Collecting duct-derived endothelin regulates arterial pressure and Na excretion via nitric oxide

Mice with a collecting duct-specific deletion of endothelin-1 are hypertensive and have impaired Na excretion. Because endothelin-1 activates NO synthase (NOS) in the collecting duct, we hypothesized that impaired renal NO production in knockout mice exacerbates the hypertensive state. Control and knockout mice were treated chronically with N(G)-nitro-l-arginine methyl ester, and blood pressure (BP) and urinary nitrate/nitrite excretion were assessed. On a normal Na diet, knockout systolic BP was 18 mm Hg greater than in controls. N(G)-nitro-l-arginine methyl ester increased BP in control mice by 30 mm Hg and 10 mm Hg in collecting duct-specific deletion of endothelin-1 knockout mice, thereby abolishing the difference in systolic BP between the groups. A high-Na diet increased BP similarly in both groups. Urinary nitrate/nitrite excretion was lower in knockout mice than in controls on normal or high Na intake. In separate experiments, renal perfusion pressure was adjusted in anesthetized mice, and urinary nitrate/nitrite and Na excretion were determined. Similar elevations of BP increased urinary Na and nitrate/nitrite excretion in control mice but to a significantly lesser extent in knockout mice. Isoform-specific NOS activity and expression were determined in renal inner medulla homogenates from control and knockout mice. NOS1 and NOS3 activities were lower in knockout than in control mice given normal or high-Na diets. However, NOS1 or NOS3 protein expressions were similar in both groups on normal or high-Na intake. These data demonstrate that collecting duct-derived endothelin-1 is important in the following: (1) chronic N(G)-nitro-l-arginine methyl ester-induced hypertension; (2) full expression of pressure-dependent changes in sodium excretion; and (3) control of inner medullary NOS1 and NOS3 activity.     

Expression of Nitric Oxide Synthase Isoforms and Nitric Oxide Production in Acute Pancreatitis and Associated Lung Injury

Background/Aims: The role of nitric oxide (NO) has been increasingly implicated in the pathophysiology of acute pancreatitis (AP). Studies have shown increased NO production in AP although not all are agreeable on whether NO is beneficial or detrimental in AP. This study aims to profile NO production and NO synthase (NOS) expression in the pancreas and lungs in the progression of AP in mice to gain insights to the role played by different NOS isoforms. Methods: AP was induced in mice by hourly administration of cerulein. NO production was determined by measuring the total nitrite and nitrate (NOx) content while NOS expression was measured by Western blot. Results: Pancreatic NO production increased sharply and was sustained throughout AP. iNOS expression was greatly increased while eNOS was downregulated at the later stages. In the lungs, there was an unexpected early increase in the constitutive NOS expression; however iNOS was also significantly overexpressed at the later time point along with a significant increase in NO. Acinar cells were found to overproduce NO in response to cerulein hyperStimulation with iNOS again being the major contributor. Conclusion: These data show that NO production and NOS expression are differentially regulated temporally and in magnitude in the pancreas and lungs in response to cerulein hyperstimulation which suggests differing roles for each NOS isoform.     

Effect of antioxidant therapy on blood pressure and NO synthase expression in hypertensive rats

Earlier studies have demonstrated evidence for increased reactive oxygen species, enhanced NO synthase (NOS) expression, and elevated NO production in spontaneously hypertensive rats (SHR). Given the negative-feedback regulation of NOS by NO, we hypothesized that enhanced NO inactivation by ROS may contribute to compensatory upregulation of NOS in SHR. The present study was designed to test this hypothesis. Eight-week-old male SHR and Wistar-Kyoto rats were treated for 3 weeks with either a placebo or the potent antioxidant, lazaroid (desmethyltirilazad, 10 mg. kg(-1). d(-1), by gastric gavage). Tail arterial blood pressure, urinary excretion of NO metabolites (ie, nitrate and nitrite), and immunodetectable NOS isotype proteins in the vascular, renal, cardiac, and cerebral tissues were measured. The placebo-treated SHR group showed a marked elevation of blood pressure and a significant upregulation of aorta, kidney, and cardiac tissue endothelial and inducible NOS (eNOS and iNOS, respectively) proteins and of brain and renal tissue neuronal NOS. Lazaroid therapy ameliorated hypertension and mitigated the upregulation of eNOS and iNOS in vascular, renal, and cardiac tissues but had limited effect on the expression of renal and brain neuronal NOS. In contrast, lazaroid therapy had no effect on blood pressure, urinary nitrate and nitrite excretion, or tissue NOS isotype expressions in the Wistar-Kyoto group. These findings support the role of oxidative stress in the genesis and/or maintenance of hypertension and compensatory upregulation of the expression of eNOS and iNOS in SHR.     

