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Inducible NOS (inducible + no)
Selected AbstractsNitric Oxide Mediates Inhibitory Effect of Interleukin-1, on Estrogen Production in Human Granulosa-Luteal CellsJOURNAL OF OBSTETRICS AND GYNAECOLOGY RESEARCH (ELECTRONIC), Issue 1 2001Hisako Tobai Abstract Objective: To investigate the effect of IL-1, on NO production and steroidogenesis in human granulosa-luteal cells obtained from women undergoing in vitro fertilization procedures. Subjects and Methods:To investigate the effect of IL-1,, granulosa-luteal cells were cultured with various doses of IL-1, (0, 0.05, 0.5, 5, 50, 100 ng/ml), IL-1, (5 ng/ml) with NG -nitro-L-arginine-methyl ester (l-NAME), selective inhibitors of NOS, sodium nitroprusside (SNP), NO donors and Genistain, a tyrosine kinase inhibitor. Results:IL-1, induced a dose-dependent stimulation of NO production and inhibited the production of estradiol in a significant way in a dose-dependent manner. l-NAME significantly decreased NO production and increased the production of estradiol and progesterone. SNP significantly increased NO production and caused decreases in the production of both estradiol and progesterone. Genistain decreased NO production and significantly increased the production of estradiol and progesterone. Inducible NOS (iNOS) messenger RNA was present in granulosa-luteal cells before treatment with IL-1,. Conclusions:IL-1, stimulated NO production, and NO inhibited the production of estradiol. [source] Downregulation of oxytocin receptors in right ventricle of rats with monocrotaline-induced pulmonary hypertensionACTA PHYSIOLOGICA, Issue 2 2010T. L. Broderick Abstract Aim:, Pulmonary hypertension (PH) in the rat leads to right ventricular (RV) hypertrophy, inflammation and increased natriuretic peptide (NP) levels in plasma and RV. Because the release of nitric oxide (NO) and atrial natriuretic peptide (ANP) is a function of the oxytocin receptor (OTR), we examined the effect of PH on gene and protein expression of OTR, NP (A, atrial; B, brain) and receptors (NPRs), nitric oxide synthases (NOS), interleukin (IL)-1,, IL-6 and tumour necrosis factor-, in the hypertrophied RV in a model of PH. Methods:, RV hypertrophy was induced in male Sprague,Dawley rats with monocrotaline (MCT; 60 mg kg,1) and was confirmed by the presence of an increased RV weight and RV-to-[left ventricle (LV) and septum] ratio. Results:, In the RV of MCT-treated rats, a ,40% reduction in OTR mRNA and protein was observed compared with the RV of control rats. This reduction was associated with increased transcripts of ANP and BNP in both ventricles and a corresponding increase in NP receptor mRNA expression for receptors A, B and C. Protein expression of inducible NOS was increased in the RV, whereas endothelial NOS transcripts were increased only in the LV of MCT-treated rats. In the RV of MCT-treated rats, downregulation of OTR was also associated with increased mRNA expression of IL-1, and IL-6. Conclusion:, Our results show that downregulation of the OTR in the RV of MCT-treated rats is associated with increased expression of NP and their receptors as well as IL-1, and IL-6. This reduction in OTR in RV myocardium may have an impact on cardiac function in the MCT-induced model of PH. [source] Role of shear stress on nitrite and NOS protein content in different size conduit arteries of swineACTA PHYSIOLOGICA, Issue 2 2009X. Guo Abstract Aim:, Inherent fundamental difference exists among arteries of different sizes. The purpose of this study was to evaluate the relation between regional difference of wall shear stress (WSS) in various sizes arteries and contents of nitrite and NO synthase (NOS) isoforms. Methods:, Five different conduit arteries in a wide range of diameter (1,8 mm) were examined in the hind limbs of 13 pigs. Blood flow rate and outer diameter were measured in vivo to determine WSS. Arterial tissues were harvested for the measurement of nitrite and NOS protein contents. The concentration of nitrite, a product of NO synthesis, was determined by high-performance liquid chromatography method. Western blot analysis was used to assess the protein contents of endothelial NOS (eNOS), inducible NOS (iNOS) and neuronal NOS (nNOS). Results:, Our data show that WSS increases with a decrease in artery diameter. Nitrite level increases with increasing WSS and hence decreases with artery diameter. The eNOS protein contents decrease with an increase in diameter. No significant difference for iNOS and nNOS protein contents was found with different artery diameter. A significant positive correlation between tissue nitrite and eNOS protein contents was also observed. Finally, the WSS-normalized eNOS is not significantly different in various size vessels. Conclusion:, Regional difference in blood flow has no effect on iNOS and nNOS protein contents in these conduit arteries. Regional difference in eNOS expression and nitrite contents may be related to the WSS-induced NO by the endothelium under normal physiological conditions. [source] Evidence for a widespread involvement of NO in control of photogenesis in bioluminescent fishACTA ZOOLOGICA, Issue 4 2010Jenny Krönström Abstract Krönström, J. and Mallefet, J. 2009. Evidence for a widespread involvement of NO in control of photogenesis in bioluminescent fish. ,Acta Zoologica (Stockholm) 91: 474,483. The presence of nitric oxide synthase (NOS) and nerve fibres in the photophores of seven bioluminescent fish species (Hygophum benoiti, Myctophum punctatum, Electrona risso, Cyclothone braueri, Vinciguerria attenuata, Maurolicus muelleri and Porichthys notatus) with endogenous photocytes, were investigated. Antibodies directed against neuronal and inducible NOS (n and iNOS respectively) and NADPH-diaphorase activity were used to reveal the locations of NOS, while antibodies directed against acetylated tubulin were used to visualize nerve fibres. The nNOS antibody labelled structures in all investigated photophores except in the organs