The NADPH-dependent thioredoxin system constitutes a functional backup for cytosolic glutathione reductase in Arabidopsis

Tight control of cellular redox homeostasis is essential for protection against oxidative damage and for maintenance of normal metabolism as well as redox signaling events. Under oxidative stress conditions, the tripeptide glutathione can switch from its reduced form (GSH) to oxidized glutathione disulfide (GSSG), and thus, forms an important cellular redox buffer. GSSG is normally reduced to GSH by 2 glutathione reductase (GR) isoforms encoded in the Arabidopsis genome, cytosolic GR1 and GR2 dual-targeted to chloroplasts and mitochondria. Measurements of total GR activity in leaf extracts of wild-type and 2 gr1 deletion mutants revealed that ≈65% of the total GR activity is attributed to GR1, whereas ≈35% is contributed by GR2. Despite the lack of a large share in total GR activity, gr1 mutants do not show any informative phenotype, even under stress conditions, and thus, the physiological impact of GR1 remains obscure. To elucidate its role in plants, glutathione-specific redox-sensitive GFP was used to dynamically measure the glutathione redox potential (E_GSH) in the cytosol. Using this tool, it is shown that E_GSH in gr1 mutants is significantly shifted toward more oxidizing conditions. Surprisingly, dynamic reduction of GSSG formed during induced oxidative stress in gr1 mutants is still possible, although significantly delayed compared with wild-type plants. We infer that there is functional redundancy in this critical pathway. Integrated biochemical and genetic assays identify the NADPH-dependent thioredoxin system as a backup system for GR1. Deletion of both, NADPH-dependent thioredoxin reductase A and GR1, prevents survival due to a pollen lethal phenotype.     

低硒大鼠心肌线粒体型硫氧还蛋白还原酶的活性和表达

目的 探讨低硒状态下大鼠心肌线粒体型硫氧还蛋白还原酶(TR2)的活性和表达变化，分析体内硒水平改变对TR2活性的影响及其可能的机制。方法 用低硒饲料喂养断乳2周的Wistar大鼠14周，分别采用生物化学方法、Western blot和Northern blot杂交检测心肌组织中TR2的活性改变及TR2蛋白、基因表达变化。结果 低硒组大鼠心肌TR2活性降低至对照组的69．3％，差异有非常显著意义。蛋白杂交的峰面积密度扫描结果显示，常硒组为3538，低硒组为2167。Northern blot结果显示，低硒组与对照组差异无显著意义。结论 低硒能够降低心肌TR2的活性和蛋白表达水平，但并不影响此酶的基因表达。提示低硒导致心肌TR2活性降低可能是由于TR2蛋白合成过程中SeCys插入减少所致。     

硒和蛋白质对大鼠心肌形态结构及谷胱甘肽过氧化物酶和线粒体型硫氧还蛋白还原酶表达的影响

目的 观察硒和蛋白质对大鼠心肌形态结构及谷胱甘肽过氧化物酶(GPX1)、线粒体型硫氧还蛋白还原酶(TR2)表达的影响.方法 选取健康纯系断乳雄性Wistar大鼠60只,按2×2析因设计分为4组,每组15只.饮水分为低硒(0 mg/L)、常硒(0.25 mg/L)2个水平;饲料分别为低蛋白(10％蛋白+0.008 mg/kg硒)、常蛋白(20％蛋白+ 0.015～0.026 mg/kg硒)2个水平.饲养1年后处死,光镜下观察心肌组织病理改变,免疫组化法和Westem blot法检测GPX1和TR2在大鼠心肌组织的表达水平.结果 各组大鼠心肌坏死率比较,差异有统计学意义(x2=11.04,P＜ 0.05),其中低硒低蛋白组[66.7％(8/12)]明显高于常硒常蛋白组[7.1％(1/14),x2=11.06,P＜ 0.05].免疫组化法检测,低硒低蛋白组、常硒低蛋白组、低硒常蛋白组、常硒常蛋白组大鼠心肌组织GPX1表达阳性率分别为0(0/12)、81.8％(9/11)、10.0％(1/10)、100.0％(14/14),其中常硒低蛋白组和常硒常蛋白组阳性率明显高于低硒低蛋白组和低硒常蛋白组(x2值分别为12.88、8.14和35.89、32.60,P均＜ 0.05);TR2阳性率分别为0(0/12)、81.8％(9/11)、0(0/10)、100.0％(14/14),其中常硒低蛋白组和常硒常蛋白组阳性率明显高于低硒低蛋白组和低硒常蛋白组(x2值分别为28.67、18.25和35.89、32.60,P均＜0.05).Western blot法检测,上述4组大鼠心肌组织GPX1蛋白表达分别为0.87±0.13、1.18±0.13、0.95±0.13、1.74±0.23,经析因分析,硒和蛋白质水平对心肌细胞GPX1表达具有影响作用(F值分别为124.93、43.16,P均＜0.05),且硒和蛋白质间有交互作用(F=24.10,P＜0.05);TR2蛋白表达分别为0.63±0.19、0.97±0.24、0.55±0.08、1.03±0.31,经析因分析,硒因素对心肌细胞TR2表达具有影响作用(F=36.97,P＜0.05).结论 常硒和常蛋白相对于低硒和低蛋白都可以提高心脏GPX1和TR2表达水平,增强心肌组织抗氧化能力,保护心肌内皮细胞,降低心肌的损伤程度,且二者联合作用较好.     

