Effect of prenatal lead exposure on nigrostriatal neurotransmission and hydroxyl radical formation in rat neostriatum: dopaminergic-nitrergic interaction

The present study was designed to explore the role of ontogenetic lead (Pb(2+)) exposure on a putative dopaminergic-nitrergic interaction in the nigrostriatal pathway. Pregnant Wistar rats were given tap water containing 250-ppm lead acetate, for the duration of pregnancy, with regular tap water (without Pb(2+)) being substituted at birth. Control rats were derived from dams that consumed tap water throughout pregnancy, and had no exposure to Pb(2+) afterwards. At 12 weeks after birth in vivo microdialysis of the neostriatum was employed to demonstrate that maternal Pb(2+) exposure was without effect on the baseline dopamine (DA) microdialysate concentration as well as amphetamine (AMPH, 1.0mg/kg i.p.)-evoked release of striatal DA. Also, prenatal Pb(2+) exposure did not enhance AMPH- and 7-nitroindazole (neuronal nitric oxide synthase inhibitor) (7-NI, 20mg/kg i.p.)-induced hydroxyl radical (HO) formation in the striatum, as indicated by analysis of the salicylate spin-trap product 2,5-dihydroxybenzoic acid. However, in rats exposed prenatally to Pb(2+), the facilitatory effect of 7-NI on DA exocytosis was attenuated. On the basis of the current study we conclude that maternal Pb(2+) exposure distorts the dopaminergic-nitrergic interaction in the nigrostriatal pathway, but without involvement of reactive oxygen species (ROS).     

钙、锌对铅致大鼠记忆障碍的保护机制探讨

目的　主要探讨饲料钙、锌干预对脑发育期低水平铅暴露致大鼠仔鼠学习记忆障碍保护作用的可能机制。方法　大鼠孕 14d至仔鼠出生 40d饮 10 0mg L含铅水 ,进食加钙 (1 2 5 % )和加锌 (10 0mg kg)配方饲料组 ,观察仔鼠生长发育情况 ,仔鼠 40d取股动脉血分析全血铅含量 ;取右半侧大脑分析脑铅含量 ;取左侧海马和小脑分析NO含量 ,右侧海马分析蛋白激酶C(PKC)活性。结果　两干预组仔鼠脑铅平均值分别为 3 0 5和 0 62 μg g ,明显低于进食普通配方饲料组 ,而加钙组血铅平均值为 0 2 9μmol L,明显低于加锌和普通配方组 ;两干预组海马和小脑一氧化氮 (NO)平均值分别为 2 7 68和 13 82 μmol g组织蛋白 ,均高于普通配方组 ,而海马细胞胞浆蛋白激酶C活性平均值分别为 0 3 3和 0 3 8pmol (min .μg组织蛋白 ) ,均低于普通配方组。结论　饲料中钙、锌干预对低水平铅暴露致大鼠学习记忆障碍的保护作用可能与降低其体内血、脑中铅蓄积水平 ,升高海马和小脑NO含量以及降低海马细胞胞浆内PKC活性有关     

Activation of ERbeta increases levels of phosphorylated nNOS and NO production through a Src/PI3K/Akt-dependent pathway in hypothalamic neurons

Estrogen plays a role in restoring homeostatic balance during the stress response by altering hypothalamic function and NO production in the brain. While we know that estrogen acts on the hypothalamus to stimulate the NO system through an ERbeta-dependent mechanism in neurons, the molecular mechanisms responsible for these effects are unknown. Because phosphorylation of nNOS at Ser(1412) increases nNOS activity which leads to increased NO production, we investigated the effects of ERbeta activation on nNOS phosphorylation at Ser(1412) and NO production in primary hypothalamic neurons. Using the selective ERbeta agonist, DPN (10nM), we show that activation of ERbeta rapidly increases phosphorylation levels of nNOS at Ser(1412) and NO production. We also show that the PI3K pathway, but not the MAPK pathway, mediates the increases in levels of Ser(1412) phosphorylation and NO production induced by ERbeta activation, as the selective PI3K inhibitor, LY294002 (10muM), blocked the effects of ERbeta activation. Finally, we demonstrate that Src kinase acts upstream of the PI3K/Akt pathway based on our finding that the selective Src inhibitor, PP2 (10muM), blocked the increases in nNOS phosphorylation levels, NO production, and PI3K/Akt activity induced by ERbeta activation. Together, our results show that Src kinase mediates ERbeta-induced increases in phosphorylation levels of nNOS at Ser(1412) and NO production by activating the PI3K/Akt pathway. These findings provide novel insight into the signaling mechanisms through which E2 stimulates the NO system in hypothalamic neurons.     