from P. notatus. The photocytes of P. notatus showed NADPH-diaphorase activity. In the myctophid species, NOS-like immunoreactivity was found in small intracellular structures of the photocytes and in nerve fibres reaching the photocytes. nNOS-positive fibres were also found among lens/filter cells in V. attenuata, and in M. muelleri the cytoplasm of lens/filter cells contained NOS-like material. In C. braueri, a cell type located at a collecting chamber for luminous products in the photophore contained NOS-like material. All photophores received an innervation reaching the photocytes, as well as other components including lens/filter areas. The results of this study comply with an involvement of nitric oxide in the control of bioluminescence in several fish species. [source] The effect of ultraviolet B irradiation on nitric oxide synthase expression in murine keratinocytesEXPERIMENTAL DERMATOLOGY, Issue 6 2000M. Sasaki Abstract: Nitric oxide (NO), which has several physiological functions in skin, is generated by NO synthase (NOS). NOS has at least three isoforms; endothelial NOS (eNOS), brain NOS (bNOS), and inducible NOS (iNOS). Ultraviolet B (UVB) irradiation has been reported to stimulate NO production in skin via induction or activation of NOS, however, the exact mechanism of NOS induction by UVB irradiation remains obscure. In this study, we investigated the direct effect of UVB on the expression of NOS isoforms in murine keratinocytes, and found a significant increase in NO production within 48 h. mRNA and protein expressions of bNOS were both enhanced by UVB irradiation in murine keratinocytes, whereas iNOS mRNA expression was suppressed at 4 and 12 h after UVB irradiation. These results suggest that the enhancement of NO production observed after UVB irradiation in murine keratinocytes may be explained in part by the upregulation of bNOS expression, but not iNOS expression. [source] NO synthase isoforms specifically modify peroxynitrite reactivityFEBS JOURNAL, Issue 19 2010Amandine Maréchal Nitric oxide synthases (NOSs) are multi-domain hemothiolate proteins that are the sole source of nitric oxide (NO) in mammals. NOSs can also be a source or a sink for peroxynitrite (PN), an oxidant that is suspected to be involved in numerous physiopathological processes. In a previous study, we showed that the oxygenase domain of the inducible NOS (iNOSoxy) reacts with PN and changes its oxidative reactivity [Maréchal A, Mattioli TA, Stuehr DJ & Santolini J (2007) J Biol Chem282, 14101,14112]. Here we report a similar analysis on two other NOS isoforms, neuronal NOS (nNOS) and a bacterial NOS-like protein (bsNOS). All NOSs accelerated PN decomposition, with accumulation of a similar heme intermediate. The kinetics of PN decomposition and heme transitions were comparable among NOSs. However, their effects on PN reactivity differ greatly. All isoforms suppressed PN two-electron oxidative activity, but iNOSoxy enhanced PN one-electron oxidation and nitration potencies, the oxygenase domain of nNOS (nNOSoxy) affected them minimally, and bsNOS abolished all PN reactivities. This led to the loss of both NOS and PN decomposition activities for nNOSoxy and iNOSoxy, which may be linked to the reported alterations in their electronic absorption spectra. Bacterial bsNOS was affected to a lesser extent by reaction with PN. We propose that these differences in PN reactivity among NOSs might arise from subtle differences in their heme pockets, and could reflect the physiological specificity of each NOS isoform, ranging from oxidative stress amplification (iNOS) to detoxification (bsNOS). [source] Effect of porto-systemic shunting on NOS expression after extended hepatectomy in ratsHEPATOLOGY RESEARCH, Issue 1 2009Hironori Hayashi Aim:, Several surgical procedures have been developed for reducing portal vein pressure to prevent postoperative liver injury. Nitric oxide synthase expression (NOS) induced by elevation of portal vein pressure is thought to play an important role in liver regeneration, but the details are not well understood. Methods:, Rats in the control group and in the subcutaneous splenic transposition (SST) group underwent 90% partial hepatectomy. Survival and portal vein pressure were analyzed. The serum IL-6 and TNF-, levels were measured by enzyme-linked immunosorbent assay (ELISA). Hepatocyte proliferation and apoptosis 12 hours after hepatectomy were analyzed immunohistochemically. The protein and messenger RNA expression of inducible and endothelial NOS were analyzed using Western blotting and quantitative reverse transcriptase polymerase chain reaction, respectively. Results:, The survival rate of the SST group was significantly higher. Portal vein pressure, TNF-, level and the apoptotic index were significantly lower in the SST group. Twelve hours after surgery, liver inducible NOS (iNOS) protein expression was significantly lower in the SST group. However, protein expression of endothelial NOS was not significantly different between the groups. Conclusion:, Inducible NOS expression after extended hepatectomy is related to the effects of porto-systemic shunting on the splanchnic circulation. Also, iNOS induction and concomitant nitric oxide generation appear to participate in the cytotoxicity of excessive portal pressure after extended hepatectomy. [source] Inhibition of nitric oxide synthase inhibitors and lipopolysaccharide induced inducible NOS and cyclooxygenase-2 gene expressions by rutin, quercetin, and quercetin pentaacetate in RAW 264.7 macrophagesJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2001Yen-Chou Chen Abstract Several natural flavonoids have been demonstrated to perform some beneficial biological activities, however, higher-effective concentrations and poor-absorptive efficacy in body of flavonoids blocked their practical applications. In the present study, we provided evidences to demonstrate that flavonoids rutin, quercetin, and its acetylated product quercetin pentaacetate were able to be used