Role of nitric oxide,tetrahydrobiopterin and peroxynitrite in glucose toxicity-associated contractile dysfunction in ventricular myocytes

Aims/hypothesis Local overproduction of nitric oxide is seen in early stages of diabetes, which can react with superoxide (O₂^–) to form peroxynitrite (ONOO^–). The aim of this study was to examine the effect of scavengers for nitric oxide, O₂^–, ONOO^– and NOS cofactor tetrahydrobiopterin (BH₄) on high glucose-induced cardiac contractile dysfunction.Methods Ventricular myocytes were cultured for 24 h with either normal (N, 5.5 mmol/l) or high (25.5 mmol/l) glucose, with or without the nitric oxide scavengers haemoglobin (100 nmol/l), PTIO (100 µmol/l), the NOS inhibitor L-NMMA (100 µmol/l), superoxide dismutase (SOD, 500 U/ml), the ONOO^– scavengers urate (100 µmol/l), MnTABP (100 µmol/l), BH₄ (10 µmol/l) and its inactive analogue NH₄ (10 µmol/l), and the GTP cyclohydrolase I inhibitor DAHP (1 mmol/l). Myocyte mechanics, NOS protein expression and activity were evaluated.Results High glucose myocytes showed reduced peak shortening, decreased maximal velocity of shortening/relengthening (± dL/dt), prolonged relengthening (TR₉₀) and normal shortening duration (TPS) associated with reduced cytosolic Ca²⁺ rise compared to normal myocytes. The high glucose-induced abnormalities were abrogated or attenuated by urate, MnTBAP, L-NMMA, BH₄, and SOD, whereas unaffected by haemoglobin, PTIO and NH₄. L-NMMA reduced peak shortening while PTIO and DAHP depressed ± dL/dt and prolonged TPS or TR₉₀ in normal myocytes. High glucose increased NOS activity, protein expression of eNOS but not iNOS, which were attenuated by L-NMMA and BH₄, respectively.Conclusion/interpretation These results suggested that NOS cofactor, NO and ONOO^– play a role in glucose-induced cardiomyocyte contractile dysfunction and in the pathogenesis of diabetic cardiomyopathy.Abbreviations PTIO 2-(4-carboxyphenyl)-4,4,5,5-tetramethyl imidazoline-1-oxyl 3-oxide - DAHP 2,4-diamino-6-hydroxy-pyrimidine - E-C excitation-contraction - MnTABP manganese (III) tetrakis (4-benzoic acid) porphyrin - ± dL/dt maximal velocity of shortening and relengthening - L-NAME N-nitro-L-arginine methyl ester - L-NMMA L-N^G-monomethyl-arginine - NOS nitric oxide synthase - PS peak shortening - ONOO^– peroxynitrite - O₂^– superoxide anion - SOD superoxide dismutase - BH₄ tetrahydrobiopterin - NH₄ tetrahydroneopterin - TPS time-to-peak shortening - TR₉₀ time-to-90% relengthening     

Effect of melatonin on cellular energy depletion mediated by peroxynitrite and poly (ADP-ribose) synthetase activation in an acute model of inflammation

