Role of neuronal nitric oxide synthase and inducible nitric oxide synthase in intestinal injury in neonatal rats

AIM: To investigate the dynamic change and role of neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS) in neonatal rat with intestinal injury and to define whether necrotizing enterocolitis (NEC) is associated with the levels of nitric oxide synthase (NOS) in the mucosa of the affected intestine tissue. METHODS: Wistar rats less than 24 h in age received an intraperitoneal injection with 5 mg/kg lipopolysaccharide (IPS). Ileum tissues were collected at 1, 3, 6, 12 and 24 h following LPS challenge for histological evaluation of NEC and for measurements of nNOS and iNOS. The correlation between the degree of intestinal injury and levels of NOS was determined. RESULTS: The LPS-injected pups showed a significant increase in injury scores versus the control. The expression of nNOS protein and mRNA was diminished after LPS injection. There was a negative significant correlation between the nNOS protein and the grade of median intestinal injury within 24 h. The expression of iNOS protein and mRNA was significantly increased in the peak of intestinal injury. CONCLUSION: nNOS and iNOS play different roles in LPS-induced intestinal injury. Caution should be exerted concerning potential therapeutic uses of NOS inhibitors in NEC.     

人胃癌组织中一氧化氮合酶的表达

目的探讨NOS与胃癌的关系.方法用NADPH-d组织化学法测定了正常胃组织、癌旁组织和癌组织中一氧化氮合酶(NOS)表达水平.结果正常胃组织中粘膜上皮细胞、各种有分泌功能的细胞及肌层神经纤维中均有NOS表达,测一个视野NOS阳性细胞的平均灰度,正常胃组织为112、癌旁组织为120、胃癌组织为145.各组间差异有显著意义.表明正常胃组织NOS活性最高,胃癌组织NOS活性最低.结论①正常胃组织有广泛的NOS分布,提示NO对维持正常胃功能具有重要作用;②胃粘膜细胞癌变过程中,NOS活性明显降低,提示NOS活性与胃粘膜细胞癌变有高度相关性.     

Nitric oxide production in rheumatoid arthritis

Abstract

Nitric oxide (NO), originally found as endothelium-derived relaxing factor (EDRF), is a free radical synthesized by NO synthases (NOS). Two isoforms exist in NOS, i.e. constitutive NOS (cNOS) and inducible NOS (iNOS). Inflammatory cytokines such as interleukin-1, interferon-γ, tumor necrosis factor-α induce iNOS expression in various cells including macrophages. Enhanced NO production is observed in arthritic conditions both in rodent models and human. The onset of arthritis in rodent models is significantly inhibited by the NOS inhibitor, N ^G-monomethyl-l-arginine. These data suggest a possible involvement of NO in the induction and/or maintenance of rheumatoid arthritis.     

梅毒患者血清一氧化氮和一氧化氮合酶水平测定

目的　检测梅毒螺旋体感染者血清中一氧化氮 (NO)和一氧化氮合酶 (NOS)的水平。方法　用分光光度法测定血清中NO水平和NOS活性 ,血清中NO3 和NO2 总量代表体内NO水平 ,NOS催化L 精氨酸和氧的反应生成NO的多少代表血清NOS活性。结果　梅毒患者NO浓度为 115± 36 3nmol/L ,NOS活性为 35 8± 7 3U/ml,二者均远远高于正常对照组。结论　梅毒螺旋体的感染引起患者体内NO水平和NOS活性升高 ,NO在梅毒感染中可能发挥重要的作用。     

Expression of inducible nitric oxide synthase in human gastric cancer

INTRODUCTIONInduciblenitricoxidesynthase(iNOS)isanenzymethatcatalyzestheformationofnitric0xide(N0)fromL-arginine.iNOSexpressionandactivityresultsintheproduction0fhighlevelsofNO[1].ThegenerationofphysiologicallevelsofNOisimp0rtantformucosalfunctionanditalsoexertsacytoprotectiveeffectonthegastr0intestinalmucosa.However,increasediNOSexpressionhasbeenobservedinpatientswithchronicinflammatorydiseasesofthegastr0intestinaltract,suchasulcerativec0litis[2'3],andgastritis['Jandithasbeenspecul…     

Subcellular distribution of nitric oxide synthase isoforms in the rat duodenum

AIM:To study the cell-type specific subcellular distribution of the three isoforms of nitric oxide synthase(NOS) in the rat duodenum.METHODS:Postembedding immunoelectronmicroscopy was performed,in which primary antibodies for neuronal NOS(nNOS),endothelial NOS(eNOS),and inducible NOS(iNOS),were visualized with protein A-gold-conjugated secondary antibodies.Stained ultrathin sections were examined and photographed with a Philips CM10 electron microscope equipped with a MEGAVIEW II camera.The specificity of t...     