with nitric oxide synthase (NOS) inhibitors (N -nitro- L -arginine (NLA) or N -nitro- L -arginine methyl ester (L -NAME)) in treatment of lipopolysaccharide (LPS) induced nitric oxide (NO) and prostaglandin E2 (PGE2) productions, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) gene expressions in a mouse macrophage cell line (RAW 264.7). The results showed that rutin, quercetin, and quercetin pentaacetate-inhibited LPS-induced NO production in a concentration-dependent manner without obvious cytotoxic effect on cells by MTT assay using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide as an indicator. Decrease of NO production by flavonoids was consistent with the inhibition on LPS-induced iNOS gene expression by western blotting. However, these compounds were unable to block iNOS enzyme activity by direct and indirect measurement on iNOS enzyme activity. Quercetin pentaacetate showed the obvious inhibition on LPS-induced PGE2 production and COX-2 gene expression and the inhibition was not result of suppression on COX-2 enzyme activity. Previous study demonstrated that decrease of NO production by L -arginine analogs effectively stimulated LPS-induced iNOS gene expression, and proposed that stimulatory effects on iNOS protein by NOS inhibitors might be harmful in treating sepsis. In this study, NLA or L -NAME treatment stimulated significantly on LPS-induced iNOS (but not COX-2) protein in RAW 264.7 cells which was inhibited by these three compounds. Quercetin pentaacetate, but not quercetin and rutin, showed the strong inhibitory activity on PGE2 production and COX-2 protein expression in NLA/LPS or L -NAME/LPS co-treated RAW 264.7 cells. These results indicated that combinatorial treatment of L -arginine analogs and flavonoid derivates, such as quercetin pentaacetate, effectively inhibited LPS-induced NO and PGE2 productions, at the same time, inhibited enhanced expressions of iNOS and COX-2 genes. J. Cell. Biochem. 82: 537,548, 2001. © 2001 Wiley-Liss, Inc. [source] Simvastatin effects on portal-systemic collaterals of portal hypertensive ratsJOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY, Issue 8 2010Hui-Chun Huang Abstract Background and Aim:, Portal-systemic collateral vascular resistance and vasoconstrictor responsiveness are crucial in portal hypertension and variceal bleeding control. Statins enhance vasodilators production, but their influence on collaterals is unknown. This study aimed to survey the effect of simvastatin on collaterals. Methods:, Partially portal vein-ligated rats received oral simvastatin (20 mg/kg/day) or distilled water from ,2 to +7 day of ligation. After hemodynamic measurements on the eighth postoperative day, baseline perfusion pressure (i.e. an index of collateral vascular resistance) and arginine vasopressin (AVP, 0.1 nM,0.1 µM) responsiveness were evaluated with an in situ perfusion model for collateral vascular beds. RT-PCR of endothelial NO synthase (eNOS), inducible NOS (iNOS), cyclooxygenase-1 (COX-1), COX-2, thromboxane A2 synthase (TXA2 -S) and prostacyclin synthase genes was performed in parallel groups for splenorenal shunt (SRS), the most prominent intra-abdominal collateral vessel. To determine the acute effects of simvastatin, collateral AVP response was assessed with vehicle or simvastatin. SRS RT-PCR of eNOS, iNOS, COX-1, COX-2 and TXA2 -S, and measurements of perfusate nitrite/nitrate, 6-keto-PGF1, and TXB2 levels were performed in parallel groups without AVP. Results:, Acute simvastatin administration enhanced SRS eNOS expression and elevated perfusate nitrite/nitrate and 6-keto-PGF1, concentrations. Chronic simvastatin treatment reduced baseline collateral vascular resistance and portal pressure and enhanced SRS eNOS, COX-2 and TXA2 -S mRNA expression. Neither acute nor chronic simvastatin administration influenced collateral AVP responsiveness. Conclusion:, Simvastatin reduces portal-systemic collateral vascular resistance and portal pressure in portal hypertensive rats. This may be related to the enhanced portal-systemic collateral vascular NO and prostacyclin activities. [source] Local isoform-specific NOS inhibition: A promising approach to promote motor function recovery after nerve injuryJOURNAL OF NEUROSCIENCE RESEARCH, Issue 9 2010Bernardo Moreno-López Abstract Physical injury to a nerve is the most frequent cause of acquired peripheral neuropathy, which is responsible for loss of motor, sensory and/or autonomic functions. Injured axons in the peripheral nervous system maintain the capacity to regenerate in adult mammals. However, after nerve transection, stumps of damaged nerves must be surgically joined to guide regenerating axons into the distal nerve stump. Even so, severe functional limitations persist after restorative surgery. Therefore, the identification of molecules that regulate degenerative and regenerative processes is indispensable in developing therapeutic tools to accelerate and improve functional recovery. Here, I consider the role of nitric oxide (NO) synthesized by the three major isoforms of NO synthases (NOS) in motor neuropathy. Neuronal NOS (nNOS) seems to be the primary source of NO that is detrimental to the survival of injured motoneurons. Endothelial NOS (eNOS) appears to be the major source of NO that interferes with axonal regrowth, at least soon after injury. Finally, NO derived from inducible NOS (iNOS) or nNOS is critical to the process of lipid breakdown for Wallerian degeneration and thereby benefits axonal regrowth. Specific inhibitors of these isoforms can be used to protect injured neurons from degeneration and promote axonal regeneration. A cautious proposal for the treatment of acquired motor neuropathy using therapeutic tools that locally interfere with eNOS/nNOS activities seems to merit consideration. © 2010 Wiley-Liss, Inc. [source] The neuronal apoptotic death in global cerebral ischemia in gerbil: Important role for sodium channel modulator,JOURNAL OF NEUROSCIENCE RESEARCH, Issue 6 2009Manoja Kumar Brahma