Involvement Of Nitric Oxide, Prostacyclin And Endothelium-Dependent Hyperpolarization In Dilatation Of Subcutaneous Arteries Of Diabetic Patients And Rats
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Date
2015-10
Authors
Mokhtar, Siti Safiah
Journal Title
Journal ISSN
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Publisher
Universiti Sains Malaysia
Abstract
Diabetic endothelial dysfunction is characterized by the impairment of endothelium-dependent relaxations. The endothelium evokes relaxation of underlying smooth muscle through the production of endothelium-dependent relaxing factors (EDRF) including nitric oxide (NO), prostacyclin and endothelium-dependent hyperpolarization (EDH). There were three main aims: firstly, to demonstrate the role of individual EDRF in the endothelial responses of normal small arteries from human and rats; secondly, to determine the effect of diabetes mellitus on the contribution of individual EDRF; and lastly, to relate the functional abnormalities in diabetic microcirculation with the changes in the expression and localization of enzymes and/ or receptors involved. The functional and molecular studies were performed in isolated blood vessels of humans and rats. In humans, subcutaneous arteries were isolated from tissues of healthy controls [n=16, mean age 31.50 (12.37)] and type-2 diabetic patients [n=20, mean age 55.3 (9.93)] who underwent lower limb surgeries. In rats, tail arteries were isolated from controls (n=14) and streptozotocin-induced diabetic rats (n=14). The rats were sacrificed after six weeks of diabetes duration. Vascular function consisted of endothelial and smooth muscle relaxation responses were determined using wire myograph system. The contributions of NO, prostacyclin and EDH were studied using selective pharmacological inhibitors. Expressions of the proteins involved, including endothelial nitric oxide synthase (eNOS), cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), prostacyclin synthase (PGIS) and prostacyclin (IP) receptor, were determined by Western blotting and immunohistochemistry. In human study, endothelium-dependent relaxations to acetylcholine (ACh) were impaired in subcutaneous arteries from diabetic patients compared to controls [Rmax; 81.17 (13.71)% versus 91.34 (4.65)%, P = 0.009]. ACh-induced NO-mediated relaxation was attenuated in subcutaneous arteries from diabetic patients compared to controls [Rmax; 31.61 (26.24)% versus 73.59 (13.79)%, P < 0.001]. However, EDH-type relaxation was greater in subcutaneous arteries from diabetic patients compared to controls [Rmax; 63.51 (17.86)% versus 43.14 (17.08)%, P = 0.003]. Relaxations of
subcutaneous smooth muscle cells to sodium nitroprusside (NO donor) and salbutamol (β-adrenoceptor agonist) were preserved, but those to prostacyclin (IP receptor agonist) were attenuated in diabetic patients compared to controls [Rmax; 43.31 (29.45)% versus 70.80 (22.84)%, P = 0.017]. Western blotting and immunostaining indicated that diabetes decreased the expressions of eNOS, PGIS and IP receptors, but increased COX-2 protein in human subcutaneous arteries. There were negative significant correlations between NO-mediated relaxations with fasting blood glucose (FBG) and glycated haemoglobin (HbA1C) in human subcutaneous arteries. In animal study, ACh-induced relaxation was decreased significantly in tail arteries of diabetic compared to control rats [Rmax; 70.81 (9.29)% versus 85.05 (11.80)%, P = 0.001]. NO-mediated relaxation was attenuated significantly in diabetic compared to control rats [Rmax; 48.47 (21.85)% versus 68.39 (23.72)%, P = 0.029]. EDH-type relaxation was slightly reduced in diabetic compared to control rats [Rmax; 17.81 (25.21)% versus 34.14 (17.19)%, P = 0.056]. Relaxations of tail smooth muscle cells to sodium nitroprusside, salbutamol and prostacyclin were comparable in control and diabetic rats. Western blotting and immunostaining indicated that diabetes diminished eNOS, but increased COX-1 and COX-2 protein expressions in rat tail arteries. ACh-induced endothelium-dependent relaxations in humans and rats small arteries are dependent on NO release and EDH, whereas prostacyclin plays a very minor role. Diabetes impairs endothelium-dependent relaxation in human subcutaneous arteries and rat tail arteries. The impairment is due to a diminished NO contribution which is associated with reduced eNOS protein expression. There are significant negative correlations between NO-mediated relaxation with FBG and HbA1C, suggesting that controlling blood sugar level can be a potential effective approach to ameliorate endothelial dysfunction in diabetes.
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Keywords
Involvement of nitric oxide , of diabetic patients and rats.