Feeding the nitric oxide synthase inhibitor L-N(omega)nitroarginine elevates serum very low density lipoprotein and hepatic triglyceride synthesis in rats

This study was conducted to study the influence of dietary L-N(omega)nitroarginine (L-NNA), a nitric oxide (NO) synthase inhibitor, on serum lipids and lipoproteins and on the activities of enzymes related to lipid metabolism in rats. Feeding rats a diet containing 0.2 g/kg L-NNA for 5 weeks elevated serum concentrations of triglyceride, cholesterol, phospholipid, and free fatty acid and reduced serum nitrate (an oxidation product of NO). The elevation in serum triglyceride was mainly due to the elevation in very low density lipoprotein (VLDL) triglyceride. Contents of cholesterol and phospholipid in the VLDL fraction also were elevated by L-NNA. L-NNA treatment caused significantly higher activity of hepatic microsomal phosphatidate phosphohydrolase (the rate-limiting enzyme in triglyceride synthesis) and lower activity of hepatic carnitine palmitoyltransferase (the rate-limiting enzyme in fatty acid oxidation). Activities of hepatic enzymes responsible for fatty acid synthesis such as glucose-6-phosphate dehydrogenase, malic enzyme, and fatty acid synthase were unaffected by L-NNA. The activity of hepatic microsomal phosphocholine cytidyltransferase (the rate-limiting enzyme in phosphatidylcholine synthesis) was reduced significantly by L-NNA. Our results suggest that lower NO production caused the elevations in hepatic triglyceride synthesis by higher esterification of fatty acid and lower fatty acid oxidation, leading to an enrichment of VLDL triglyceride.     

Abrogation of bleomycin-induced lung fibrosis by nitric oxide synthase inhibitor, aminoguanidine in mice.

The effects of aminoguanidine (AG), a specific inhibitor of inducible nitric oxide synthase, on the bleomycin (BL)-induced lung fibrosis was evaluated in mice. The animals were placed into five groups: saline (SA)-instilled drinking water (SA+H(2)O), saline-instilled drinking water containing 0.5%AG (SA+0.5%AG), BL-instilled drinking water (BL+H(2)O), BL-instilled drinking water containing 0.2%AG (BL+0.2%AG), and BL-instilled drinking water containing 0.5%AG (BL+0.5%AG). The mice had free access to H(2)O or H(2)O containing AG and lab chow ad lib 2 days prior to intratracheal (IT) instillation of BL (0.07U/mouse/100 microL) or an equivalent volume of sterile isotonic saline. The mice in the SA+0.5%AG group consumed the greatest amount of AG without any ill effects than the mice in any other group. There were no differences in any of the measured biochemical determinants between the SA+H(2)O and SA+0.5%AG control groups. The IT instillation of BL in the BL+H(2)O group caused significant increases in the lipid peroxidation, hydroxyproline content, and prolyl hydroxylase activity of lungs and influx of inflammatory cells in the broncheoalveolar lavage fluid (BALF) as compared to both control groups. The intake of aminoguanidine by mice in the BL+0.5%AG group caused significant reductions in the BL-induced increases in all measured biochemical indices of lung fibrosis without any effects on the influx of inflammatory cells in the BALF. In fact, AG in both BL-treated groups additionally increased the total cell counts in the BALF from mice in the BL+0.2%AG and BL+0.5%AG groups as compared to the BL+H(2)O group. Histopathological evaluation of the lungs revealed that the mice in the BL+0.5%AG group had markedly fewer fibrotic lesions than mice in the BL+H(2)O group. These results demonstrate that aminoguanidine minimizes the BL-induced lung fibrosis at both the biochemical and the morphological level and support our earlier hypothesis that the production of nitric oxide plays a significant role in the pathogenesis lung fibrosis caused by BL.     