Abstract Global ischemia was induced in gerbil by bilateral occlusion of the common carotid arteries for 5 min. Sodium ionophore monensin or sodium channel blocker tetrodotoxin (TTX) was administered at doses of 10 ,g/kg, i.p., 30 min before ischemia induction; the dose was repeated after 22 hr. Subsequently, brain infarct occurred, determined at 24 hr after occlusion. Large, well-demarcated infarcts were observed in both hemispheres, an important observation because it critically influences the interpretation of the data. Because nitric oxide (NO) production is thought to be related to ischemic neuronal damage, we examined increases in Ca2+ influx, which lead to the activation of nitric oxide synthase (NOS). Then we evaluated the contributions of neuronal NOS, endothelial NOS, and inducible NOS to NO production in brain cryosections. The cytosolic release of apoptogenic molecules like cytochrome c and p53 were confirmed after 24 hr of reflow. TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labeling) labeling detected the apoptotic cells, which were confirmed in neuron-rich cell populations. After 24 hr, all the ischemic changes were amplified by monensin and significantly attenuated by TTX treatment. Additionally, the nesting behavior and histological outcomes were examined after 7 day of reflow. The neuronal damage in the hippocampal area and significant decrease in nesting scores were observed with monensin treatment and reduced by TTX pretreatment after day 7 of reflow. To our knowledge, this report is the first to highlight the involvement of the voltage-sensitive Na+ channel in possibly regulating in part NO system and apoptosis in a cytochrome c,dependent manner in global ischemia in the gerbil, and thus warrants further investigation. © 2008 Wiley-Liss, Inc. [source] Loss of lipopolysaccharide-induced nitric oxide production and inducible nitric oxide synthase expression in scrapie-infected N2a cellsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2003Heléne Lindegren Abstract In scrapie-infected cells, the conversion of the cellular prion protein to the pathogenic prion has been shown to occur in lipid rafts, which are suggested to function as signal transduction platforms. Neuronal cells may respond to bacterial lipopolysaccharide (LPS) treatment with a sustained and elevated nitric oxide (NO) release. Because prions and the major LPS receptor CD14 are colocalized in lipid rafts, the LPS-induced NO production in scrapie-infected neuroblastoma cells was studied. This study shows that LPS induces a dose- and time-dependent increase in NO release in the murine neuroblastoma cell line N2a, with a 50-fold increase in NO production at 1 ,g/ml LPS after 96 hr, as measured by nitrite in the medium. This massive NO release was not caused by activation of the neuronal NO synthase (nNOS), but by increased expression of the inducible NOS (iNOS) mRNA and protein. However, in scrapie-infected N2a cells (ScN2a), the LPS-induced NO production was completely abolished. The absence of LPS-induced NO production in ScN2a was due not to abolished enzymatic activity of iNOS but to a complete inhibition of the LPS-induced iNOS gene expression as measured by Western blot and RT-PCR. These results indicate that scrapie infection inhibits the LPS-mediated signal transduction upstream of the transcriptional step in the signaling cascade and may reflect the important molecular and cellular changes induced by scrapie infection. © 2002 Wiley-Liss, Inc. [source] In-vitro anti-inflammatory activity of Pinus sylvestris and Plantago lanceolata extracts: effect on inducible NOS, COX-1, COX-2 and their products in J774A.1 murine macrophagesJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 3 2005E. Vigo Extracts of the plant species Pinus sylvestris L. and Plantago lanceolata L. have been used in traditional medicine for the treatment of certain respiratory diseases, but little is known about their precise effects and mechanisms of action. In this study, we investigated the effect of these plant extracts on the production of nitric oxide (NO) and prostaglandin E2, NO synthase (NOS) type II, cyclooxygenase-1 (COX-1) and COX-2 mRNA expression in the murine macrophage cell line J774A.1. We found that Pinus sylvestris and Plantago lanceolata extracts inhibited NO production in a concentration-dependent manner in this cell line, without obvious cytotoxic effects as tested by MTT assay. The Plantago lanceolata extract at all doses used, and the Pinus sylvestris extract at high doses, showed significant scavenging of NO radicals released by the NO donor PAPA-NONOate. Our data also show that pre-treatment with these extracts significantly inhibits inducible NOS (iNOS) mRNA production in this cell line, without affecting COX-1 mRNA expression. COX-2 mRNA levels and PGE2 levels induced by lipopolysaccharide/interferon-, were not modified upon pre-treatment with the extracts. Thus, our results suggest that the anti-inflammatory properties of Pinus sylvestris and Plantago lanceolata extracts may reflect decreased NO production, possibly due to inhibitory effects on iNOS gene expression or to NO-scavenging activity. [source] Endothelial, but not the inducible, nitric oxide synthase is detectable in normal and portal hypertensive ratsLIVER INTERNATIONAL, Issue 6 2002Michael Martin Stumm Abstract:Background: Chronic portal hypertension is accompanied by a nitric oxide (NO) dependent vasodilation. Three isoforms of NO producing synthases (NOS) are characterized: neuronal NOS (nNOS), endothelial NOS (eNOS) and inducible NOS (iNOS). Sources of increased NO levels in chronic hypertension is disputed. Methods: To determine eNOS and iNOS expression in different organs of portal hypertensive and control rats, we divided Sprague-Dawley rats in 6 groups: (1) Partial portal vein ligated rats, (2) Bile duct ligated rats, (3) Carbon tetrachloride treated rats, (4) Sham operated rats, (5) Untreated control rats, and (6) LPS treated rats. Immunohistochemistry (IHC) and