• Biomedical Science Letters
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• v.19 no.3
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• pp.180-187
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• 2013

Hypoxia is an integral part of the environment during luteolysis. In this study we examined whether hypoxia could directly stimulate endothelial cells to produce nitric oxide (NO). Endothelial cells were cultured in hypoxic (5% $O_2$) or normoxic (20% $O_2$) conditions and the levels of total NO, inducible NO and endothelial NO was measured. We found that hypoxia but not normoxia upregulated NO production. The increased NO levels correlated with increased inducible NO synthase (iNOS) expression whereas expression of endothelial NOS (eNOS) expression remained constant. Addition of the iNOS specific inhibitor 1400W to hypoxic cultures prevented NO production suggesting that hypoxia-induced NO production in endothelial cells was due mainly to upregulation of iNOS. We also found that prostaglandin $F_{2{\alpha}}$ (PGF) production was unaffected by hypoxia suggesting that upregulation of NO was not due to increased synthesis of PGF. In summary, we report that endothelial cells cultured under hypoxic conditions produce NO via the iNOS pathway. This study provides the importance of the relation between the hypoxic environment and the induction of NO by endothelial cells during regression of the corpus luteum in the ovary.

• Toxicological Research
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• v.24 no.3
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• pp.169-174
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• 2008

This study examined the mechanisms underlying the effects of the vasorelaxation active substance(VAS), dimethyladenosine-5'-L-arabinose, and its partial purification fraction on nitric oxide synthase in improving erectile dysfunction with particular focus on the nitric oxide (NO)-cGMP pathways. Two rat models, 9-month-old SD rats and 11-month-old SD rats, were given VAS(40 mg/kg per day) for 4 days, The aqueous fraction of silworm male pupae extract; semi-purified VAS(100 mg/kg per day) for 10 days, respectively. The NOS activities of the following three enzymes were examined: neuronal NO synthase(nNOS), inducible NOS(iNOS), endothelial NOS(eNOS), vascular endothelial growth factor on endothelial cells(VEGF) and anti-inflammation effect of Tumor necrosis factor-$\alpha$. The results showed increases in the nitric oxide synthase activities. Western blotting of the tissue homogenate showed an increase in the nNOS level in the brain and tongue, and an increase in the endothelial NO synthase(eNOS) level in penis. However, there was little association with VEGF production in HUVEC endothelial cells and no relationship with TNF-$\alpha$ which showed low levels.

• Korean Journal of Veterinary Research
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• v.43 no.4
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• pp.551-554
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• 2003

Nitric oxide synthase (NOS) has been reported in uterus. We examined the expression of the NOS isoforms, constitutive endothelial (eNOS) and inducible NOS (iNOS), in bovine uterus by immunohistochemistry. eNOS immunoreactivity was localized predominantly to the endothelial cells that line uterine microvessels and to endometrial glandular epithelial cells, but was barely detectable in endometrial stromal cells. iNOS immunostaining was detected in glandular epithelial and stromal cells in the endometrium and in the endothelial cells of myometrial blood vessels. These findings suggest that both eNOS and iNOS may play important roles in the physiology of the uterus, possibly by generating NO.

• Journal of Microbiology and Biotechnology
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• v.18 no.2
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• pp.392-395
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• 2008

In an attempt to delineate the direct effect of arsenite-induced endothelial dysfunction on nitric oxide (NO) production, confluent bovine aortic endothelial cells (BAEC) were incubated with arsenite, and endothelial NO synthase expression and NO production were measured. Exposure of arsenite decreased NO production for up to 24h. This decrease was accompanied by decreases in cAMP, protein kinase A (PKA) activity, and furthermore, significant reduction of pCREB. In conclusion, this study is the first to demonstrate that exposure of arsenite decreases NO production by a reduction of pCREB and PKA activity that may be mediated by cAMP, leading to endothelial dysfunction.