Impaired duodenal bicarbonate secretion in diabetic rats. Salutary effect of nitric oxide synthase inhibitor

We previously reported the impaired HCO₃⁻ secretion and the increased mucosal susceptibility to acid in the duodenum of streptozotocin (STZ)-induced diabetic rats. In this study, we investigated the salutary effect of the NO synthase inhibitor L-NAME (N^G-nitro-L-arginine methyl ester) on these changes and compared it with those of insulin. Animals were injected streptozotocin (STZ: 70 mg/kg, ip) and used after 1, 3–4, and 5–6 weeks of diabetes with blood glucose levels of > 300 mg/dL. Under urethane anesthesia the HCO₃⁻ secretion was measured in the proximal duodenal loop using a pH-stat method and by adding 10 mM HCl. L-NAME (20 mg/kg × 2) or insulin (4 units/rat) was administered sc for 4–5 weeks, starting 1 week after STZ treatment. The duodenal HCO₃⁻ secretory responses to various stimuli such as mucosal acidification (10 mM HCl for 10 min), 16,16-dimethyl prostaglandin E₂ (dmPGE₂: 10 μg/kg, iv), and vagal stimulation (0.5 mA, 2 ms, 3 Hz) were significantly decreased in STZ-treated rats, depending on the duration of diabetes. Repeated administration of L-NAME, starting from 1 week after STZ treatment, significantly reduced blood glucose levels toward normal values and restored the HCO₃⁻ responses to various stimuli in STZ rats, the effects being similar to those observed after supplementation of insulin. Diabetic rats developed duodenal lesions after perfusion of the duodenum with 150 mM HCl for 4 h, but this ulcerogenic response was significantly inhibited by the repeated treatment with L-NAME as well as insulin. We conclude that L-NAME is effective in ameliorating hyperglycemic conditions in STZ-diabetic rats, similar to insulin, and restores the impaired HCO₃⁻ secretion and the increased mucosal susceptibility to acid in diabetic rat duodenums.     

Hyperactivity caused by a nitric oxide synthase inhibitor is countered by ultra-wideband pulses.

Potential action of ultra-wideband (UWB) electromagnetic field pulses on effects of N(G)-nitro- L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase (NOS), on nociception and locomotor activity was investigated in CF-1 mice. Animals were injected IP with saline or 50 mg/kg L-NAME and exposed for 30 min to no pulses (sham exposure) or UWB pulses with electric field parameters of 102+/-1 kV/m peak amplitude, 0.90+/-0.05 ns duration, and 160+/-5 ps rise time (mean+/-S.D.) at 600/s. Animals were tested for thermal nociceptive responses on a 50 degrees C surface and for spontaneous locomotor activity for 5 min. L-NAME by itself increased mean first-response (paw lift, shake, or lick; jump) and back-paw-lick response latencies and mean locomotor activity. Exposure to UWB pulses reduced the L-NAME-induced increase in back-paw-lick latency by 22%, but this change was not statistically significant. The L-NAME-induced hyperactivity was not present after UWB exposure. Reduction and cancellation of effects of L-NAME suggest activation of opposing mechanism(s) by the UWB pulses, possibly including increase of nitric oxide production by NOS. The action, or actions, of UWB pulses appears to be more effective on locomotor activity than on thermal nociception in CF-1 mice.     

The role of inducible nitric oxide synthase inhibitor on the arteriolar hyporesponsiveness in hemorrhagic-shocked rats

Hemorrhagic shock (HS) has been implicated in the induction of inducible nitric oxide synthase (iNOS) that leads to increase production of nitric oxide (NO). Recently, NO has been implicated to cause hyporesponsiveness of blood vessel in vitro towards vasoconstrictors in refractory (decompensated) HS. In our in vivo model, we examined the effects of aminoguanidine (AG), a known iNOS inhibitor, with angiotensin II (ANG II), a vasoconstrictor, following hemorrhagic shock decompensatory phase (HSDP) on percentage survival, vascular responsiveness, mean arterial blood pressure (MABP), heart rate and mean nitrate/nitrite levels in anaesthetized rats. HSDP (3 h) was achieved via constant pressure method (40-45 mmHg). MABP and heart rate was measured via the left carotid artery. Plasma collected from HSDP rats was used to measure nitrate/nitrite levels. Vascular hyporeactivity to ANG II was carried out using HSDP aortic strips, precontracted with KCl and noradrenaline. Sham-operated rats served as controls. HSDP rats decreased percentage survival, vascular contractility to ANG II and noradrenaline, MABP, heart rate while showing increased levels of nitrate/nitrite. Infusion of AG with ANG II, increased percentage survival and had reversed these cardiovascular effects of HSDP rats. This study indicates that excessive NO formation from iNOS activity induces vascular hyporeactivity and decompensation in HSDP. This might suggest that selective NOS inhibitor, AG, when coupled with ANG II, show reduction in NO's effect in HSDP.     