immunoblotting (IB) using antibodies against eNOS or iNOS were carried out on samples from thymus, aorta, heart, lung, oesophagus, liver, spleen, kidney, pancreas, small and large intestine. Results: IHC revealed an even eNOS expression in all groups. Expression of iNOS was restricted to macrophages in organs of LPS treated and the thymus of rats. IB mirrored these results. Conclusion: In chronic portal hypertension, the main source for NO production depends on eNOS activity. [source] Multi-frequency EPR and Mössbauer spectroscopic studies on freeze-quenched reaction intermediates of nitric oxide synthase ,MAGNETIC RESONANCE IN CHEMISTRY, Issue S1 2005C. Jung Abstract It is believed by analogy to chloroperoxidase (CPO) from Caldariomyces fumago that the electronic structure of the intermediate iron-oxo species in the catalytic cycle of nitric oxide synthase (NOS) corresponds to an iron(IV) porphyrin- , -cation radical. Such species can also be produced by the reaction of ferric NOS with external oxidants within the shunt pathway. We present multi-frequency EPR (9.6, 94, 285 GHz) and Mössbauer spectroscopic studies on freeze-quenched intermediates of the oxygenase domain of nitric oxide synthase which has reacted with peroxy acetic acid within 8,200 ms. The intermediates of the oxygenase domain of both the cytokine inducible NOS (iNOSox) and the neuronal NOS (nNOSox) show an organic radical signal in the 9.6-GHz spectrum overlapping with the spectrum of an unknown species with g -values of 2.24, 2.23 and 1.96. Using 94- and 285-GHz EPR the organic radical signal is assigned to a tyrosine radical on the basis of g -values (i.e. Tyr*562 in nNOSox and Tyr*341 in iNOSox). Mössbauer spectroscopy of 57Fe-labeled unreacted nNOSox shows a ferric low-spin heme-iron (, = 0.38 mms,1, ,EQ = 2.58 mms,1). The reaction of nNOSox with peroxy acetic acid for 8 ms leads to the disappearance of the magnetic background characteristic for native nNOSox and a new species with , = 0.27 mms,1 and ,EQ = 2.41 mms,1 is detected at 4.2 K which does not resemble the parameters typical for a Fe(IV) center. It is proposed that this intermediate species corresponds to a ferric low-spin species which magnetically couples to an amino acid radical (presumably Trp*409). Copyright © 2005 John Wiley & Sons, Ltd. [source] NO message from muscleMICROSCOPY RESEARCH AND TECHNIQUE, Issue 3 2001Zarko Grozdanovic Abstract The synthesis of the free radical gas nitric oxide (NO) is catalyzed by the enzyme NO synthase (NOS). NOS converts arginine and molecular oxygen to NO and citrulline in a reaction that requires NADPH, FAD, FMN, and tetrahydrobiopterin as cofactors. Three types of NOS have been identified by molecular cloning. The activity of the constitutively expressed neuronal NOS (nNOS) and endothelial NOS (eNOS) is Ca2+/calmodulin-dependent, whereas that the inducible NOS (iNOS) is Ca2+ -insensitive. The predominant NOS isoform in skeletal muscle is nNOS. It is present at the sarcolemma of both extra- and intrafusal muscle fibers. An accentuated accumulation of nNOS is found in the endplate area. This strict sarcolemmal localization of nNOS is due its association with the dystrophin-glycoprotein complex, which is mediated by the syntrophins. The activity of nNOS in skeletal muscle is regulated by developmental, myogenic, and neurogenic influences. NO exerts several distinct effects on various aspects of skeletal muscle function, such as excitation-contraction coupling, mitochondrial energy production, glucose metabolism, and autoregulation of blood flow. Inside the striated muscle fibers, NO interacts directly with several classes of proteins, such as soluble guanylate cyclase, ryanodine receptor, sarcoplasmic reticulum Ca2+ -ATPase, glyceraldehyde-3-phosphate dehydrogenase, and mitochondrial respiratory chain complexes, as well as radical oxygen species. In addition, NO produced and released by contracting muscle fibers diffuses to nearby arterioles where it acts to inhibit reflex sympathetic vasoconstriction. Microsc. Res. Tech. 55:148,153, 2001. © 2001 Wiley-Liss, Inc. [source] 6-Shogaol suppressed lipopolysaccharide-induced up-expression of iNOS and COX-2 in murine macrophagesMOLECULAR NUTRITION & FOOD RESEARCH (FORMERLY NAHRUNG/FOOD), Issue 12 2008Min-Hsiung Pan Abstract Ginger, the rhizome of Zingiber officinale, is a traditional medicine with carminative effect, antinausea, anti-inflammatory, and anticarcinogenic properties. In this study, we investigated the inhibitory effects of 6-shogaol and a related compound, 6-gingerol, on the induction of nitric oxide synthase (NOS) and cyclooxygenase-2 (COX-2) in murine RAW 264.7 cells activated with LPS. Western blotting and reverse transcription-PCR analyses demonstrated that 6-shogaol significantly blocked protein and mRNA expression of inducible NOS (iNOS) and COX-2 in LPS-induced macrophages. The in vivo anti-inflammatory activity was evaluated by a topical 12- O -tetradecanoylphorbol 13-acetate (TPA) application to mouse skin. When applied topically onto the shaven backs of mice prior to TPA, 6-shogaol markedly inhibited the expression of iNOS and COX-2 proteins. Treatment with 6-shogaol resulted in the reduction of LPS-induced nuclear translocation of nuclear factor-,B (NF,B) subunit and the dependent transcriptional activity of NF,B by blocking phosphorylation of inhibitor ,B (I,B), and p65 and subsequent degradation of I,B,. Transient transfection experiments using NF,B reporter constructs indicated that 6-shogaol inhibits the transcriptional activity of NF,B in LPS-stimulated mouse macrophages. We found that 6-shogaol also inhibited LPS-induced activation of PI3K/Akt and extracellular signal-regulated kinase 1/2, but not p38 mitogen-activated protein kinase (MAPK). Taken together, these results show that 6-shogaol downregulates inflammatory iNOS and COX-2 gene expression in macrophages by inhibiting the activation of NF,B by interfering with the activation PI3K/Akt/I,B kinases IKK and MAPK. [source] AQP1 and