• Journal of Ginseng Research
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• v.37 no.1
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• pp.64-73
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• 2013

Korean red ginseng water extract (KG-WE) has known beneficial effects on the cardiovascular system via inducting nitric oxide (NO) production in endothelium. Endothelial arginase inhibits the activity of endothelial nitric oxide synthase (eNOS) by substrate depletion, thereby reducing NO bioavailability and contributing to vascular diseases including hypertension, aging, and atherosclerosis. In the present study, we demonstrate that KG-WE inhibits arginase activity and negatively regulates NO production and reactive oxygen species generation in endothelium. This is associated with increased dimerization of eNOS without affecting the protein expression levels of either arginase or eNOS. In a vascular tension assay, when aortas isolated from wild type mice were incubated with KG-WE, NO-dependent enhanced vasorelaxation was observed. Furthermore, KG-WE administered via by drinking water to atherogenic model mice being fed high cholesterol diet improved impaired vascular function. Taken together, these results suggest that KG-WE may exert vasoprotective effects through augmentation of NO signaling by inhibiting arginase. Therefore, KG-WE may be useful in the treatment of vascular diseases derived from endothelial dysfunction, such as atherosclerosis.

• Journal of Applied Biological Chemistry
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• v.48 no.4
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• pp.178-182
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• 2005

Nitric oxide (NO) produced from endothelial cells plays a critical role in vascular physiology. The regulation of endothelial NO synthase (eNOS) involves various mechanisms including multiple Ser/Thr phosphorylations. Recently, eNOS-Ser617 was newly recognized to be phosphorylated in response to humoral factors including vascular endothelial growth factor. However, it remains unknown whether and how eNOS-Ser617 phosphorylation is stimulated by shear stress, the primary stimulus of endothelial NO production. This issue was explored in the present study using cultured bovine aortic endothelial cells (BAECs). Over-expression of a constitutively active protein kinase B(Akt) mutant in BAECs increased Ser617 phosphorylation while constitutively active protein kinase A mutant had no effect. When BAECs were subjected to an arterial level of laminar shear stress, eNOS-Ser617 phosphorylation was clearly increased in a time-dependent manner. Shear stress also stimulated Akt phosphorylation at Thr308, one of the key regulatory sites. The time courses of eNOS-Ser617 and Akt-Thr308 phosphorylations appeared to be very similar. These results suggested that eNOS-Ser617 phosphorylation, mediated by Akt, is a physiological response to the mechanical shear stress, involved in the regulation of NO production in endothelial cells.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.5
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• pp.441-447
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• 2016

Despite the complex vascular effects of dexmedetomidine (DEX), its actions on human pulmonary resistance arteries remain unknown. The present study tested the hypothesis that DEX inhibits vascular tension in human pulmonary arteries through the endothelial nitric oxide synthase (eNOS) mediated production of nitric oxide (NO). Pulmonary artery segments were obtained from 62 patients who underwent lung resection. The direct effects of DEX on human pulmonary artery tension and changes in vascular tension were determined by isometric force measurements recorded on a myograph. Arterial contractions caused by increasing concentrations of serotonin with DEX in the presence or absence of L-NAME (endothelial nitric oxide synthase inhibitor), yohimbine (${\alpha}_2$-adrenoceptor antagonist) and indomethacin (cyclooxygenase inhibitor) as antagonists were also measured. DEX had no effect on endothelium-intact pulmonary arteries, whereas at concentrations of $10^{-8}{\sim}10^{-6}mol/L$, it elicited contractions in endothelium-denuded pulmonary arteries. DEX (0.3, 1, or $3{\times}10^{-9}mmol/L$) inhibited serotonin-induced contraction in arteries with intact endothelium in a dose-dependent manner. L-NAME and yohimbine abolished DEX-induced inhibition, whereas indomethacin had no effect. No inhibitory effect was observed in endothelium-denuded pulmonary arteries. DEX-induced inhibition of vasoconstriction in human pulmonary arteries is mediated by NO production induced by the activation of endothelial ${\alpha}_2$-adrenoceptor and nitric oxide synthase.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.1
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• pp.83-90
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• 2017