DNA single-strand breakage and activation of the nuclear enzyme poly (ADP-ribose) synthetase (PARS) triggers an energy-consuming, inefficient repair cycle, which contributes to peroxynitrite-induced cellular injury. Recently, we proposed that during an acute model (pleurisy), cellular injury is mediated by peroxynitrite formation and consequent PARS activation. Here, we investigated whether in vivo melatonin treatment inhibits cellular injury induced by peroxynitrite production and PARS activation in macrophages collected from rats subjected to carrageenan-induced pleurisy. Macrophages harvested from the pleural cavity exhibited a significant production of peroxynitrite, as measured by the oxidation of the fluorescent dye dihydrorhodamine 123. Furthermore, carrageenan-induced pleurisy caused a suppression of macrophage mitochondrial respiration, DNA strand breakage, activation of PARS, and reduction of cellular levels of NAD+. In vivo treatment with melatonin [12.5 or 25 or 50 mg/kg, intraperitoneally (i.p.), 30 min before carrageenan] significantly reduced peroxynitrite formation in a dose-dependent manner and prevented the appearance of DNA damage, decrease in mitochondrial respiration, loss of cellular levels of NAD+, and PARS activation. Our study supports the view that the antioxidant and anti-inflammatory effect of melatonin is also correlated with the inhibition of peroxynitrite production and PARS activation. In conclusion, melatonin may be a novel pharmacological approach to prevent cell injury in acute inflammation.     

Protective effect of melatonin on cellular energy depletion mediated by peroxynitrite and poly (ADP-ribose) synthetase activation in a non-septic shock model induced by zymosan in the rat

Abstract: DNA single-strand breakage and activation of the nuclear enzyme poly (ADP-ribose) synthetase (PARS) triggers an energy-consuming, inefficient repair cycle, which contributes to peroxynitrite-induced cellular injury. Recently it was proposed that zymosan, a non-bacterial agent, causes cellular injury by inducing the production of peroxynitrite and consequent PARS activation. Here we investigated whether in vivo melatonin treatment inhibits cellular injury induced by peroxynitrite production and PARS activation in macrophages collected from rats subjected to zymosan-induced shock. Macrophages harvested from the peritoneal cavity exhibited a significant production of peroxynitrite, as measured by the oxidation of the fluorescent dye dihydrorhodamine 123. Furthermore, zymosan-induced shock caused a suppression of macrophage mitochondrial respiration, DNA strand breakage, activation of PARS and reduction of cellular levels of NAD⁺. In vivo treatment with melatonin (25 and 50 mg/kg, intraperitoneally, 1 hr after zymosan injection) significantly reduced in dose-dependent manner peroxynitrite formation and prevented the appearance of DNA damage, the decrease in mitochondrial respiration, the loss of cellular levels of NAD⁺, and the PARS activation. Our study supports the view that the antioxidant and antiinflammatory effect of melatonin is also correlated with the inhibition of peroxynitrite production and PARS activation. In conclusion, melatonin may be a novel pharmacological approach to prevent cell injury in inflammation.     

The beneficial effects of melatonin against heart mitochondrial impairment during sepsis: inhibition of iNOS and preservation of nNOS

While it is accepted that the high production of nitric oxide (NO˙) by the inducible nitric oxide synthase (iNOS) impairs cardiac mitochondrial function during sepsis, the role of neuronal nitric oxide synthase (nNOS) may be protective. During sepsis, there is a significantly increase in the expression and activity of mitochondrial iNOS (i‐mtNOS), which parallels the changes in cytosolic iNOS. The existence of a constitutive NOS form (c‐mtNOS) in heart mitochondria has been also described, but its role in the heart failure during sepsis remains unclear. Herein, we analyzed the changes in mitochondrial oxidative stress and bioenergetics in wild‐type and nNOS‐deficient mice during sepsis, and the role of melatonin, a known antioxidant, in these changes. Sepsis was induced by cecal ligation and puncture, and heart mitochondria were analyzed for NOS expression and activity, nitrites, lipid peroxidation, glutathione and glutathione redox enzymes, oxidized proteins, and respiratory chain activity in vehicle‐ and melatonin‐treated mice. Our data show that sepsis produced a similar induction of iNOS/i‐mtNOS and comparable inhibition of the respiratory chain activity in wild‐type and in nNOS‐deficient mice. Sepsis also increased mitochondrial oxidative/nitrosative stress to a similar extent in both mice strains. Melatonin administration inhibited iNOS/i‐mtNOS induction, restored mitochondrial homeostasis in septic mice, and preserved the activity of nNOS/c‐mtNOS. The effects of melatonin were unrelated to the presence or the absence of nNOS. Our observations show a lack of effect of nNOS on heart bioenergetic impairment during sepsis and further support the beneficial actions of melatonin in sepsis.     