Nitric oxide synthase activity in rat cardiac mitochondria

Abstract. Recent publications shown mitochondrial localization of the enzyme nitric oxide synthase (NOS) in a number of tissues. However, conflicting results about mitochondrial NOS (mtNOS) immunoreactivity and enzymatic activity are available to date in the literature. In this study we purified mitochondria from rat hearts and analysed these preparations for NOS immunoreactivity and activity, showing the presence of either a constitutive (the endothelial isoform) and an inducible NOS immunoreactivity. A basal NOS activity (64.2 ± 5.1 pmol/mg protein/30 min) was detectable. 1 mM N^G-Monomethyl-L-arginine (L-NMMA), a competitive inhibitor of all NOS isoforms, caused a drop in NOS activity to 33.8 ± 1.9 pmol/mg protein/30 min. Simultaneous administration of 10 µM (S)-2-amino-(1-iminoethylamino)-5- thiopentanoic acid (GW274150), a specific NOS2 inhibitor, together with removal of Ca²⁺ and calmodulin (CaM) from the assay buffers, known to interfere with the activity of constitutive NOS isoforms, caused a reduction in NOS activity (17.4 ± 1.2 pmol/mg protein/30 min). 10 µM GW274150 reduced NOS activity to 41.6 ± 4 pmol/mg protein/30 min, while Ca²⁺/CaM withdrawal reduced basal NOS activity to 45.8 ± 5 pmol/mg protein/30 min. This dual mtNOS machinery is suggested to be involved in modulating cardiac O₂ consumption in different (patho)physiological conditions.     

NO及NOS在低氧大鼠肺组织和主动脉中的差异性及其意义

目的观察低氧性肺动脉高压大鼠肺组织匀浆、主动脉匀浆及出肺血和入肺血中一氧化氮（NO）的含量、一氧化氮合酶（NOS）的活性。方法将24只雄性SD大鼠随机分成4组,即常氧2周组、常氧3周组、低氧2周组、低氧3周组。采用间断负压低氧法制备大鼠低氧性肺动脉高压模型;右心室导管法测定最高肺动脉压（PAP）;左颈总动脉插管测量左颈总动脉压代表动脉血压（Psa）;计算右心室肥厚指数[RV/（LV+S）];采用硝酸还原酶法测定各组大鼠出、入肺血及肺组织和主动脉匀浆中的NO含量;用化学比色法测定各组大鼠肺组织和主动脉匀浆中NOS的活性;应用免疫组化染色法观察各组大鼠肺组织及主动脉eNOS在蛋白质水平表达的变化。结果低氧组大鼠的PAP[（43.4±4.4）mmHg,（51.8±4.2）mmHg,1mmHg=0.133kPa],RV/（LV+S）（32.3±1.0,37.0±1.6）均高于其正常对照组[（20.8±2.4）mmHg,（21.8±3.9）mmHg;21.3±1.0,20.3±1.2,P<0.01)],且随缺氧时间延长而增高（P<0.01）,而Psa与常氧对照组比无差别。低氧组大鼠的出、入肺血及肺组织匀浆中的NO含量、NOS活性及肺组织eNOS的表达量均较其常氧对照组显著降低（P<0.01）,出肺血与入肺血的NO含量无差别;主动脉匀浆中NO含量和NOS的活性及大鼠主动脉的eNOS染色在各组间未见明显差异。结论低氧时,大鼠肺组织中NO的含量及NOS的活性均较常氧时降低,而主动脉中二者的表达在低氧和常氧时却没有差异,这种差异性可能是低氧时引起肺动脉高压却很少导致高血压的机制之一。     