AQP3, Psoriasin, and Nitric Oxide Synthases 1,3 are Inflammatory Mediators in Erythema Toxicum NeonatorumPEDIATRIC DERMATOLOGY, Issue 5 2003Giovanna Marchini M.D., Ph.D. Its etiology and physiologic significance are still unclear. The purpose of this study was to extend the search for possible inflammatory mediators of the rash. We performed immunohistochemistry on punch biopsy cryosections from lesions of four, 1-day-old infants and from four matched controls without rash, using antibodies against the water channel proteins aquaporin-1 (AQP1) and aquaporin-3 (AQP3), psoriasin, and the nitric oxide synthase (NOS) enzymes, neuronal NOS (nNOS), inducible NOS (iNOS), and endothelial NOS (eNOS). All sections from the lesions showed a dense, nodular cellular infiltrate located near the hair follicle. The vessels in the dermis showed a high incidence of AQP1 and eNOS. Strong staining for AQP1, AQP3, and psoriasin, as well as nNOS, iNOS, and eNOS were seen in the entire epidermal layer. The infiltrate in the dermis contained numerous cells expressing AQP1, AQP3, nNOS, iNOS, and eNOS. Double immunofluorescence staining showed that AQP3 was located in CD1a-expressing Langerhans cells and other dendritic cells in the dermis, as well as in CD14-expressing macrophages, CD15-expressing neutrophils, and EG2-expressing eosinophils surrounding the hair follicle. Our findings show that AQP1 and AQP3, psoriasin, and NOSs are involved in the activation of the skin immune system at birth. [source] Inducible and endothelial constitutive nitric oxide synthase gene polymorphisms in Kawasaki diseasePEDIATRICS INTERNATIONAL, Issue 2 2003Vahid Khajoee AbstractBackground: Nitric oxide (NO) is secreted by immune and vascular endothelial cells, and appears to play important roles in the pathophysiology of Kawasaki disease (KD). Thus, genetic variations in NO synthase (NOS) genes may be involved in the development of coronary artery lesions (CAL) in KD. Methods: The present study investigated the association of endothelial constitutive NOS (ecNOS) and inducible NOS (iNOS) gene polymorphisms with the development of CAL in KD in a Japanese population. Results: The genotype distributions of 27-bp tandem repeat polymorphism within intron 4 of ecNOS gene did not show any significant difference between controls and KD patients with or without CAL. In addition, there was no significant association between whole-allele distribution of iNOS gene promoter (penta-repeat CCTTT) polymorphism and KD with or without CAL. Conclusion: These results did not support any association of ecNOS and iNOS gene polymorphisms to the development of CAL in KD patients in a Japanese population. [source] Lung eNOS and iNOS are Reoxygenation Time-Dependent Upregulated After Acute HypoxiaTHE ANATOMICAL RECORD : ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY, Issue 6 2010Alma Rus Abstract Nitric oxide plays a critical role in many physiological and physiopathological processes in the lung. Changes in the NO/NOS (Nitric Oxide/Nitric Oxide Synthase) system after hypoxia situations remain controversial in this organ, so that the aim of this work is to perform a complete study of this system in the hypoxic lung after different reoxygenation times ranging from 0 h to 5 days posthypoxia. This is a novel follow-up study carried out in Wistar rats submitted for 30 min to acute hypobaric hypoxia. We measured endothelial and inducible NOS (eNOS, iNOS) mRNA and protein expression, location, and in situ NOS activity as well as nitrated protein expression and location. In addition, NO levels were indirectly quantified (NOx) as well as the apoptosis level. Results showed an increase in eNOS mRNA, protein, activity as well as eNOS positive immunostaining at 0 h posthypoxia, coinciding with raised NOx levels. Contrary, iNOS, nitrated protein expression and apoptosis level augmented during the final reoxygenation times. The lung NO/NOS system provokes two responses to the hypoxia/reoxygenation processes: (i) eNOS is responsible of the immediate response, producing NO, which causes vasodilation and bronchodilation, and (ii) iNOS is related to the second late response, which seems to be involved in some of the deleterious consequences that hypoxia induces in the lung. Anat Rec, 2010. © 2010 Wiley-Liss, Inc. [source] Post-translational Regulation of Endothelial Nitric Oxide Synthase (eNOS) by Estrogens in the Rat VaginaTHE JOURNAL OF SEXUAL MEDICINE, Issue 5 2010Biljana Musicki PhD ABSTRACT Introduction., Estrogens control vaginal blood flow during female sexual arousal mostly through nitric oxide (NO). Although vascular effects of estrogens are attributed to an increase in endothelial NO production, the mechanisms of endothelial NO synthase (eNOS) regulation by estrogens in the vagina are largely unknown. Aims., Our hypothesis was that estrogens regulate eNOS post-translationally in the vagina, providing a mechanism to affect NO bioavailability without changes in eNOS protein expression. Methods., We measured eNOS phosphorylation and eNOS interaction with caveolin-1 and heat shock protein 90 (HSP90) in the distal and proximal vagina of female rats at diestrus, 7 days after ovariectomy and 2 days after replacement of ovariectomized rats with estradiol-17, (15 µg). Main Outcome Measures., Molecular mechanisms of eNOS regulation by estrogen in the rat vagina. Results., We localized phospho-eNOS (Ser-1177) immunohistochemically to the endothelium lining blood vessels and vaginal sinusoids. Estrogen withdrawal decreased phosphorylation of eNOS on its positive regulatory site (Ser-1177) and increased eNOS binding to its negative regulator caveolin-1 (without affecting eNOS/HSP90 interaction), and they were both normalized by estradiol replacement. Protein expressions of phosphorylated Akt (protein kinase B) and extracellular signal-regulated protein kinase 1/2 (ERK1/2) were not affected by estrogen status, suggesting that the effect of estrogens on eNOS (Ser-1177) phosphorylation was not mediated by activated AKT