白藜芦醇对高脂处理人脐静脉内皮细胞SIRT1表达、eNOS活性及NO分泌的影响

目的:探讨白藜芦醇对高脂处理人脐静脉内皮细胞(HUVEC)组蛋白去乙酰化酶1(SIRT1)表达、内皮型一氧化氮合酶(eNOS)活性及一氧化氮(NO)分泌的影响。方法:将HUVEC分为正常对照组、高脂组及高脂+白藜芦醇预处理组,培养24h后采用逆转录-聚合酶链反应法及Western印迹法分别检测培养细胞中SIRT1的mRNA及蛋白表达水平;分别以硝酸还原酶法、化学比色法检测培养液中的NO分泌量和eNOS活性。结果:与高脂组比较,白藜芦醇可增强SIRT1的mRNA及蛋白表达水平,升高NO分泌量及增强eNOS活性(P<0.05)。结论:白藜芦醇可能通过增强SIRT1表达及促进NO分泌而具有保护内皮细胞免受高脂损伤的作用。     

Role of miR-199b-5p in regulating angiogenesis in mouse myocardial microvascular endothelial cells through HSF1/VEGF pathway

Our study explored effects of miR-199b-5p on angiogenesis in mouse myocardial microvascular endothelial cells (MMVECs) and the involved working mechanisms. We applied explant culture to incubate C57/BL6 mouse MMVECs. Lipofection was used to transfect miR-199b-5p mimic, miR-199b-5p inhibitor and miR-199b-5p scramble respectively. MMVECs were divided into miR-199b-5p up-regulation, miR-199b-5p down-regulation and control groups based on above sequence. Expressions of miR-199b-5p, heat shock factor protein 1 (HSF1) mRNA were assessed by real-time quantitative polymerase chain reaction (RT-QPCR). Expressions of HSF1 and vascular endothelial growth factor (VEGF) were assessed by Western Blotting. Cell proliferation was assessed by CCK8. Tubule formation assay was conducted to assess formation of blood vessels. Results showed that miR-199b-5p up/down-regulation groups exhibited no obvious differences in the expressions of HSF1 mRNA compared to control group. However, miR-199b-5p up-regulation group recorded lower expressions of HSF1 and VEGF in the level of protein, and reduced cell proliferation and tubule formation. Whereas, miR-199b-5p down-regulation group presented the contrary results. The experiment indicated that miR-199b-5p can regulate proliferation and angiogenesis in mouse MMVECs through the pathway of HSF1/VEGF.     

Involvement of NO/cGMP pathway in toluene-induced locomotor hyperactivity in female rats

Rationale Nitric oxide (NO) is implicated in the acute locomotor activating effects of some addictive drugs such as amphetamine, caffeine, and PCP, but has not been investigated in the case of toluene.Objectives This study determined the contribution of the NO-cyclic GMP (cGMP) pathway to locomotor stimulant effects of toluene.Methods Locomotor activity was measured for 90 min immediately following toluene (500–1,000 mg/kg, IP) or corn oil treatments in Sprague-Dawley female rats. A NO generator, sodium nitroprusside (SNP) (3 and 6 mg/kg), a NO precursor, l-arginine (l-Arg) (250 mg/kg), a NO synthase inhibitor,N^G-nitro-l-arginine methyl ester (l-NAME) (5–20 mg/kg, IP), and a soluble guanylyl cyclase inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (10 mg/kg) were injected 5 min before toluene (750 mg/kg, IP) treatment. The combination effects of SNP with l-NAME, l-arginine with l-NAME, SNP with ODQ and l-arginine with ODQ on toluene-induced locomotor hyperactivity were also determined.Results The locomotor hyperactivity induced by toluene was significantly inhibited by SNP and l-arginine, but enhanced by l-NAME and ODQ. SNP and l-arginine completely reversed the combined effects of l-NAME and toluene to a basal level and abolished the enhancing effects of ODQ.Conclusions The results suggested that NO/cGMP-dependent mechanism might be involved in toluene-induced locomotor activity in rats.     