Tumor necrosis factor–α–induced hypertension in pregnant rats results in decreased renal neuronal nitric oxide synthase expression

BackgroundPreeclampsia is associated with increases in plasma levels of tumor necrosis factor–α (TNF-α), a cytokine known to contribute to endothelial dysfunction. We recently reported that a twofold elevation in plasma TNF-α produces significant reductions in renal function and hypertension in pregnant rats. The purpose of this study was to determine the role of the nitric oxide (NO) system in TNF-α–induced hypertension in pregnant rats.MethodsTumor necrosis factor–α (50 ng/day) was chronically infused starting at day 14 of gestation. Mean arterial pressure, 24-h urinary nitrite/nitrate excretion, and renal nitric oxide synthase (NOS) protein expression by Western blot analysis was measured at day 19 of gestation.ResultsA twofold increase in plasma TNF-α levels in pregnant rats resulted in a significant increase in arterial pressure (97 ± 3.6 v 116 ± 2.1 mm Hg, pregnant versus TNF-α pregnant, respectively, P < .05), but no significant change in urinary nitrite/nitrate excretion (22.0 ± 1.9 v 20.8 ± 2.5 μmol/24 h, pregnant versus TNF-α pregnant, respectively), a measure of whole body NO production. As abnormalities in renal production of NO would not be reflected in the measure of whole body NO production, changes in renal NOS protein levels were determined. The protein expression of both neuronal (nNOS) and inducible (iNOS) nitric oxide synthase were significantly decreased in the medulla of TNF-α pregnant rats (nNOS: 10.6 ± 0.7 v 8.2 ± 0.8 densitometric units, P < .05; and iNOS: 19.2 ± 0.9 v 15.4 ± 0.8 densitometric units, P < .05, pregnant versus TNF-α pregnant, respectively).Conclusion:The hypertension associated with a chronic twofold increase in TNF-α in pregnant rats is associated with significant decreases in renal nNOS and iNOS protein production.     

Nitric oxide production during adjuvant-induced and collagen-induced arthritis

Objective. To investigate the role of nitric oxide (NO) production and NO synthase (NOS) induction during adjuvant-induced arthritis (AIA) and collagen-induced arthritis (CIA) in Dark Agouti rats. Methods. Urinary nitrate excretion and immune NOS (iNOS) messenger RNA (mRNA) expression were measured in the joint, lymph node, spleen, and liver tissues following the induction of either AIA or CIA. Results. Urinary nitrate excretion and iNOS mRNA expression increased substantially during joint inflammation in both models of arthritis. However, the increases in urinary nitrate excretion and iNOS mRNA expression observed in the joint, liver, and spleen tissues during AIA were greater than those observed during CIA, although iNOS induction in the lymph nodes was similar for both models. A prior injection with Mycobacterium bovis heat-shock protein resulted in suppression of arthritis and NO production in AIA, but not in CIA. Conclusion. Differences in NO production during AIA versus CIA are a reflection of the fundamental pathophysiologic differences between these 2 models of arthritis. Thus, NO production in these 2 models could not be merely a nonspecific reaction to the adjuvant injection, nor simply a byproduct of local inflammation in the joint.     