Blood pressure response to hypoxia: role of nitric oxide synthase

BACKGROUND: Chronic exposure to hypobaric hypoxia has been shown to increase arterial pressure in genetically normal rats. The associated increase in blood pressure is unrelated to the hypoxia-induced erythrocytosis and persists indefinitely after restoration of normoxia. It is accompanied by a marked reduction in urinary excretion of nitric oxide metabolites (NOx) and is ameliorated by L-arginine supplementation. In view of the latter observations, we hypothesized that hypoxia-induced hypertension may be associated with downregulation of NO synthase (NOS). METHODS: Male Sprague Dawley rats were randomized to the hypoxic and control groups. Rats assigned to the hypoxic group were placed in chambers with air pressure maintained at 390 mm Hg. Animals assigned to the control group were kept in the chamber at 760 mm Hg air pressure. Animals were kept in their respective conditions for up to 21 days. Group of animals were tested at days 2, 3, 7, and 21. RESULTS: The hypoxic group exhibited a steady increase in arterial pressure beginning at day 3. This was accompanied by a transient increase followed by a significant decline in kidney NOS-I, NOS-II, and NOS-III abundance. A similar biphasic change was observed with NOS-II and NOS-III in the cardiac and vascular tissues. The changes in NOS abundance in the given tissues were associated with parallel changes in nitrotyrosine abundance, which reflects local NO production. The latter finding provides functional evidence for the changes observed in NOS abundance. CONCLUSIONS: Chronic hypoxia-induced hypertension in rats is associated with marked downregulation of NOS isotypes, which can, in part, account for the previously reported L-arginine-responsive hypertension in this model.     

高同型半胱氨酸血症诱导eNOS脱偶联损害冠状动脉内皮功能

目的探讨高同型半胱氨酸血症（HHcy）患者冠状动脉内皮功能是否被损伤,以及这种损伤是否通过内皮型一氧化氮合酶（eNOS）脱偶联实现的。方法 71例参与者被分成健康对照组（n=50）和HHcy组（n=21）,利用多普勒超声心动测定腺苷诱导下冠状动脉左前降支的舒张功能的改变,冠脉血流速度储备（CFVR）由最大血流速度与基线水平的比值计算得出。采用ELISA以及高效液相色谱法测定血浆一氧化氮（NO）、四氢生物蝶呤（BH4）的水平。结果与健康对照组相比,HHcy组患者血浆NO、BH4的水平降低（P〈0.05）;HHcy组CFVR低于健康对照组（P〈0.05）;血浆Hcy水平与NO及CFVR呈负相关（P〈0.05）。结论 HHcy可能通过降低BH4生物利用度,诱导eNOS脱偶联,而导致冠状动脉内皮功能损伤,进而促进不良冠脉事件的发生。     

肝癌组织中一氧化氮合酶及其基因表达的研究

目的研究肝癌组织中一氧化氮合酶(iNOS)及其基因表达与肝癌发生发展的关系。方法用免疫组化和原位杂交的方法对21例肝癌及癌旁组织中的诱导型一氯化氮合酶(iNOS)及其基因表达进行原位检测和观察。结果:NOS 阳性反应物质呈黄色或棕黄色,位于细胞浆中。非癌殖织(肉眼观距癌组织边缘>1.5)多呈阴性或弥漫弱阳性,但部分非癌组织中可见 iNOS 呈阳性的细胞呈点状分布;癌旁组织多呈阳性,提示 iNOS 表达与肝组织癌变有关。癌组织核心多呈阴性或弥漫弱阳性,但分化中和差的癌组织核心也分别有一例 NOS 呈强阳性;周边癌组织呈局灶阳性,侵入纤维组织中的弥敢癌细胞星强阳性,提示 NOS 的表达与肝癌组织的侵润能力有关。肝癌组织 iNOSmRNA 阳性细胞的分布与 iN-OS 蛋白的表达基本相似。结论 iNOS 蛋白及其基因表达与肝组织癌变及肝癌侵润能力有关。     