N-acetylcysteine inhibits in vivo nitric oxide production by inducible nitric oxide synthase.

This in vivo study evaluates the effect of N-acetylcysteine (NAC) administration on nitric oxide (NO) production by the inducible form of nitric oxide synthase (iNOS). NO production was induced in the rat by the ip administration of 2 mg/100 g lipopolysaccharide (LPS). This treatment caused: (1) a decrease in body temperature within 90 min, followed by a slow return to normal levels; (2) an increase in plasma levels of urea, nitrite/nitrate, and citrulline; (3) the appearance in blood of nitrosyl-hemoglobin (NO-Hb) and in liver of dinitrosyl-iron-dithiolate complexes (DNIC); and (4) increased expression of iNOS mRNA in peripheral blood mononuclear cells (PBMC). Rat treatment with 15 mg/100 g NAC ip, 30 min before LPS, resulted in a significant decrease in blood NO-Hb levels, plasma nitrite/nitrate and citrulline concentrations, and liver DNIC complexes. PBMC also showed a decreased expression of iNOS mRNA. NAC pretreatment did not modify the increased levels of plasma urea or the hypothermic effect induced by the endotoxin. The administration of NAC following LPS intoxication (15 min prior to sacrifice) did not affect NO-Hb levels. These results demonstrate that NAC administration can modulate the massive NO production induced by LPS. This can be attributed mostly to the inhibitory effect of NAC on one of the events leading to iNOS protein expression. This hypothesis is also supported by the lack of effect of late NAC administration.     

一氧化氮合酶抑制剂L-NAME对大鼠脑缺血耐受诱导的影响

采用大鼠四血管闭塞全脑缺血耐受模型和脑组织切片形态学方法,观察应用一氧化氮合酶(NOS)抑制剂L—NAME对大鼠海马CAl区脑缺血耐受(BIT)诱导的影响,在整体水平探讨一氧化氮(NO)在BIT诱导中的作用。54只Wistar大鼠凝闭双侧推动脉后分为6组：(1)假手术组(n＝6);分离双侧颈总动脉,但不阻断脑血流;(2)损伤性缺血组(n＝6)：全脑缺血10min;(3)预缺血 损伤性缺血组(n＝6)：脑缺血预处理(CIP)3min,再灌注72h后行全脑缺血10min;(4)L—NAME组;分别于CIP前1h和后1、12及36h腹腔注射L—NAME(5mg／kg),每个时间点6只动物,其余步骤同预缺血 损伤性缺血组;(5)L—NAME L—精氨酸组(n＝6)：于CIP前1h腹腔注射L—NAME(5mg／kg)和L—精氨酸(300mg／kg),其它步骤同L—NAME组;(6)L—NAME 损伤性缺血组(n＝6)：于腹腔注射L—NAME(5mg／kg)72h后行全脑缺血10min。实验结果表明,(1)单纯10min全脑缺血可使海马CAl区组织学分级增加(表明损伤加重),神经元密度降低(P＜0．01);(2)预缺血 损伤性缺血组的海马CAl区组织学分级、神经元密度与假手术组相比,无显著性差别(P＞0．05);(3)L—NAME组中,应用L—NAME后海马CAl区组织学分级增加,神经元密度降低,与预缺血 损伤性缺血组相比有显著性差异(P＜0．05),表明L—NAME可阻断CIP对神经元的保护作用;(4)L—NAME L—精氨酸组与L—NAME组相比,海马CAl区组织损伤明显减轻(P＜0．05),但与预缺血 损伤性缺血组相比仍有显著性差别(P＜0．05),提示L-精氨酸可部分逆转L—NAME的作用;(5)L—NAME 损伤性缺血组的组织学表现与损伤性缺血组相同(P＞0．05)。这些结果表明,在整体情况下N0参与BIT的诱导。与CIP前1h及后1、12h给予L—NAME组相比,CIP后36h给予L—NAME对CIP保护作用的阻断效应明显减弱,提示N0在CIP后较早阶段即开始参与BIT的诱导。     