Reduced nerve blood flow in diabetic rats: relationship to nitric oxide production and inhibition of aldose reductase

This study examined links between impaired nitric oxide production in the sciatic endoneurium, nerve blood flow, and polyol pathway flux, to test the hypothesis that reduced nerve blood flow might be compromised by competition for NADPH between aldose reductase and nitric oxide synthase. Sciatic nerves of streptozotocin-diabetic rats showed reduced laser Doppler flux (by 51 % or 63 %; both p<0.05)—indicative of reduced nerve blood flow—and reduced motor nerve conduction velocity (17 % in two experiments; p<0.05). Acute interruption of nitric oxide production in the sciatic nerves of control rats, via endoneurial injection of Nω-nitro-D -arginine methyl ester ( L -NAME), caused a local reduction (of 64 %; p<0.001) in nerve Doppler flux. This was reversed by either L -arginine or sodium nitroprusside. The response to L -NAME was greatly reduced in diabetic rats (only 22 % reduction; p<0.01), though both L -arginine and SNP caused marked increases in flux. Chronic inhibition of aldose reductase in diabetic rats (with either sorbinil or imirestat at a range of doses) had little effect on resting sciatic nerve Doppler flux, though both inhibitors normalized conduction velocity. Both aldose reductase inhibitors reduced sorbitol pathway intermediates in a dose-related manner. These findings do not support the proposition that aldose reductase inhibitors normalise conduction velocity by mechanisms dependent upon either normalization of endoneurial nitric oxide or nerve blood flow. Instead, a mechanism based upon more direct effects on axon or Schwann cell function is favoured. © 1998 John Wiley & Sons, Ltd.     

Effects of interleukin‐6 treatment on neurovascular function,nerve perfusion and vascular endothelium in diabetic rats

Aim: Interleukin‐6 (IL‐6), a member of the neuropoietic cytokine family, participates in neural development and has neurotrophic activity. Recent research has also indicated actions to improve vasa nervorum function in diabetes. Both these facets are potentially relevant for treatment of diabetic neuropathy. The aim of this study was to determine whether IL‐6 treatment corrected changes in neurovascular function in streptozotocin‐induced diabetic rats. Methods: After 1 month of diabetes, rats were given IL‐6 for 1 month. The rats were subjected to sensory testing and measurements of nerve conduction velocities and nerve blood flow by hydrogen clearance microelectrode polarography. Further groups were used to study responses of the isolated gastric fundus and renal artery. Results were statistically analysed using ANOVA and post hoc tests. Results: Diabetic rats showed mechanical hyperalgesia, thermal hyperalgesia, and tactile allodynia. The former was unaffected by IL‐6 treatment, whereas the latter two measures were corrected. Immunohistochemical staining of dorsal root ganglia for IL‐6 did not reveal any changes with diabetes or treatment. The results showed that 22 and 17.4% slowing of sciatic motor and saphenous sensory nerve conduction velocities, respectively, with diabetes were improved by IL‐6. Sciatic endoneurial perfusion was halved by diabetes and corrected by IL‐6. A 40.6% diabetic deficit in maximal non‐adrenergic, non‐cholinergic relaxation of gastric fundus to nerve stimulation was unaffected by IL‐6. Renal artery endothelium‐dependent relaxation was halved by diabetes, the endothelium‐derived hyperpolarizing factor (EDHF) component being severely attenuated. IL‐6 did not affect nitric oxide‐mediated vasorelaxation, but markedly improved EDHF responses. Conclusions: IL‐6 improved aspects of small and large nerve fibre and vascular endothelium dysfunction in diabetic rats. The functional benefits related to increased nerve blood flow via an EDHF mechanism, and IL‐6 could have therapeutic potential in diabetic neuropathy and vasculopathy, which should be further evaluated.     