复方灯盏花滴丸对海马神经元一氧化氮和一氧化氮合酶的影响

目的研究复方灯盏花滴丸(FDD)对谷氨酸致大鼠原代海马神经元的NO和一氧化氮合酶的影响。方法取SD大鼠32只,每8只分别用生理盐水、尼莫地平、FDD高、低剂量灌胃,85h后,取每只大鼠股动脉血,制成5%的含药血清。另取新生24h SD乳鼠大脑原代培养海马神经元并鉴定后,随机分为对照组、尼莫地平组、FDD高、低剂量组和模型组,前4组分别加入上述配制对应的药物血清,模型组不加含药血清。以谷氨酸制成损伤模型。采用MTT法测定细胞存活率,酶法检测细胞释放NO量,细胞内总一氧化氮合酶(tNOS)和诱导型一氧化氮合酶(iNOS)活性。结果与对照组比较,模型组细胞存活率下降,NO、tNOS和iNOS活性增加(P0.01),与模型组比较,FDD高、低剂量组均能明显增加细胞存活率、减少NO释放、降低tNOS活性和iNOS活性(P0.05,P0.01)。结论 FDD对谷氨酸致原代培养大鼠海马神经元损伤有保护作用,其作用机制可能与降低一氧化氮合酶的活性、尤其iNOS活性及减少NO释放有关。     

Cytochrome c oxidase regulates endogenous nitric oxide availability in respiring cells: a possible explanation for hypoxic vasodilation

One of the many routes proposed for the cellular inactivation of endogenous nitric oxide (NO) is by the cytochrome c oxidase of the mitochondrial respiratory chain. We have studied this possibility in human embryonic kidney cells engineered to generate controlled amounts of NO. We have used visible light spectroscopy to monitor continuously the redox state of cytochrome c oxidase in an oxygen-tight chamber, at the same time as which we measure cell respiration and the concentrations of oxygen and NO. Pharmacological manipulation of cytochrome c oxidase indicates that this enzyme, when it is in turnover and in its oxidized state, inactivates physiological amounts of NO, thus regulating its intra- and extracellular concentrations. This inactivation is prevented by blocking the enzyme with inhibitors, including NO. Furthermore, when cells generating low concentrations of NO respire toward hypoxia, the redox state of cytochrome c oxidase changes from oxidized to reduced, leading to a decrease in NO inactivation. The resultant increase in NO concentration could explain hypoxic vasodilation.     

Distribution of nitric oxide synthase in stomach wall in rats

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Nitric oxide: not just a negative inotrope

Nitric oxide (NO) appears to play a role in modulating cardiac function in both health and disease. Early studies in isolated rodent cardiac myocytes demonstrated a depressant effect of NO supplied by NO donors (exogenous) as well as NO generated within myocytes (endogenous). There is increasing evidence for a functional NO generating system within the human myocardium, which appears upregulated in certain disease states. Induction of the high output nitric oxide synthase isoform (iNOS) has been demonstrated in the failing myocardium, though its functional significance remains unproven. More recently published data have contradicted the notion that NO acts solely as a negative inotrope demonstrating positive inotropy in both isolated rodent and human ventricular myocytes in response to a range of NO donors. Different NO donors have different NO release kinetics and generate a range of NO species (NO., NO+ and NO-) which may interact at a number of subcellular targets. The observed response of any cardiac preparation to an NO donor represents the net effect of activation of different effector targets and may explain the contradictory reported effects of NO. To realise the therapeutic potential of NO will require specific targeting at a subcellular level.     

High nitric oxide synthase activity in endothelial cells in ulcerative colitis

Endothelial nitric oxide (NO) synthase, a unique NO synthase (NOS) isoform that is expressed constitutively by the vascular endothelium both in vivo and in vitro, is believed to be essential to systemic and/or local vascular integrity. NOS expression by endothelial cells may indicate vascular activation. We successfully established a simple method for the culture of microvascular endothelial cells from a small amount of tissue and investigated ulcerative colitis (UC), in which condition vascular factors have not been studied extensively. We cultured endothelial cells from the mesenteries of surgical patients with UC and assayed NOS activity by reduced nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry. Strong NOS activity was demonstrated in the cells from all UC patients (5/5), whereas no activity was detected in the cells from human umbilical veins and the mesenteries of colon cancer patients (0/10 and 0/5, respectively). This strong NOS activity was not diminished by incubation with a high concentration of glucocorticoid, suggesting that it was constitutive. These results indicate a close relationship of vascular activation (high NOS activity) with the pathogenesis of UC.