Inhibition of the morphine withdrawal syndrome by a nitric oxide synthase inhibitor,NG-nitro-L-arginine methyl ester

The hypothesis that an arginine-nitric oxide (NO) synthase-NO system mediates the morphine abstinence syndrome was tested in adult male rats implanted subcutaneosly for 3 days with one morphine (75 mg) pellet followed by naloxone-precipitated withdrawal (0.5 mg/kg). Injection with a NO synthase inhibitor, N^G-nitro-L-arginine methyl ester (NAME, 100 mg/kg subcutaneous), shortly before naloxone-induced withdrawal significantly inhibited abstinence signs by 25–80%. Continuous infusion of NAME via subcutaneous osmotic pumps during the development of morphine physical dependence and during naloxone-precipitated withdrawal also inhibited morphine abstinence signs. In addition, treatment with isosorbide dinitrate, a NO donor, induced a quasi morphine-abstinence syndrome (QMAS) that was significantly suppressed by implantation of a morphine pellet 3 days before isosorbide dinitrate treatment. These results indicate that NO mediates part of the expression of the morphine abstinence syndrome.     

Nitric oxide produced by inducible nitric oxide synthase is associated with mammary tumorigenesis in irradiated rats.

This study evaluated whether nitric oxide (NO) derived from nitric oxide synthase (NOS) induced by radiation is associated with tumorigenesis in the mammary glands. When rats were exposed to whole-body irradiation with gamma-rays (1.5 Gy) immediately after weaning and then treated with diethylstilbestrol, as an irradiated control, the tumor incidence (85%) was increased 7.6-fold in comparison with that (11.1%) of the non-irradiated control. The tumor incidence declined to 28.6% in the rats injected intraperitoneally with phenyl-N-tert-butylnitrone (PBN, 160 mg/kg), an inhibitor of inducible NOS (iNOS) expression and also a spin trapping agent, 30 min before irradiation. Also, the tumor incidence (25%) in rats orally administered with N-(3-(aminomethyl)-benzyl)-acetamide (1400W, 2.3+/-0.1 mg/day), a highly selective inhibitor of iNOS, dissolved in drinking water for 3 days after the irradiation was less than one-third of that in the irradiated control. On treatment with PBN or 1400W, no adenocarcinoma developed. Many of the mammary tumors that developed in the irradiated rats were positive for the estrogen receptor (ER). In contrast, ER was not detected in the tumors yielded from irradiated rats administered with PBN or 1400W. These results indicate that iNOS-derived NO may participate in the formation of estrogen-dependent mammary adenocarcinomas following radiation.     

Role of neuronal nitric oxide synthase in hypoxia-induced anapyrexia in rats.

Anapyrexia (a regulated decrease in body temperature) is a response to hypoxia that occurs in organisms ranging from protozoans to mammals, but very little is known about the mechanisms involved. Recently, it has been shown that the NO pathway plays a major role in hypoxia-induced anapyrexia. However, very little is known about which of the three different nitric oxide synthase isoforms (neuronal, endothelial, or inducible) is involved. The present study was designed to test the hypothesis that neuronal nitric oxide synthase (nNOS) plays a role in hypoxia-induced anapyrexia. Body core temperature (T(c)) of awake, unrestrained rats was measured continuously using biotelemetry. Rats were submitted to hypoxia, 7-nitroindazole (7-NI; a selective nNOS inhibitor) injection, or both treatments together. Control animals received vehicle injections of the same volume. We observed a significant (P < 0.05) reduction in T(c) of approximately 2.8 degrees C after hypoxia (7% inspired O(2)), whereas intraperitoneal injection of 7-NI at 25 mg/kg caused no significant change in T(c). 7-NI at 30 mg/kg elicited a reduction in T(c) and was abandoned in further experiments. When the two treatments were combined (25 mg/kg of 7-NI and 7% inspired O(2)), we observed a significant attenuation of hypoxia-induced anapyrexia. The data indicate that nNOS plays a role in hypoxia-induced anapyrexia.     