Reactions of the melatonin metabolite AMK (N1-acetyl-5-methoxykynuramine) with reactive nitrogen species: formation of novel compounds, 3-acetamidomethyl-6-methoxycinnolinone and 3-nitro-AMK

The melatonin metabolite N1-acetyl-5-methoxykynuramine (AMK) was found to be unstable in air when adsorbed on a thin-layer silica gel chromatography plate, a result that is in good agreement with the relatively high reactivity of this compound. Three novel main products were separated from the reaction mixture and identified by mass spectrometry and nuclear magnetic resonance data as: (i) 3-acetamidomethyl-6-methoxycinnolinone (AMMC), (ii) 3-nitro-AMK (AMNK, N1-acetyl-5-methoxy-3-nitrokynuramine), and (iii) N-[2-(6-methoxyquinazolin-4-yl)-ethyl]-acetamide (MQA). AMMC and AMNK are shown to be nonenzymatically formed also in solution, by nitric oxide (NO) in the first case, and by a mixture of peroxynitrite and hydrogen carbonate, in the second one. The use of three different NO donors, PAPA-NONOate, S-nitroso-N-acetylpenicillamine and sodium nitroprussiate led to essentially the same results, with regard to a highly preferential formation of AMMC; AMNK was not detected in these reaction systems. Competition experiments with the NO scavenger N-acetylcysteine indicate a somewhat lower reactivity compared with the competitor. Peroxynitrite led to AMNK formation in the presence of physiological concentrations of hydrogen carbonate at pH 7.4, but not in its absence, indicating that nitration involves a mixture of carbonate radicals and NO2, formed from the peroxynitrite-CO2 adduct. No AMMC was detected after AMK exposure to peroxynitrite. Both AMNK and AMMC exhibited a much lower reactivity toward 2,2'-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) cation radicals than did AMK. In a competition assay for hydroxyl radicals, AMMC showed prooxidant properties, whereas AMNK was a moderate antioxidant. AMMC and AMNK should represent relatively stable physiological products, although their rates of synthesis are still unknown and may be low. Formation of these compounds may contribute to the disappearance of AMK from tissues and body fluids.     

高血压病血管内皮功能的改变及苯那普利治疗的影响

目的：探讨高血压病（EH）血管内皮功能的改变及苯那普利治疗的影响，进一步揭示EH的形成与发展机制。方法：观察EH患血浆一氧化氮（NO）和血管紧张素转换酶（ACH）的水平及苯那普利干预前、后血压的变化，并与正常组作对照。结果：EH患NO较对照组显下降（P＜0．01），ACE显升高（P＜0．01），苯那普利治疗后收缩压，舒张压都显下降（P＜0．01），NO显升高（P＜0．05），ACE显下降（P＜0．05）。结论：EH患存在血管内皮功能不良，苯那普利有显降压疗效，可显修复和改善EH患不良的血管内皮结构和功能。     

Involvement of Rho kinase (ROCK) in sepsis-induced acute lung injury

Indirect acute lung injury is associated with high morbidity and mortality. We investigated the link between Rho kinase (ROCK) activation and apoptotic cell death in sepsis induced acute lung injury. This hypothesis was tested by administering a specific, selective inhibitor of ROCK (Y-27632) to rats subjected to cecal ligation and puncture (CLP). Rats were randomly divided into 4 groups as; sham-operated, sham + Y-27632, CLP and CLP + Y-27632. Twenty-four hours later, each experiment was terminated and lungs analyzed. Histopathology was assessed by hematoxylin-eosin staining and the presence of apoptosis was evaluated through the TUNEL assay. Pulmonary activity of caspase 3 and ROCK 1 & 2 were measured by western blot. Interstitial edema, severely damaged pulmonary architecture with massive infiltration of the inflammatory cells and an increase in lung tissue TBARS levels as well as 3-NT to total tyrosine ratios were observed in untreated CLP animals. Pretreatment of animals with Y-27632, reduced lung injury in the CLP induced septic rats in each of these parameters of lung injury (p<0.05). Western immunoblot revealed active caspase cleavage and increased expression of active fragment of ROCK 1 & 2 in the CLP group. TUNEL assay showed an increase in percentage of apoptotic cells when comparing the CLP group with the CLP + Y-27632 group. These results suggest an important role of Rho kinase in sepsis induced lung injury by a mechanism that might be related to oxidative and/or nitrosative stress mediated caspase cleavage leading to apoptosis.     