铅对海马神经元型一氧化氮合酶及其基因表达的影响

目的　观察实验性铅中毒对小鼠海马神经元型一氧化氮合酶 (nNOS)及其基因表达的影响 ,探讨铅致学习记忆损害的分子毒理学机制。方法　小鼠用 5g L醋酸铅溶液染毒。采用逆转录 聚合酶链反应 (RT PCR)方法 ,半定量分析染毒 2、4和 8周小鼠海马nNOS基因表达的变化。采用尼克酰胺腺嘌呤二核苷酸磷酸黄递酶 (NADPH d)组织化学染色法分析铅染毒对小鼠海马NOS阳性神经元的影响。结果　nNOSmRNA存在于正常小鼠海马组织中 ;铅染毒后仅 2周 ,海马nNOSmRNA的含量显著减少 ;随着染毒时间的增加 ,海马nNOSmRNA的含量逐渐下降。镜下观察 ,视野中正常小鼠海马平均NOS阳性细胞数为 ( 5 7 63± 11 5 )个 ,而铅染毒组仅为 ( 2 9 3 5± 9 10 )个。结论　铅可使小鼠海马nNOS及其基因表达下降     

肝硬化患者血一氧化氮、一氧化氮酶检测及其与内毒素的关系

目的　探讨肝硬化患者血一氧化氮 (NO)、一氧化氮酶 (NOS)的含量及其与内毒素的关系。方法　分别应用酶法和比色法测定 4 9例肝硬化患者血 NO、NOS及内毒素的含量 ,并与 35名正常人作对照。结果　肝硬化患者血 NO、NOS、内毒素水平均明显高于正常人组 (P<0 .0 5～ 0 .0 1)。结论　肝硬化患者血 NO NOS水平的增高与内毒素含量的增高有着密切的关系     

新生儿缺氧缺血性脑病患者血NO/NOS和TNF测定的临床意义

目的　探讨了新生儿缺氧缺血性脑病患者血清 NO/ NOS和 TNF含量及其临床意义。方法　应用生化法和放射免疫分析法对 38例新生儿缺氧缺血性脑病患者进行了血清 NO/ NOS和 TNF含量检测 ,并与 35名正常新生儿作比较。结果　血清患儿组血清 NO水平低于正常人组 (P<0 .0 1 ) ,而 NOS和 TNF水平则显著高于正常人组(P<0 .0 1 )。结论　测定血清 NO/ NOS和 TNF含量与患儿的危重程度发生发展、预后有关     

Intermedin alleviates the inflammatory response and stabilizes the endothelial barrier in LPS-induced ARDS through the PI3K/Akt/eNOS signaling pathway

Inflammatory storms and endothelial barrier dysfunction are the central pathophysiological features of acute respiratory distress syndrome (ARDS). Intermedin (IMD), a member of the calcitonin gene-related peptide (CGRP) family, has been reported to alleviate inflammation and protect endothelial cell (EC) integrity. However, the effects of IMD on ARDS have not been clearly elucidated. In the present study, clinical ARDS data were used to explore the relationship between serum IMD levels and disease severity and prognosis, and we then established a model to predict the possibility of hospital survival. Mouse models of ARDS and LPS-challenged endothelial cells were used to analyze the protective effect and underlying mechanism of IMD. We found that in patients with ARDS, increased serum IMD levels were associated with reduced disease severity and increased rates of hospital survival. IMD alleviated the LPS-induced inflammatory response by decreasing proinflammatory cytokines, NF-κB p65 expression and NF-κB p65 nuclear translocation. In addition, IMD stabilized the endothelial barrier by repairing adherens junctions (AJs), cytoskeleton and capillary leakage. IMD exerted protective effects against ARDS on pulmonary endothelial cells, at least partly, through PI3K/Akt/eNOS signaling, while IMD’s anti-inflammation effect was mediated through an eNOS-independent mechanism. Our study may provide new therapeutic insight for ARDS treatment.     