Effects of hydroxyurea and L-arginine on the production of nitric oxide metabolites in cultures of normal and sickle erythrocytes

Previous in vitro studies suggest that erythrocytes may be a source of nitric oxide (NO) produced by nitric oxide synthase (NOS) or by oxyhemoglobin-mediated oxidation of hydroxyurea (HU). This study was performed to determine the roles of HU and NOS in the production of NO by normal and sickle erythrocytes. Red blood cells (RBCs) from normal adult hemoglobin (HbAA) and homozygous sickle cell subjects (HbSS) were incubated with PBS containing 0.2 mM hydrogen peroxide (control) for 2 h at 37 degrees C in the presence and absence of l-arginine, the substrate for NOS, and with l-arginine plus HU in the presence and absence of l-NMMA, a specific inhibitor of NOS. The nitrate and nitrite metabolites of NO, expressed as [NOx], were measured. [NOx] in the HbAA and HbSS RBC cultures was not significantly different in the presence and absence of 1.0 mM l-arginine (p>0.1). [NOx] in the HbAA and HbSS cultures treated with a clinically relevant dose of HU (1.0 mM) plus 1.0 mM l-arginine was significantly greater than that in controls incubated with PBS and with l-arginine p < 0.01. However, [NOx] in the HbAA and HbSS cultures treated with 50 microg/ml l-NMMA was not significantly different than that in the cultures treated with HU plus l-arginine in the absence of l-NMMA. These findings suggest that NOx production by erythrocytes may be increased by treatment with HU and may not be decreased by inhibiting NOS. Therefore, we conclude that a therapeutic dose of HU may increase the plasma concentration of NO by a mechanism that does not require erythrocytes NOS activity.     

Immune Agranulocytosis and Clarithromycin

Previous in vitro studies suggest that erythrocytes may be a source of nitric oxide (NO) produced by nitric oxide synthase (NOS) or by oxyhemoglobin-mediated oxidation of hydroxyurea (HU). This study was performed to determine the roles of HU and NOS in the production of NO by normal and sickle erythrocytes. Red blood cells (RBCs) from normal adult hemoglobin (HbAA) and homozygous sickle cell subjects (HbSS) were incubated with PBS containing 0.2 mM hydrogen peroxide (control) for 2 h at 37°C in the presence and absence of L-arginine, the substrate for NOS, and with L-arginine plus HU in the presence and absence of L-NMMA, a specific inhibitor of NOS. The nitrate and nitrite metabolites of NO, expressed as [NOx], were measured. [NOx] in the HbAA and HbSS RBC cultures was not significantly different in the presence and absence of 1.0 mM L-arginine (p > 0.1). [NOx] in the HbAA and HbSS cultures treated with a clinically relevant dose of HU (1.0 mM) plus 1.0 mM L-arginine was significantly greater than that in controls incubated with PBS and with L-arginine p < 0.01. However, [NOx] in the HbAA and HbSS cultures treated with 50 μg/ml L-NMMA was not significantly different than that in the cultures treated with HU plus L-arginine in the absence of L-NMMA. These findings suggest that NOx production by erythrocytes may be increased by treatment with HU and may not be decreased by inhibiting NOS. Therefore, we conclude that a therapeutic dose of HU may increase the plasma concentration of NO by a mechanism that does not require erythrocytes NOS activity     

Regulation of bone mass and bone turnover by neuronal nitric oxide synthase

Nitric oxide (NO) is produced by NO synthase (NOS) and plays an important role in the regulation of bone cell function. The endothelial NOS isoform is essential for normal osteoblast function, whereas the inducible NOS isoform acts as a mediator of cytokine effects in bone. The role of the neuronal isoform of NOS (nNOS) in bone has been studied little thus far. Therefore, we investigated the role of nNOS in bone metabolism by studying mice with targeted inactivation of the nNOS gene. Bone mineral density (BMD) was significantly higher in nNOS knockout (KO) mice compared with wild-type controls, particularly the trabecular BMD (P < 0.01). The difference in BMD between nNOS KO and control mice was confirmed by histomorphometric analysis, which showed a 67% increase in trabecular bone volume in nNOS KO mice when compared with controls (P < 0.001). This was accompanied by reduced bone remodeling, with a significant reduction in osteoblast numbers and bone formation surfaces and a reduction in osteoclast numbers and bone resorption surfaces. Osteoblasts from nNOS KO mice, however, showed increased levels of alkaline phosphatase and no defects in proliferation or bone nodule formation in vitro, whereas osteoclastogenesis was increased in nNOS KO bone marrow cultures. These studies indicate that nNOS plays a hitherto unrecognized but important physiological role as a stimulator of bone turnover. The low level of nNOS expression in bone and the in vitro behavior of nNOS KO bone cells indicate that these actions are indirect and possibly mediated by a neurogenic relay.     