Acetylsalicylate (ASA)-induced mitochondrial dysfunction and its potentiation by Ca2+

Although it has been suggested that acetylsalicylate (ASA)-induced mitochondrial dysfunction plays an important role in the pathogenesis of Reye's syndrome, administration of ASA alone does not cause this syndrome in therapeutic doses. We noted, however, that Ca²⁺ plays important roles in the regulation of cellular functions. ASA at concentrations of 250 μM or less, which had little effect on succinate-linked respiration, impaired Ca²⁺ accumulation in liver mitochondria by causing an increase in Ca²⁺ release. ASA plus Ca²⁺, which in concentrations of 150 μM or less alone had little effect on mitochondrial respiration, inhibited state 3 respiration and dinitrophenol-induced uncoupling of respiration. In addition, ASA plus Ca²⁺ increased state 4 respiration and ATPase activity. These results indicate that ASA plus Ca²⁺ impaired mitochondrial ATP synthesis, and suggest that ASA and ASA-induced Ca²⁺ increases in cytosol form a vicious circle of effects. Furthermore, oral administration of ASA (150 mg/kg for 5 days running) to rats did not affect mitochondrial structure or liver function, but resulted in aberrations of mitochondrial respiration. These results suggest that even therapeutic doses of ASA may induce alteration in mitochondrial function.     

黄芩苷对糖尿病大鼠组织醛糖还原酶活性及视网膜细胞凋亡的影响

取大鼠40只,随机分为NC组、DM组、依帕司他治疗组、黄芩苷治疗组。治疗16周后,测定视网膜组织醛糖还原酶（AR）活性及Bcl-2和Bax蛋白表达水平,发现AR活性在DM组明显升高,治疗组明显降低（P均〈0.01）。视网膜Bcl-2、Bax蛋白表达在DM组明显增加,治疗组明显减少。结果表明AR激活促进Bcl-2、Bax蛋白的表达,诱导细胞凋亡,参与DR的发生发展。醛糖还原酶抑制剂抑制AR活性,降低Bax、Bcl-2的表达,从而抑制细胞凋亡,延缓DR的发展。     

The association of aldose reductase gene (AKR1B1) polymorphisms with diabetic neuropathy in adolescents.

AIMS: Variants in the aldose reductase gene (AKR1B1) have been implicated in the development of diabetic retinopathy and nephropathy, with the most convincing data identifying a (CA)(n) repeat microsatellite allele (Z-2), which has a functional role in gene expression. In this study the association between polymorphisms in the AKR1B1 gene and diabetic neuropathy was investigated. METHODS: The pupillary response to light was used as the major outcome in this study along with abnormal hot thermal threshold. Three hundred and sixty-three adolescents underwent genotyping of the AKR1B1 gene. The microsatellite (CA)(n) repeat was sequenced and two single nucleotide polymorphisms, -106C-->T and -12C-->G, were investigated by restriction fragment length polymorphism. RESULTS: Seventy-six percent of participants had pupillary abnormalities (45% with two, 15% with three abnormalities). Presence of the Z-2/Z-2 genotype increased the risk nearly three-fold for pupillary abnormalities [odds ratio (OR) 3.02, 95% confidence interval (CI) 1.14, 7.98). The susceptibility genotypes (Z-2/Z-2 with -106C/-106C, Z-2/Z with -106C/-106C or Z/Z with -106C/-106C) were associated with resting pupil diameter abnormalities when compared with the protective genotypes (Z+2/Z+2 or -106T/-106T) (OR 2.83, 95% CI 1.25, 6.41). The combination of Z+2/-106T reduced the risk of abnormal heat discrimination (OR 0.48, 95% CI 0.23, 0.99). CONCLUSIONS: In this study we have shown that Z-2/Z-2 genotype is significantly associated with the development of pupillary abnormality, an early indicator of diabetic autonomic neuropathy, in adolescent Australian patients with Type 1 diabetes.     