血浆高密度脂蛋白胆固醇及NOS NO水平与冠心病发病相关性研究

目的研究低高密度脂蛋白胆固醇(HDL-C)血症与血浆一氧化氮合酶(NOS)和一氧化氮(NO)的相关性,探讨低HDL-C致动脉粥样硬化的可能机制。方法使用全自动生化分析仪测定血脂,选取79例低HDL-C患者和70名正常人群作为对照组,分别用化学比色法测定血浆NOS和用硝酸酶还原法测定血浆NO。结果低HDL-C组血浆NOS和血浆NO水平均显著低于健康对照组(P<0.05)。结论血浆低HDL-C可导致血浆NOS和NO水平显著降低,从而导致冠心病的发病率增高。     

大鼠创伤性脑损伤早期脑组织NO含量和NOS活性的变化

目的 探讨一氧化氮（NO）和神经元型NO合酶（nNOS）是否参与创伤性脑损伤（TBI）的发病机理。方法 采用落体法大鼠TBI动物模型,观察大鼠TBI早期脑组织NO含量、NOS活性的动态变化和特异性nNOS抑制剂7-硝基吲唑（7-NI）对其的影响。结果大鼠TBI后30min脑组织NO含量和NOS活性显著升高,伤后2h两者回落,但仍高于对照组;伤后6h两者接近对照组;伤后12hNO含量仍接近对照组,而NOS活性再次高于对照组。结论 NO和nNOS在创伤性脑损伤机制中可能起重要作用。     

Investigation of a possible interaction between the heme oxygenase/biliverdin reductase and nitric oxide synthase pathway in murine gastric fundus and jejunum

This study investigated the possible interaction between the heme oxygenase (HO)/biliverdin reductase (BVR) and nitric oxide synthase (NOS) pathway in murine gastric fundus and jejunum, since previous studies have shown that both HO-2 and BVR are expressed in interstitial cells of Cajal (ICCs) and co-localized with neuronal NOS in a large proportion of myenteric neurons along the gastrointestinal tract. Neither HO inhibition by chromium mesoporphyrin (CrMP) nor co-incubation with CO or biliverdin/bilirubin affected nitrergic neurotransmission - i.e. relaxations induced by non-adrenergic non-cholinergic (NANC) nerve stimulation or exogenous NO - under normal physiological conditions. However, biliverdin/bilirubin reversed the inhibitory effect of the superoxide generator LY83583 on exogenous NO-induced relaxations in both tissues. When gastric fundus muscle strips were depleted of the endogenous antioxidant Cu/Zn superoxide dismutase (SOD) by the Cu-chelator DETCA, electrically induced NANC relaxations were also affected by LY82583; however, biliverdin/bilirubin could not substitute for the loss of Cu/Zn SOD when this specific antioxidant enzyme was depleted. In jejunal muscle strips, the combination DETCA plus LY83583 nearly abolished contractile phasic activity and, hence, did not allow studying nitrergic relaxation in these experimental conditions. In conclusion, this study does not establish a role for HO/CO in inhibitory NANC neurotransmission in murine gastric fundus and jejunum under normal physiological conditions. However, the antioxidants biliverdin/bilirubin might play an important role in the protection of the nitrergic neurotransmitter against oxidative stress.     

硒和/或维生素E对实验性高脂血症大鼠心、肝、肾及血清中一氧化氮及一氧化氮合酶的影响

目的 :观察硒和/或维生素E(VE)对实验性高脂血症大鼠心、肝、肾、血清一氧化氮 (NO)及一氧化氮合酶 (NOS)的影响。方法 :对大鼠喂以高脂饲料致实验性高脂血症 ,然后分别分组给予硒和/或VE ,4wk后取血及心、肝、肾组织匀浆 ,采用硝酸还原酶等方法测定上述组织的NO和NOS含量。结果 :高脂饲料可致血清、心、肝、肾组织中NO含量及NOS活性降低 ;硒和/或VE能不同程度地增加这些组织中NO含量及心、肝、肾中的NOS活性 (P<0 05或P<0 01) ,且两者合用比单用作用更明显。结论 :硒和/或VE可致实验性高脂血症大鼠心、肝、肾及血清中的NO和NOS发生改变。     

Mechanisms mediating the cardioprotective effect of carvedilol in cadmium induced cardiotoxicity. Role of eNOS and HO1/Nrf2 pathway