Activation of inducible nitric oxide synthase/nitric oxide by curli fibers leads to a fall in blood pressure during systemic Escherichia coli infection in mice

Septic shock, a major cause of death, is characterized by a pathophysiologic increased production of nitric oxide (NO), which leads to vasodilation and myocardial toxicity. Septic Escherichia coli frequently express proteinaceous curli fibers. In this study, curliated E. coli induced high levels of NO by directly inducing type 2 nitric oxide synthase (NOS2) both in vitro and in vivo. More severe hypotension and higher plasma nitrite/nitrate levels were seen in wild type mice systemically infected with curliated E. coli than in animals infected with E. coli mutants that lacked curli proteins. Blood pressure remained stable in NOS2-deficient mice with curliated bacteria. Increased heart rates, transient hypothermia, and loss of gross activity were seen in all mice, regardless of curli expression. Study results suggest that expression of curli fibers by E. coli activates the NO/NOS2 arm of the innate immune system, which leads to a significant fall in blood pressure.     

Limited oral bioavailability and active epithelial excretion of paclitaxel (Taxol) caused by P-glycoprotein in the intestine

In mice, the mdr1a and mdr1b genes encode drug-transporting proteins that can cause multidrug resistance in tumor cells by lowering intracellular drug levels. These P-glycoproteins are also found in various normal tissues such as the intestine. Because mdr1b P-glycoprotein is not detectable in the intestine, mice with a homozygously disrupted mdr1a gene [mdr1a(−/−) mice] do not contain functional P-glycoprotein in this organ. We have used these mdr1a(−/−) mice to study the effect of gut P-glycoprotein on the pharmacokinetics of paclitaxel. The area under the plasma concentration-time curves was 2- and 6-fold higher in mdr1a(−/−) mice than in wild-type (wt) mice after i.v. and oral drug administration, respectively. Consequently, the oral bioavailability in mice receiving 10 mg paclitaxel per kg body weight increased from only 11% in wt mice to 35% in mdr1a(−/−) mice. The cumulative fecal excretion (0–96 hr) was markedly reduced from 40% (after i.v. administration) and 87% (after oral administration) of the administered dose in wt mice to below 3% in mdr1a(−/−) mice. Biliary excretion was not significantly different in wt and mdr1a(−/−) mice. Interestingly, after i.v. drug administration of paclitaxel (10 mg/kg) to mice with a cannulated gall bladder, 11% of the dose was recovered within 90 min in the intestinal contents of wt mice vs. <3% in mdr1a(−/−) mice. We conclude that P-glycoprotein limits the oral uptake of paclitaxel and mediates direct excretion of the drug from the systemic circulation into the intestinal lumen.     

Nitric oxide and TNF-α trigger colonic inflammation and carcinogenesis in Helicobacter hepaticus-infected, Rag2-deficient mice

Recombinase-activating gene-2-deficient (Rag2^−/−) mice lacking functional lymphocytes provide a useful model of chronic inflammatory bowel disease-emulating events in human colon cancer. Infection of Rag2^−/− mice with Helicobacter hepaticus led to accumulation of macrophages and neutrophils in the colon, a process temporally related to up-regulation of tissue inducible nitric oxide synthase (iNOS) expression at the site of infection and increased nitric oxide (NO) production, as evidenced by urinary excretion of nitrate. Progressive development of increasingly severe inflammation, hyperplasia, dysplasia, and cancer accompanied these changes. Concurrent administration of an iNOS inhibitor prevented NO production and abrogated epithelial pathology and inhibited the onset of cancer. The presence of Gr-1⁺ neutrophils and elevated tumor necrosis factor-α (TNF-α) expression in colon were required for increased iNOS expression and cancer, whereas interleukin-10 (IL-10) down-regulated TNF-α and iNOS expression and suppressed cancer. Anti-inflammatory CD4⁺ regulatory lymphocytes also down-regulated iNOS and reduced cancer formation. Collectively, these results confirm essential roles for inflammation, increased TNF-α expression, and elevated NO production in colon carcinogenesis.     