L－精氨酸逆转一氧化氮抑制后的犬冠脉血流动力学改变和内皮素－1含量升高

观察了犬冠脉内灌注Ｎ－单甲基左旋精氨酸（Ｌ－ＮＭＭＡ）后再灌注Ｌ－精氨酸（Ｌ－Ａｒｇ）和单独灌注Ｌ－ＮＭＭＡ前后冠脉血流动力学、冠脉血流储备以及冠脉对不同浓度的乙酰胆碱（Ａｃｈ）反应的变化，同时用放免法测定冠脉前降支（ＬＡＤ）伴行静脉血中内皮素－１（ＥＴ－１）含量。结果发现，Ｌ－Ａｒｇ完全逆转了灌注Ｌ－ＮＭＭＡ引起的血流动力学改变，使心率回升，下降的基础冠脉流量（ＣＢＦ），从２０±８ｍｌ／ｍｉｎ回升至２８±７ｍｌ／ｍｉｎ，Ｐ＜０．０５），降低的冠脉储备恢复，从５１±１０ｍｌ／ｍｉｎ升至９４±１５ｍｌ／ｍｉｎ，Ｐ〈０．０１），ＥＴ－１的含量不再升高，从１５．５±３．０ｎｇ／Ｌ下降至５．０±２．０ｎｇ／Ｌ，Ｐ〈０．０１），Ａｃｈ介导的ＣＢＦ增加不再受到抑制（Ｐ〈０．０１）。结果提示提供外源性Ｌ－Ａｒｇ可增加一氧化氮（ＮＯ）的产生，使由于ＮＯ抑制而产生的血流动力学改变和ＥＴ－１升高发生逆转。     

The effect of bronchodilation and spirometry on fractional exhaled nitric oxide (FeNO50), bronchial NO flux (JawNO) and alveolar NO concentration (CANO) in children and young adults

Objective: Fractional exhaled nitric oxide (FeNO), bronchial nitric oxide (JawNO) and alveoar nitric oxide (C_ANO) are biomarkers of eosinophilic inflammation, usually measured simultaneously with spirometry and bronchodilation. Our aim was to investigate the effect of bronchodilation and spirometry on FeNO, C_ANO and JawNO in children and young adults with well-controlled asthma and in healthy volunteers. Methods: FeNO was measured in 95 subjects (62 controls, 33 asthmatics). C_ANO and JawNO were assessed in 41 of the subjects (35 healthy, 6 asthmatics.) Measurements were performed before spirometry (1), right after spirometry (2), 20 min after the first spirometry and bronchodilation (3), right after the post-bronchodilation spirometry (4) and 30 min after the last spirometry (5). Results: The presence of well-controlled asthma was not associated with different pattern of reaction after spirometry and bronchodilation. A statistically significant difference was observed only between FeNO4 and FeNO5, as well as between C_ANO1 and C_ANO3 (19.14 ± 1.68 vs 20.62 ± 1.85 ppb, p = 0.001 and 4.42 ± 0.40 vs 3.09±0.32 ppb, p = 0.001, respectively). Conclusions: Spirometry and bronchodilation have an insignificant effect on FeNO and JawNO. Even if a slight change occurs in FeNO and JawNO, this does not modify clinician's decision and therapeutic strategy. C_ANO values (C_ANO1) are significantly decreased 20 min after spirometry and bronchodilation.     

高血压病患者动脉平滑肌细胞NO的变化及其与发病的关系

为探讨高血压病（ＥＨ）患者一氧化氮（ＮＯ）浓度变化特点与高血压病发病的关系，本研究以复合胶元酶法分离培养ＥＨ患者和血压正常者（ＮＴ）离体动脉平滑肌细胞（ＶＳＭＣ），检测细胞培养液的ＮＯ含量和ＶＳＭＣ的一氧化氮合酶（ＮＯＳ）活性，观察ＥＨ患者动脉ＶＳＭＣ分泌ＮＯ的特点。结果显示:①ＥＨ组ＶＳＭＣ的ＮＯ含量和ＮＯＳ活性均显著低于ＮＴ组（Ｐ＜０．０１）；②ＥＨ患者和ＮＴＶＳＭＣ的ＮＯ含量和ＮＯＳ活性随培养时间延长均显著地增加（Ｐ＜０．０１）；③ＥＨ患者和ＮＴ的ＮＯ含量随ＮＯＳ活性增强而增加，并呈显著的正相关（Ｐ＜０．０１）。表明ＥＨ患者ＶＳＭＣ的ＮＯＳ－ＮＯ系统可能存在功能或结构的异常，并参与高血压病的发生和发展