Advanced age is one of the risk factors for vascular diseases that are mainly caused by impaired nitric oxide (NO) production. It has been demonstrated that endothelial arginase constrains the activity of endothelial nitric oxide synthase (eNOS) and limits NO generation. Hence, arginase inhibition is suggested to be vasoprotective in aging. In this study, we examined the effects of intravenous injection of Piceatannol, an arginase inhibitor, on aged mice. Our results show that Piceatannol administration reduced the blood pressure in aged mice by inhibiting arginase activity, which was associated with NO production and reactive oxygen species generation. In addition, Piceatannol administration recovered $Ca^{2+}$/calmodulin-dependent protein kinase II phosphorylation, eNOS phosphorylation and eNOS dimer stability in the aged mice. The improved NO signaling was shown to be effective in attenuating the phenylephrine-dependent contractile response and in enhancing the acetylcholine-dependent vasorelaxation response in aortic rings from the aged mice. These data suggest Piceatannol as a potential treatment for vascular disease.

• Journal of Physiology & Pathology in Korean Medicine
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• v.20 no.3
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• pp.680-684
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• 2006

This study was designed to investigate effects of KH-304 in improving erectile dysfunction (ED), particularly in terms of nitric oxide (NO)-cGMP pathways. After oral administration of the KH-304 water extract, 1OOmg, 300mg, 500mg or 700mg per 1 kg of Dody weigh for 10days, We examined the expression and activity of two enzyme: neuronal NO synthase (nNOS), endothelial NO synthase (eNOS) and that act upon the major NO-cGMP signaling pathway in penile tissue. Effect of KH-304 on COMP degradation was also examined using bovine vascular smooth muscle cells pretreated with an NO donor, S-nitroso-N-Acetylpenicillamine (SNAP), Also, it examined the endothelial NO synthase (eNOS) for seaching effecting period (100mg, 300mg/kg for 10 and 30days) and peak intracavernous pressures (ICPS) in penile tissues rabbit copus cavernosum contracted by 10-6 M phenylephrine. The severely reduced peak intracavernous pressures (ICPS) in penile tissues were restored completely after KH-304 treatment, and KH-304 treatment significantly made the latency period earlier. Furthermore, the penile expression levels of nNOS, eNOS dependent NOS activities and COMP concentrations were increased significantly in the KH-304 100, 300mg treated rats. These results suggest that KH-304 with high expression of NOS may be useful in erectile dysfunction.

• Environmental Analysis Health and Toxicology
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• v.29
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• pp.10.1-10.6
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• 2014

Objectives Cigarette smoking had been recorded as the main cause of impaired endothelium-dependent vasodilation in smokers by reducing nitric oxide (NO), a production of endothelial nitric oxide synthase (eNOS). However, the mechanism of NO impairment via eNOS activity is unclear until now. In this study, cell passage is suggested to be a relevant factor to eNOS expression under cigarette smoking stress. Methods Bovine aortic endothelial cells (BAECs) were chosen as the research subject with passages ranking from 6 to 9 (6P to 9P). After exposure of cigarette smoking extract (CSE) solution, MTT assay and Western blot method were performed to check the cell viability as well as eNOS protein concentration. In these experiments, four concentrations of CSE at 0.5, 1, 2, and 4% were selected for treatment. Results Our results showed that cells almost died at 4% of CSE. Besides, eNOS protein mass had a linear decrease under the increase of CSE concentration. In addition, the effect of CSE on eNOS expression was dissimilar between different passages. Conclusions This study indicated that CSE had effect on both cell viability and eNOS expression. Besides, a reduction in protein mass was matched with the decrease of cell viability due to CSE tress. Last but not least, the response of eNOS protein to different concentration of CSE at different passages was disparate, making the hypothesis about cell passage related inhibition of eNOS caused by CSE solution.