Nitric oxide synthase inhibitors attenuate ozone-induced airway inflammation in guinea pigs. Possible role of interleukin-8

Nitric oxide (NO) is increased in exhaled air of asthmatics. We hypothesized that endogenous NO contributes to airway inflammation and hyperresponsiveness, and that interleukin-8 (IL-8) might be involved in this mechanism. In human transformed bronchial epithelial cells in vitro, NO donors increased IL-8 production dose-dependently. In addition, tumor necrosis factor-alpha (TNF-alpha) plus IL-1beta plus interferon-gamma (IFN-gamma) increased IL-8 in culture supernatant of epithelial cells; the combination of NO synthase (NOS) inhibitors, aminoguanidine (AG) plus N(G)-nitro-L-arginine methyl ester (L-NAME) attenuated the cytokine-induced IL-8 production in epithelial cells. In guinea pigs in vivo, ozone exposure induced airway hyperresponsiveness to acetylcholine and increased neutrophils in bronchoalveolar lavage fluid (BALF), and these changes persisted for at least 5 h. Pretreatment with NOS inhibitors had no effect on airway hyperresponsiveness or neutrophil accumulation immediately after ozone, but significantly inhibited the changes 5 h after ozone. NOS inhibitors also attenuated the increases of nitrite/nitrate levels in BALF and the IL-8 mRNA expression in epithelial cells and in neutrophils in guinea pig airways 5 h after ozone. These results suggest that endogenous NO may play an important role in the persistent airway inflammation and hyperresponsiveness after ozone exposure, presumably partly through the upregulation of IL-8.     

Polyphenol-containing azuki bean (Vigna angularis) extract attenuates blood pressure elevation and modulates nitric oxide synthase and caveolin-1 expressions in rats with hypertension

Background and aimsAzuki beans (Vigna angularis) contain polyphenols such as proanthocyanidins that exhibit potential radical scavenging activities. We herein investigated the effects of polyphenol-containing azuki bean extract (ABE) on elevated blood pressure, nitric oxide (NO) production, and expressions of endothelial NO synthase (eNOS), inducible NOS (iNOS), and caveolin-1 proteins in the aorta and kidney of chronically hypertensive rats.Methods and resultsSpontaneously hypertensive rats (SHRs/Izm) with approximately 200 mm Hg systolic blood pressure (SBP) were randomly divided into 2 groups fed either 0% or 0.9% ABE-containing diet. Age-matched normotensive Wistar–Kyoto rats were used as the control. The content of 24-h urinary nitrate/nitrite (NOx) excretion was measured to evaluate NO production. After 8 weeks of treatment, the eNOS, iNOS, and caveolin-1 protein expressions in the aorta and kidney were analyzed by western blotting. The SBP of the ABE-treated SHR was significantly lower than that of the untreated SHR. The level of 24-h urinary NOx excretion was significantly higher in the ABE-treated SHR than in the untreated SHR. The eNOS and iNOS expressions in the aorta and kidney were remarkably upregulated in the untreated SHR but suppressed in the ABE-treated SHR. The vascular and renal caveolin-1 expressions were upregulated in the ABE-treated SHR.ConclusionsABE reduced the elevated blood pressure and increased NO production in long-term treatment. It may be associated with the modulation of eNOS and iNOS protein expressions in the aorta and kidney during the development of hypertension.     