Cadmium (Cd) is a highly toxic heavy metal with several harmful effects including cardiotoxicity. For the first time, we aimed to evaluate the possible cardioprotective effect of carvedilol (CAR) in Cd induced cardiotoxicity and study the mechanisms involved in such protection including endothelial nitric oxide synthase (eNOS) and HO1/Nrf2 pathway. CAR (1,10 mg/kg/d) was administered orally for 4 weeks with Cd induced cardiac injury (3 mg/kg/d) orally for 4 weeks. We measured cardiac enzymes, mean arterial pressure changes, heme oxygenase-1 (HO1) and total antioxidant capacity (TAC). Moreover; cardiac tissue malondialdehyde (MDA), tumor necrosis factor alpha (TNFα), western blotting of caspase3 and eNOS levels and histopathology were evaluated. Immunoexpression of eNOS in cardiac tissue, gene expression changes of HO1, and nuclear factor erythroid 2-related factor 2 (Nrf2) using real time polymerase chain reactions (rtPCR) were detected. Our results showed that CAR could significantly decrease Cd induced cardiotoxicity.     

The effects of nitric oxide in acute lung injury

Acute lung injury (ALI) is a common clinical problem associated with significant morbidity and mortality. Ongoing clinical and basic research and a greater understanding of the pathophysiology of ALI have not been translated into new anti-inflammatory therapeutic options for patients with ALI, or into a significant improvement in the outcome of ALI. In both animal models and humans with ALI, there is increased endogenous production of nitric oxide (NO) due to enhanced expression and activity of inducible NO synthase (iNOS). This increased presence of iNOS and NO in ALI contributes importantly to the pathophysiology of ALI. However, inhibition of total NO production or selective inhibition of iNOS has not been effective in the treatment of ALI. We have recently suggested that there may be differential effects of NO derived from different cell populations in ALI. This concept of cell-source-specific effects of NO in ALI has potential therapeutic relevance, as targeted iNOS inhibition specifically to key individual cells may be an effective therapeutic approach in patients with ALI. In this paper, we will explore the potential role for endogenous iNOS-derived NO in ALI. We will review the evidence for increased iNOS expression and NO production, the effects of non-selective NOS inhibition, the effects of selective inhibition or deficiency of iNOS, and this concept of cell-source-specific effects of iNOS in both animal models and human ALI.     

银杏黄酮苷对氧化损伤内皮细胞产生NO、eNOS和ICAM-1的影响

目的观察银杏黄酮苷对氧化损伤的人脐静脉内皮细胞（HUVEC）产生NO、内皮型一氧化氮合酶（eNOS）和人可溶性细胞间黏附分子（ICAM-1）的影响。方法体外培养HUVEC,传至3代后,以不同浓度的银杏黄酮苷分别作用于HUVEC,然后进行氧化损伤处理。以硝酸还原酶法测定培养液上清中的NO水平,免疫细胞化学法检测内皮细胞eNOS的表达,ELISA法测定细胞培养液中ICAM-1的含量。结果HUVEC在氧化损伤（H2O2100μmol/L,2h）后产生NO的量显著减少（P〈0.01）,eNOS表达下调,ICAM-1表达上调;银杏黄酮苷可以剂量依赖性的增加内皮细胞NO生成量,上调eNOS的表达,下调ICAM-1表达。结论银杏黄酮苷可能通过增加HUVEC eNOS的表达增加N0的释放、抑制ICAM-1的表达等机制对内皮细胞起保护作用。     

用一氧化氮光学生物传感器研究供体药物体外释放的通路

目的研究一氧化氮(NO)供体药物体外释放的通路。方法用自制NO光学生物传感器通过模拟硝普钠(SNP)释放NO的3种通路所需要的体外环境,测定了不同条件的生理溶液中SNP释放的NO。结果光照和巯基化合物是引起SNP体外释放NO的主要因素。在酸化的生理液中,NO在40.0~360.0μmol.L-1的浓度范围内,线性关系良好(r=0.9992),检测限为4.0μmol.L-1,日内精密度RSD=2.3%,日间精密度RSD=3.4%。干扰试验发现,L-Cys和L-GSH在中性环境下对CytC膜有干扰,但在酸性条件下无干扰。与经典方法Griess比色法比较,无显著性差异。结论所用生物传感器系统为NO供体药物的体外释放研究提供了一种较好的手段。