Identification of inducible calmodulin-dependent nitric oxide synthase in the liver of rats.

A calmodulin-dependent nitric oxide synthase was significantly induced in the liver of rats treated intravenously with heat-killed Propionibacterium acnes and 5 days later with Escherichia coli lipopolysaccharide. The apparent calmodulin-dependent and -independent isozymes were separated by Mono Q column chromatography after their partial purification by 2',5'-ADP-agarose affinity chromatography. Both enzymes had a molecular weight of 125,000 as determined by SDS-polyacrylamide gel electrophoresis and required NADPH, tetrahydrobiopterin, and dithiothreitol as cofactors. Their activities were completely inhibited by the specific nitric oxide synthase inhibitors NG-monomethyl-L-arginine and N omega-nitro-L-arginine at 80 and 800 microM, respectively. The peptide maps of these two isozymes with lysylendopeptidase and their reverse-phase column chromatographic profiles were indistinguishable. In the presence of bovine calmodulin, the purified calmodulin-dependent isozyme behaved as a calmodulin-independent isozyme on Mono Q column chromatography. The purified calmodulin-independent isozyme was converted to a calmodulin-dependent isozyme by EDTA and EGTA. Calmodulin blot analysis using 125I-calmodulin showed that the two isozymes bound calmodulin equally efficiently.     

Cadmium reduces nitric oxide production by impairing phosphorylation of endothelial nitric oxide synthase.

Cadmium (Cd) perturbs vascular health and interferes with endothelial function. However, the effects of exposing endothelial cells to low doses of Cd on the production of nitric oxide (NO) are largely unknown. The objective of the present study was to evaluate these effects by using low levels of CdCl2 concentrations, ranging from 10 to 1000 nmol/L. Cd perturbations in endothelial function were studied by employing wound-healing and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. The results suggest that a CdCl2 concentration of 100 nmol/L maximally attenuated NO production, cellular migration, and energy metabolism in endothelial cells. An egg yolk angiogenesis model was employed to study the effect of Cd exposure on angiogenesis. The results demonstrate that NO supplementation restored Cd-attenuated angiogenesis. Immunofluorescence, Western blot, and immuno-detection studies showed that low levels of Cd inhibit NO production in endothelial cells by blocking eNOS phosphorylation, which is possibly linked to processes involving endothelial function and dysfunction, including angiogenesis.     

Expression of endothelial nitric oxide synthase in ciliated epithelia of rats.

Endothelial nitric oxide synthase (eNOS), originally found in the endothelium of vascular tissue, also exists in other cell types, including ciliated epithelia of airways. The eNOS is ultrastructurally localized to the basal body of the microtubules of the cilia, and nitric oxide (NO) stimulates ciliary beat frequency (CBF). We examined whether the expression of eNOS is present in ciliated cells of other organs. Western blotting analysis revealed that eNOS was expressed in the rat cerebrum, lung, trachea, testis, and oviduct. Immunohistochemical staining showed that eNOS was localized in the ciliated epithelia of airways, oviduct, testis, and ependymal cells of brain in addition to the endothelium and smooth muscle of the vasculature. To confirm the activation of eNOS in the ciliated epithelia, we examined the effect of L-arginine (L-Arg), the substrate of NOS, on the production of nitrite and nitrate (NOx) in the cultured explants of rat trachea. L-Arg (100 microM) increased NOx levels significantly (p<0.05). In explants exposed to inhibitors of NOS, the effect of l-Arg on the production of NOx was blocked. These findings suggest that epithelial NO plays an important role in signal transduction associated with ciliary functions.     