Neuronal nitric oxide synthase is associated with airway obstruction in BALB/c mice exposed to ozone

BACKGROUND: The functional role of nitric oxide (NO) and the various nitric oxide synthase (NOS) isoforms in asthma is controversial. OBJECTIVE: To investigate the role of NO in mice exposed to ozone, three known isoforms of NOS [inducible NOS (iNOS), neuronal NOS (nNOS), and endothelial NOS (eNOS)] were studied. METHODS: The expression of iNOS, nNOS, and eNOS was determined in lung by Western blot analysis after exposure to filtered air and ozone (0.12, 0.5, 1 or 2 ppm) for 3 h. Using barometric whole-body plethysmography and increase in enhanced pause (P(enh)) as an index of airway obstruction, we measured airway responses to ozone exposure. Bronchoalveolar lavage (BAL) was performed. Nitrate and nitrite were measured using a modified Griess reaction. RESULTS: The nitrate concentration in BAL fluid, which indicates the in vivo generation of NO in airways, from the ozone-exposed group was significantly greater than that from the group exposed to filtered air (631.0 +/- 86.4 vs. 152.1 +/- 16.9 micromol/l, p < 0.05). The nitrate concentration in BAL fluid was increased more in mice exposed to 2-ppm ozone than that in mice exposed to filtered air or 0.12-, 0.5-, or 1-ppm ozone. Increases in P(enh) after exposure to ozone or filtered air were significantly higher in the ozone-exposed groups than in the group exposed to filtered air (p < 0.01). Increases in P(enh) were dependent on the ozone concentration. Although the protein levels of eNOS and iNOS determined were within normal levels, the amount of nNOS protein was markedly elevated in airway tissue homogenates of the group exposed to 2-ppm ozone. CONCLUSION: These findings demonstrate that the nNOS isoform may be involved in airway obstruction in mice exposed to ozone.     

Effect of mycophenolate mofetil on severity of nephritis and nitric oxide production in lupus-prone MRL/lpr mice

Mycophenolate mofetil (MMF), an immunosuppressive drug commonly used in organ transplantation, is increasingly being used to treat autoimmune diseases including systemic lupus erythematosus (SLE). Excessive production of nitric oxide (NO) by inducible nitric oxide synthase (iNOS) has been implicated in the pathogenesis of lupus nephritis. We evaluated the effect of MMF on the severity of nephritis and the production of NO in lupus-prone MRL/lpr mice. Eight-week-old female MRL/lpr mice (n = 20) were treated with MMF (100 mg/kg/day) by oral gavage for 12 weeks. Control mice (n = 20) received vehicle on the same schedule. The mice were killed after 12 weeks of treatment. Treatment with MMF significantly decreased the amount of proteinuria, prolonged survival and reduced the histological severity of glomerulonephritis. Urinary nitrite/nitrate excretion in the MMF-treated mice was significantly reduced during the first 8 weeks of treatment. However, by the end of the 12 weeks' treatment period, there was no significant difference between vehicle and MMF-treated mice in terms of urinary nitrite/nitrate excretion, intra-renal production of NO, expression of iNOS protein and induction of iNOS mRNA. We conclude that MMF is effective in attenuating the severity of nephritis in MRL/lpr mice. The beneficial effects of MMF on lupus nephritis during the early phase of the disease might be partly attributed to the inhibition of NO production. The inhibitory effect of MMF on NO production diminishes as the disease progresses. MMF probably has additional, as yet undefined mode of actions to fully account for its beneficial effects on lupus nephritis.     

Persistence of the nitric oxide pathway in the aorta of hypercholesterolemic apolipoprotein-E-deficient mice

The markers of the bioavailability of NO (endothelium-dependent relaxation to acetylcholine and cyclic GMP content) in the thoracic aorta of apolipoprotein-E-deficient (ApoE KO) mice, 20 weeks old with enriched cholesterol diet or 35 weeks old on regular chow, are not decreased, in contrast with other models of atherosclerosis. However, severe hypercholesterolemia, the presence of atherosclerotic lesions and increased NADPH oxidase activity have been reported as early as at 20 weeks of age. The present experiments were designed to test if an increased capacity of NO production in these mice explains this paradox. The expressions of the 3 isoforms of NO synthase (NOS) were compared in ApoE KO and C57Bl/6J mice using Western blot and localized by immunohistochemistry. No adaptive increase in the expression of NOS was detected in ApoE KO mice. NO bioavailability could also be preserved by upregulation of enzymes involved in redox status such as CuZn or Mn superoxide dismutase and catalase. However, these enzymes were less expressed in ApoE KO mice than in control mice. These results highlight that ApoE KO mice represent an atypical model of atherosclerosis.     

