (IR) may result in part from rho-kinase activation, and cyclooxygenase (COX) activation, and resultant reactive oxygen species (ROS) may be involved. METHODS: We tested this hypothesis in male Wistar rats, subjected to 60 min of bilateral clamping of the renal arteries and 60 min of reperfusion or a sham procedure, and treated by the rho-kinase inhibitor Y27632 (1 mg/kg) and/or the nonspecific COX inhibitor diclofenac (10 mg/kg). Renal blood flow was measured by fluorescent microspheres, and ROS in the arterial endothelium was quantified by dihydroethidium staining. Endothelium-dependent vasodilation was determined by an acetylcholine concentration-response curve in the presence or absence of diclofenac (10 microM). RESULTS: Y27632 increased renal blood flow and reduced ROS in vivo, and improved endothelium-dependent vasodilation in vitro, following IR with or without diclofenac. Following IR, diclofenac had no effect on renal blood flow and ROS in vivo, but improved endothelium-dependent vasodilation in vitro. CONCLUSION: Activation of rho-kinase impairs endothelium-dependent vasodilation and perfusion following renal IR, independently of COX and resultant ROS. In contrast, the vasodilatory effect of rho-kinase inhibition may be partly mediated by decreasing ROS, unrelated to COX and resultant vasoconstricting prostanoids.

3beta-OH-tibolone acutely dilates rat muscle arterioles similar to 3alpha-OH-tibolone.

In an earlier study, we focused on the vasoactive effect of 3alpha-OH-tibolone on spontaneously constricted isolatedfemale rat gracilis muscle arterioles. Vasodilator effects (from 10 to 10 M) of 3alpha-OH-tibolone were similar to those of 17beta-estradiol. It was reported that 3beta-OH-tibolone like estradiol altered GABAB activation in neurons through a membrane estrogen receptor, whereas the 3alpha-OH metabolite did not. We therefore hypothesized that the 3beta-OH metabolite may also have a vasodilating effect in our isolated arteriole model. The results indicate that 3beta-OH-tibolone induces a vasodilator effect in small arterioles that is comparable with that of 3alpha-OH-tibolone at the same concentration. This is intriguing because the binding affinity of 3alpha-OH-tibolone to the estrogen receptor is almost twice that of 3beta-OH-tibolone. Other mechanisms may play a role.

Endothelium-dependent dilatory effects of 3alpha-OH-tibolone in gracilis muscle arterioles of the female Wistar rat.

OBJECTIVE: Tibolone is a synthetic steroid used for the treatment of the symptoms of menopause that, once metabolized, has estrogenic, progestogenic, and androgenic properties. We investigated the direct vasodilatory effects of the major active tibolone metabolite 3alpha-OH-tibolone and its sulfated form on female rat skeletal muscle arterioles, which play an important role in the control of blood pressure. DESIGN: In isolated, pressurized spontaneously constricted arterioles (mean passive diameter 83 +/- 3 microm), we investigated the vasodilatory effect of 3alpha-OH-tibolone and its sulfated form. To study the role of the endothelium and in particular that of nitric oxide, we repeated the experiments with 3alpha-OH-tibolone after removal of the endothelium and on vessels pretreated with the nitric oxide synthesis inhibitor, Nomega-nitro-L-arginine (L-Na). Finally we compared the vasodilatory effect of 3alpha-OH-tibolone with 17beta-estradiol. RESULTS: A dose-dependent dilatation to 3alpha-OH-tibolone was observed starting at a concentration of 10 M. With the sulfated form of 3alpha-OH-tibolone, dilatation was only present at the highest concentration (10 M). In the denuded vessels, the vasodilatory effect was absent at concentrations from 10 to 10 M. The dilatation induced by 3alpha-OH-tibolone was not significantly reduced by L-Na. The vasodilatory effect of 3alpha-OH-tibolone did not differ from that of 17beta-estradiol. CONCLUSIONS: 3alpha-OH-tibolone has a dose-dependent vasodilatory effect on isolated skeletal muscle arterioles from the rat. The sulfated form has no vasodilatory effect in this setup. This finding suggests that during this short incubation time there was no conversion of the sulfated metabolite into its active form by the vascular endothelium. The vasodilatory effect of 3alpha-OH-tibolone is endothelium dependent at physiologic concentrations and comparable to that of 17beta-estradiol.

Rho kinase regulates renal blood flow by modulating eNOS activity in ischemia-reperfusion of the rat kidney.

Renal ischemia-reperfusion (I/R) results in vascular dysfunction characterized by a reduced endothelium-dependent vasodilatation and subsequently impaired blood flow. In this study, we investigated the role of Rho kinase in endothelial nitric oxide synthase (eNOS)-mediated regulation of renal blood flow and vasomotor tone in renal I/R. Male Wistar rats were subjected to 60-min bilateral clamping of the renal arteries or sham procedure. One hour before the clamping, the Rho kinase inhibitor Y27632 (1 mg/kg) was intravenously infused. After I/R, renal blood flow was measured using fluorescent microspheres. I/R resulted in a 62% decrease in renal blood flow. In contrast, the blood flow decrease in the group treated with the Rho kinase inhibitor (YI/R) was prevented. Endothelium-dependent vasodilatation of renal arcuate arteries to ACh was measured ex vivo in a pressure myograph. These experiments demonstrated that the in vivo treatment with the Rho kinase inhibitor prevented the decrease in the nitric oxide (NO)-mediated vasodilator response. In addition, after I/R renal interlobar arteries showed a decrease in phosphorylated eNOS and vasodilator-stimulated phosphoprotein, a marker for bioactive NO, which was attenuated by in vivo Rho kinase inhibition. These findings indicate that in vivo inhibition of Rho kinase in renal I/R preserves renal blood flow by improving eNOS function.