Systemic injection of a nitric oxide synthase inhibitor suppresses sleep responses to sleep deprivation in rats

We hypothesized that nitric oxide (NO) may play a role in homeostatic sleep regulation. To test this hypothesis, we studied the sleep deprivation (SD)-induced homeostatic sleep responses after intraperitoneal administration of an NO synthase inhibitor, Nomega-nitro-L-arginine methyl ester (L-NAME, a cumulative dose of 100 mg/kg). Amounts and intensity of sleep were increased in response to 8 h of SD in control rats (n = 8). Sleep amounts remained above baseline for 16 h after SD followed by a negative rebound. Rapid eye movement sleep (REMS) and non-REMS (NREMS) intensities were elevated for 16 and 4 h, respectively. L-NAME treatment (n = 8) suppressed the rebound increases in NREMS amount and intensity. REMS rebound was attenuated by L-NAME in the first dark period after SD; however, a second rebound appeared in the subsequent dark period. REMS intensity did not increase after SD in L-NAME-injected rats. The finding that the NO synthase inhibitor suppressed rebound increases in NREMS suggests that NO may play a role as a signaling molecule in homeostatic regulation of NREMS.     

Epigallocatechin gallate preserves endothelial function by reducing the endogenous nitric oxide synthase inhibitor level

Asymmetric dimethylarginine (ADMA), the endogenous nitric oxide synthase inhibitor, is thought to be a key factor contributing to endothelial dysfunction. Tea catechins can cause an endothelium-dependent vasorelaxation. The present study examined the effect of epigallocatechin gallate (EGCG), the major component of tea catechins, on endothelial dysfunction induced by native low density lipoprotein (LDL) in rats and oxidized LDL (ox-LDL) in cultured endothelial cells, and whether the protective effect of EGCG is related to reduction of ADMA level. A single injection of LDL (4 mg x kg(-1), i.v.) markedly reduced endothelium-dependent relaxation and the serum nitrite/nitrate (NO) level, and increased serum concentrations of ADMA, malondialdehyde (MDA), and tumor necrosis factor-alpha (TNF-alpha). EGCG (10 or 50 mg x kg(-1), i.p.) significantly attenuated the inhibition of vasodilator response to acetylcholine and the decreased serum nitrite/nitrate level, and reduced the elevated levels of ADMA, MDA, and TNF-alpha. Exposure of endothelial cells to ox-LDL (100 microg x mL(-1)) for 24 h markedly increased the medium levels of lactate dehydrogenase (LDH), ADMA, TNF-alpha, and MDA, and decreased the level of nitrite/nitrate in the medium and the activity of dimethylarginine dimethylaminohydrolase (DDAH) in the endothelial cells. EGCG (10 and 100 microg x mL(-1)) significantly decreased the levels of LDH, ADMA, TNF-alpha, and MDA, and increased the level of nitrite/nitrate and the activity of DDAH. These results suggest that EGCG protects endothelial dysfunction induced by native LDL in vivo or by ox-LDL in endothelial cells, and the protective effect of EGCG on the endothelium is related to decrease in ADMA level via increasing of DDAH activity.     

Preferential loss of body fat during starvation in dietary obese rats.

This study was undertaken to examine whether diet-induced obesity alters the amount and/or composition of weight lost during starvation. The amount and composition of weight lost during a 4-day period of starvation was determined before and at 17, 30 and 42 weeks after rats (350 g of body weight) were given a high fat diet (HFD). To control for effects of aging, a second group of rats, fed standard laboratory chow, was also subjected to similar periods of starvation. Although total weight loss during starvation was never greater for HFD rats than for chow-fed rats, the former group showed a clear patter of increasing loss of body fat and total energy and conservation of fat-free tissues with periods of starvation later in life. In addition, chow-fed rats showed substantial energy conservation during each period of starvation (i.e. they lost less energy each day than their pre-starvation energy requirements). In contrast, HFD rats demonstrated substantial energy conservation only at 17 weeks and not at 30 or 42 weeks; during the last period of starvation, their average daily loss of carcass energy exceeded their pre-starvation energy requirements. This suggests the increased fat mass of these rats may have led to increased fuel availability and to an increased metabolic rate during starvation. If these results are applicable to humans, the more obese subjects are likely to show greater total loss of energy than lean subjects, but show a lesser loss of lean body mass, at least initially. If protein requirements are reflected by the ability to mobilize protein during food restriction, protein requirements would be substantially lower in the dietary obese rats than in controls. In summary, diet-induced obesity leads to preferential loss of body fat and conservation of lean mass during starvation.     