or ERK1/2. eNOS phosphorylation on its negative regulatory site (Ser-114) was increased in the vagina by estrogen withdrawal and normalized by estradiol replacement, implying that the maintenance of low phosphorylation of eNOS on this site by estradiol may limit eNOS interaction with caveolin-1 and preserve the enzyme's activity. Total eNOS, inducible NOS, caveolin-1, and HSP90 protein expressions were not affected by ovariectomy or estradiol replacement in the distal or proximal vagina. Conclusions., These results define novel estrogen signaling mechanisms in the vagina which involve eNOS phosphorylation and eNOS,caveolin-1 interaction. Musicki B, Liu T, Strong TD, Lagoda GA, Bivalacqua TJ, and Burnett AL. Post-translational regulation of endothelial nitric oxide synthase (eNOS) by estrogens in the rat vagina. J Sex Med 2010;7:1768,1777. [source] NADPH-Diaphorase Activity and NO Synthase Expression in the Olfactory Epithelium of the BovineANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 3 2010S. Wenisch With 2 figures Summary NADPH-diaphorase (NADPH-d) staining of the bovine olfactory epithelium was compared with the immunohistochemical localization of nitric oxide synthase (NOS), soluble guanylyl cyclase, and cGMP (cyclic guanosine 3,,5,-monophosphate). Out of the three isoforms, only the inducible NOS (NOS-II) was found at the epithelial surface correlating with the strong labelling for NADPH-d. In contrast, light diaphorase staining associated with deeper epithelial regions did not coincide with any NOS immunoreactivity. As there is overlapping expression of NOS-II, soluble guanylyl cyclase and cGMP at the luminal surface morphologically occupied by dendritic knobs of olfactory receptor neurons and microvillar endings of supporting cells, the nitric oxide (NO)/cGMP pathway is likely to be involved in modulating the odour signals during olfactory transduction. [source] Experimental ischaemia-reperfusion injury induces vascular endothelial growth factor expression in the rat testisANDROLOGIA, Issue 4 2009H. Hashimoto Summary Testicular torsion causes ischaemia-reperfusion (I-R) injury of testis and might lead to male infertility. Its injury initiates a pathophysiological cascade, including an activation of inflammatory cytokines and generation of nitric oxide and other reactive oxygen species. Vascular endothelial growth factor (VEGF) mediates angiogenesis and promotes endothelial cell survival. The aim of our study was to investigate the time course expression of VEGF, VEGF-receptor (R)1, VEGF-R2, nitric oxide synthases (NOS) in experimental I-R injury of rat testis. In torsion side testis, the expression of VEGF protein and mRNA significantly increased in a time-dependent manner (P < 0.001 and P < 0.001, respectively). Although the expression of VEGF-R1 mRNA was increased in a similar way (P < 0.001), VEGF-R2 mRNA expression was not detected. In immunohistochemistry, the increase in VEGF protein staining was observed in testicular vascular endothelial cells and germ cells at 24 h after reperfusion. Significant activation of inducible NOS and endothelial NOS was investigated at 12 and 24 h after reperfusion (P < 0.01 and P < 0.001, respectively). This is the first report to show the time course expression of VEGF in experimental I-R rat testis. [source] Inducible nitric oxide synthase expression and its prognostic significance in colorectal cancerAPMIS, Issue 2 2010KYRIAKOS ZAFIRELLIS Zafirellis K, Zachaki A, Agrogiannis G, Gravani K. Inducible nitric oxide synthase expression and its prognostic significance in colorectal cancer. APMIS 2010; 118: 115,24. Nitric oxide synthases (NOS) are expressed in colorectal cancer. The aim of this study was to examine the inducible NOS (iNOS) expression in colorectal cancer and to investigate its prognostic relevance. Tissue sections of primary tumors from 132 patients undergoing curative resection for colorectal cancer were immunohistochemically examined for iNOS expression. The expression pattern of iNOS was correlated with various clinicopathological characteristics and survival. iNOS immunoreactivity was observed in the cytoplasm of tumor epithelial cells in 60 patients (45.5%) and positively correlated with lymph node involvement (p = 0.019). No significant correlation was found between iNOS expression and various clinicopathological characteristics, including age, gender, tumor location, tumor size, tumor grade, T stage, and Union International Contra la Cancrum (UICC) stage. Survival analysis showed a significant correlation between iNOS-positive tumors and poor disease-specific survival (p < 0.0001), with independent prognostic significance in multivariate analysis (HR = 4.42; p < 0.0001). Patients with stage II disease and iNOS-positive tumors had significantly worse disease-specific survival than those with iNOS-negative tumors (p < 0.0001). In addition, patients with stage III disease and iNOS-positive tumors had significantly worse disease-specific survival than those with iNOS-negative tumors (p = 0.001). The ability of iNOS to predict outcome in colorectal cancer patients may be independent of other known prognostic factors, providing a new molecular marker with significant potential for clinical utility. [source] Differential expression of nitric oxide synthases in human scalp epidermal and hair follicle pigmentary units: implications for regulation of melanogenesisBRITISH JOURNAL OF DERMATOLOGY, Issue 2 2005H.M. Sowden Summary Background, Nitric oxide (NO) is a ubiquitous gaseous lipophilic molecule generated from the conversion of l -arginine to l -citrulline by the NO synthases (NOSs). Ultraviolet radiation (UVR)-induced NO production appears to stimulate epidermal melanogenesis. However, given their relative protection from UVR, it is unclear whether NO plays a similar role in hair bulb melanocytes. Objectives, We aimed to identify the expression profiles of the NOS isoforms endothelial NOS (eNOS), neuronal NOS (nNOS) and inducible NOS (iNOS) and of phosphorylated