Alterations in nitrate/nitrite and nitric oxide synthase in preovulatory follicles in gonadotropin-primed immature rat

Nitric oxide synthase (NOS) and nitric oxide (NO) play important roles in ovulation. The purpose of this study was to investigate the changes of intrafollicular nitrate/nitrite concentration and NOS mRNA expression in preovulatory follicles during equine CG (eCG) and human CG (hCG) induced ovulation in immature rats. Immature Sprague-Dawley rats received 15 IU eCG and then 15 IU hCG 48 h later. Rats were killed immediately before, 5 h after or 10 h after hCG injection, and their preovulatory follicles were dissected. Follicular fluid, granulosa cell, and theca cell layers were collected from preovulatory follicles and assayed for NO or NOS mRNA or for in vitro incubation study. Nitrate/nitrite concentration in the follicular fluid decreased significantly 5 and 10 h after hCG injection. Inducible NOS (iNOS) mRNA expression, which was greater in granulosa cell than in the theca cell layer, decreased significantly 5 and 10 h after hCG injection. However, endothelial NOS (eNOS) mRNA expression was detected mainly in the theca cell layer and further increased 5 and 10 h after hCG injection but remained low in granulosa cells. In vitro treatment of granulosa cells with 10(-4) or 5x10(-4) M S-nitroso-L-acetyl penicillamine (NO donor) decreased progesterone production and increased DNA fragmentation. We concluded that the decrease in nitrate/nitrite concentration in preovulatory follicles after hCG injection was due mainly to decreased iNOS expression in granulosa cells. These changes in nitrate/nitrite concentration may prevent apoptosis in preovulatory follicles.     

Enhanced mucosal permeability and nitric oxide synthase activity in jejunum of mast cell deficient mice

Background—Recent reports have described amodulating influence of nitric oxide (NO) on intestinal mucosalpermeability and have implicated a role for mast cells in this NOmediated process.
Aims—To assess further the contribution of mastcells to the mucosal permeability changes elicited by the NO synthase(NOS) inhibitor N^G-nitro-L-arginine methylester(L-NAME), using mast cell deficient (W/W^V) andmast cell replete mice (+/+).
Methods—Chromium-51 EDTA clearance (from blood tojejunal lumen), jejunal NOS and myeloperoxidase (MPO) activities, andplasma nitrate/nitrite levels were monitored.
Results—The increased EDTA clearance elicited byintraluminal L-NAME in W/W^V mice (4.4-fold) wassignificantly greater than the response observed in control (+/+) mice(1.8-fold). The exacerbated response in W/W^v mice wasgreatly attenuated by pretreatment with either dexamethasone (1.3-fold)or the selective inducible NOS inhibitor, aminoguanidine (1.4-fold),and partially attenuated by the mast cell stabiliser, lodoxamide(2.9-fold). Jejunal inducible NOS activity was significantly higher inW/W^V than in +/+ mice, while jejunal MPO was lower inW/W^V mice than in +/+ mice, suggesting that the higherinducible NOS in W/W^V does not result from the recruitmentof inflammatory cells into the gut. The higher inducible NOS activityin the jejunum of W/W^V was significantly reduced bydexamethasone treatment.
Conclusions—Our results suggest that mast cellsnormally serve to inhibit inducible NOS activity tonically in the gutand that inhibitors of NOS elicit a larger permeability response when this tonic inhibitory influence is released by mast cell depletion.

Keywords:aminoguanidine; c-kit; dexamethasone; epithelium; neutrophils