Role of endogenous nitric oxide synthase inhibitor in gastric mucosal injury

To explore the role of the endogenous nitric oxide synthase (NOS) inhibitor asymmetric dimethylarginine (ADMA) in gastric mucosal injury, 3 models of gastric mucosal injury induced by ethanol, indomethacin, or cold stress were used in rats. The cultured human gastric mucosal epithelial cell line GES-1 infected by Helicobacter pylori (Hp) was selected to mimic human gastric mucosal injury. Gastric mucosal ulcer index (UI), levels of ADMA and NO, and activity of dimethylarginine dimethylaminohydrolase (DDAH) were determined in the mucosal injury models; in Hp-infected or ADMA-treated GES-1 cells, levels of ADMA, NO, and TNF-alpha and activity of DDAH were measured. The results showed that UI and levels of ADMA were markedly increased and accompanied by significantly decreased DDAH activity in the mucosal injury models. Incubation of GES-1 cells with Hp increased levels of TNF-alpha and ADMA and decreased activity of DDAH. Administration of ADMA also increased levels of TNF-alpha. The results suggest that ADMA plays an important role in facilitating gastric mucosal injury, an effect which is associated with inhibiting NO synthesis and inducing inflammatory reaction.     

Day- and nighttime injection of a nitric oxide synthase inhibitor elicits opposite sleep responses in rats

Previous studies suggest that nitric oxide (NO) may play a role in sleep regulation, particularly in the homeostatic process. The present studies were undertaken to compare the sleep effects of injecting a NO synthase (NOS) inhibitor when homeostatic sleep pressure is naturally highest (light onset) or when it is at its nadir (dark onset) in rats. Sleep, electroencephalogram delta-wave activity during nonrapid eye movement sleep (NREMS), also known as slow-wave activity (SWA), and brain temperature responses to three doses of the NOS inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME ; 5, 50, and 100 mg/kg) injected intraperitoneally at light or dark onset were examined in rats (n = 6 to 8). The effects of 5 mg/kg L-NAME were determined in both normal and vagotomized (VX) rats. Light onset administration of 50 mg/kg L-NAME decreased NREMS amounts and suppressed SWA and increased rapid eye movement sleep (REMS) amounts. At dark onset, L-NAME injection also dose dependently suppressed SWA; however, unlike light onset injections, both NREMS and REMS amounts were increased after all three doses. Sleep responses to 5 mg/kg L-NAME were not different in control and VX rats, suggesting that the sleep effects of L-NAME are not mediated through the activation of sensory vagal mechanisms. The present findings suggest that timing of the injection is a major determinant of the sleep responses observed after systemic L-NAME injection in rats.     

Phosphatidylinositol 3-kinase inhibitor suppresses inducible nitric oxide synthase expression in bronchiole epithelial cells in asthmatic rats

Inducible nitric oxide synthase (iNOS) is known to produce nitric oxide (NO), which is a main contributor to asthmatic airway inflammation. Recent studies have shown that phosphatidylinositol 3-kinase (PI3K) is ubiquitously expressed in airway epithelial cells and its inhibition could relieve airway inflammation and hyperresponsiveness. This study aimed to explore the interaction of PI3K and NO signaling in allergic asthma. We investigated the effects of PI3K inhibitor wortmannin on iNOS expression in bronchiole epithelial cells and NO, IL-4 and IFN-γ levels in lung tissues of asthmatic rat model, which was prepared by 10% OVA solution sensitization and 1% OVA aerosol challenge. Our results showed that the ratio of eosinophils to total cells in BALF, PI3K activity, NO and IL-4 levels in lung tissues was increased after OVA sensitization and challenge, but then was attenuated by the administration of wortmannin. In contrast, IFN-γ level in lung tissues was decreased after OVA sensitization and challenge and increased after the administration of wortmannin. The expression of iNOS protein in bronchiole epithelial cells, iNOS mRNA level and iNOS activity in lung tissues was markedly upregulated after OVA sensitization and challenge, but the upregulation was significantly antagonized by wortmannin. Taken together, these data provide evidence that PI3K functions upstream to modulate iNOS/NO signaling, which then promotes the development of airway inflammation in asthmatic animal model. PI3K inhibitor wortmannin could lead to reduced iNOS expression and NO production, therefore inhibiting airway inflammatory responses.