eNOS and nitrotyrosine within the epidermal and follicular melanin units of normal human haired scalp during the hair growth cycle. Methods, This study employed single and double immunohistochemical and immunofluorescence staining techniques using haired scalp from 10 healthy individuals (six women and four men). Results, Melanocytes in the basal layer of the epidermis expressed eNOS, nNOS and nitrotyrosine. By contrast, melanogenically active melanocytes of the anagen hair bulb were wholly negative for these markers. However, other follicular melanocytes not actively involved in pigment production, including undifferentiated melanocytes located in the outer root sheath and melanocytes surviving the apoptosis-driven hair follicle (HF) regression during catagen/telogen, expressed eNOS, nNOS and nitrotyrosine. While iNOS was only weakly expressed in the basal layer of the human epidermis, it was highly expressed in keratinocytes of the inner root sheath (IRS), where it colocalized with trichohyalin, a differentiation-associated protein of the IRS that requires enzyme-catalysed conversion of arginine to citrulline. Conclusions, The NOS isoforms and nitrotyrosine are differentially expressed in different cutaneous melanocyte subpopulations. Results of this study suggest a possible role for eNOS, nNOS, iNOS and nitrotyrosine in melanocyte biology, particularly with respect to melanogenesis and melanocyte survival during HF regression. Another example of possible NO involvement in HF biology is the postsynthetic modification of trichohyalin in differentiating keratinocytes of the IRS. These results suggest that NO may influence several aspects of HF biology. [source] Gastric mucosal resistance to acute injury in experimental portal hypertensionBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2001Sara Calatayud The gastric mucosa of portal hypertensive rats exhibits important microvascular changes and a nitric oxide (NO)-dependent hyperemia. This study analyses whether portal hypertensive mucosa exhibits changes in its ability to withstand aggression. Portal hypertension was induced by partial portal vein ligation (PPVL) or common bile duct ligation (CBDL) and gastric damage was induced by oral administration of ethanol or aspirin. Experiments were performed in conscious or anaesthetized rats and some animals were pre-treated with the NO-synthesis inhibitor L -NAME. Conscious PPVL or CBDL rats showed an increased resistance to the damaging effects of ethanol. Oral administration of aspirin produced less gastric damage in PPVL conscious rats than in the control group. The protective effects of portal hypertension were maintained in animals anaesthetized with ketamine and absent when pentobarbital was employed. Pre-treatment with L -NAME restored the damaging effects of ethanol and aspirin in PPVL rats without modifying the level of damage in control animals. Gastric bleeding induced by oral aspirin, as measured by the luminal release of 51Cr-labelled erythrocytes, was significantly greater in PPVL rats than in control animals. Semi-quantitiative analysis by RT , PCR of the mRNA for endothelial NO-synthase (eNOS), neuronal NOS (nNOS) and inducible NOS (iNOS) levels showed that the expression of iNOS was slightly increased in both the gastric mucosa and smooth muscle of PPVL rats. No changes were observed in eNOS and nNOS expression. Conscious portal hypertensive rats exhibit an enhanced resistance to acute gastric damage which is absent under the influence of some types of anaesthesia and seems related to an increased synthesis of nitric oxide. However, mucosal lesions in these animals show an augmented bleeding per area of injury. British Journal of Pharmacology (2001) 132, 309,317; doi:10.1038/sj.bjp.0703785 [source] Increased nitric oxide production in nasal epithelial cells from allergic patients , RT-PCR analysis and direct imaging by a fluorescence indicator: DAF-2 DA*CLINICAL & EXPERIMENTAL ALLERGY, Issue 6 2001S. Takeno Background Nitric oxide (NO) is believed to participate in the regulation of airway clearance and non-specific cellular immunity. Recent studies have suggested that airway epithelial cells of allergic and non-allergic individuals may differ in their ability to produce this molecule. Objective The aim of this study was to detect the difference in NO production in human nasal epithelial cells between normal subjects and patients with perennial allergic rhinitis (AR), and to assess the relationship between the expression of nitric oxide synthase (NOS) isoforms and the severity of the disease. Methods Nasal epithelial cells were obtained from the inferior turbinate. The expression of mRNAs encoding constitutive endothelial NOS (eNOS) and inducible NOS (iNOS) was studied by reverse transcription-polymerase chain reaction (RT-PCR). Direct NO production in living cells was visualized and quantified by a fluorescent indicator, DAF-2 DA. Results RT-PCR analysis demonstrated that AR patients with a RAST score of 5 or 6 showed significant increases in the levels of iNOS mRNA and slight reductions in those of eNOS mRNA. Patients with a RAST score of 2,4 also revealed the same tendency however, the difference was not significant. DAF-2 DA imaging demonstrated that epithelial cells, especially the ciliated cells, produced a larger amount of NO than non-epithelial inflammatory cells. Preincubation with L-NAME resulted in an approximate 40% decrease in both groups. Conclusion These results directly indicate that nasal epithelial cells of AR patients overall produce higher levels of NO through the concomitant expression of different NOS isoforms. Continuous NO production by the epithelial cells in normal subjects further support the hypothesis that NO derived from epithelium may play dual roles in the regulation of nasal airway clearance and in the host defense. In addition, the use of DAF-2 DA provides a reliable method to visualize and quantify the direct NO production of living cells. [source] | ||||||||||||||||||||||||||||