Exogenous NO Research Articles

A calcium-insensitive attenuated nitrosative stress response contributes significantly in the radioresistance of Sf9 insect cells

Lepidopteran insects/insect cells display 50–100 times higher radioresistance than humans, and are evolutionarily closest to mammals amongst all radioresistant organisms known. Compared to mammalian cells, Lepidopteran cells (TN-368, Sf9) display more efficient antioxidant system and DNA repair and suffer considerably less radiation-induced DNA/cytogenetic damage and apoptosis. Recent studies indicate that a considerably lower radiation-induced oxidative stress may significantly reduce macromolecular damage in Lepidopteran cells. Since nitrosative stress contributes in radiation-induced cellular damage, we investigated its nature in the γ-irradiated Sf9 cells (derived from Spodoptera frugiperda; order Lepidoptera; family Noctuidae) and compared with BMG-1 human cell line having significant NOS expression. Radiation induced considerably less ROS/RNS in Sf9 cells, which remained unchanged on treatment with NOS inhibitor l-NMMA. Surprisingly, growth of Sf9 cultures or irradiation could not induce NO or its metabolites, indicating negligible basal/radiation-induced NOS activity that remained unchanged even after supplementation with arginine. Cytosolic calcium release following high-dose (1000–2000 Gy at 61.1 cGy s −1) γ-irradiation or H 2O 2 (250 μM) treatment also failed to generate NO in Sf9 cells having high constitutive levels of calmodulin, whereas BMG-1 cells displayed considerable calcium-dependent NO generation even following 10 Gy dose. These results strongly imply the lack of calcium-mediated NOS activity in Sf9 cells. Addition of exogenous NO from GSH-NO caused considerable increase in radiation-induced apoptosis, indicating significant contribution of constitutively attenuated nitrosative stress response into the radioresistance of Lepidopteran cells. Our study demonstrates for the first time that a calcium-insensitive, attenuated nitrosative stress response may contribute significantly in the unusual radioresistance displayed by Lepidopteran insect cells.

Physiological effects of exogenous nitric oxide on Brassica juncea seedlings under NaCl stress

The study was conducted to investigate the physiological effects of exogenous NO on potherb mustard (Brassica juncea Coss.) seedlings under salt stress. The plants were grown in Hogland nutrient solution for 15 d and treated with 150 mM NaCl, NO donor sodium nitropruside (SNP) and NO scavenger methylene blue (MB-1) for 4 d. The NaCl stress increased superoxide dismutase, peroxidase and ascorbate peroxidase activities and malondialdehyde (MDA) and free proline contents, and decreased soluble protein content. However, the application of exogenous NO limited the production of MDA and free proline, while markedly promoted SOD, POD and APX activity.

Rationale and design of a trial on the role of bosentan in Fontan patients: Improvement of exercise capacity?

Background The Fontan circulation is a palliative procedure performed in patients with complex congenital heart disease (CHD), making transpulmonary blood flow dependent on the systemic venous pressure. In a Fontan circulation a low pulmonary vascular resistance (PVR) is crucial, as is epitomized by the observation that a high PVR is a strong predictor of mortality. Long-term follow-up has shown that PVR may rise many years after the Fontan procedure has been performed, possibly due to micro-emboli from a dilated right atrium or from the venous system. Other mechanisms of increased PVR might be aging, obstructed airways caused by lymphatic dysfunction, lack of pulsatile pulmonary flow causing a release of endothelium-derived vasoactive molecules, and prolonged overexpression of vasoconstrictors such as endothelin-1. Mean plasma level of endothelin-1 has been shown to be significantly higher in Fontan patients compared to healthy controls. In patients with pulmonary arterial hypertension (PAH), therapy with bosentan, an endothelin-1 receptor antagonist, has demonstrated to improve exercise capacity and to reduce the elevated PVR. In addition, reduction of PVR is shown early and late after the Fontan procedure on treatment with exogenous NO, another advanced PAH therapy. However, the long term effect of reducing the PVR by bosentan treatment on exercise capacity in Fontan patients is still unknown. Methods We designed a prospective, multicenter, randomized open label trial to study the effect of bosentan in Fontan patients. The primary endpoint will be the change in maximum exercise capacity (peak V'O2). Conclusion We hypothesize that treatment with bosentan, an endothelin-1 receptor antagonist, improves maximum exercise capacity and functional capacity in adult Fontan patients.

1371 NF-κ/INOS/CAMP SIGNALING CASCADE MEDIATES INFLAMMATORY CYTOKINES-INDUCED UPREGULATION OF CONNEXIN43 EXPRESSION AND FUNCTION IN URINARY TRACT INFECTION

You have accessJournal of UrologyInfections/Inflammation of the Genitourinary Tract: Kidney & Bladder1 Apr 20111371 NF-κ/INOS/CAMP SIGNALING CASCADE MEDIATES INFLAMMATORY CYTOKINES-INDUCED UPREGULATION OF CONNEXIN43 EXPRESSION AND FUNCTION IN URINARY TRACT INFECTION Kai Li, Jian Yao, Norifumi Sawada, Hiroshi Nakagomi, Tatsuya Miyamoto, Satoru Kira, Hideki Kobayashi, Takayuki Tsuchida, Masanori Kitamura, and Masayuki Takeda Kai LiKai Li Chuo City, Japan More articles by this author , Jian YaoJian Yao Chuo City, Japan More articles by this author , Norifumi SawadaNorifumi Sawada Chuo City, Japan More articles by this author , Hiroshi NakagomiHiroshi Nakagomi Chuo City, Japan More articles by this author , Tatsuya MiyamotoTatsuya Miyamoto Chuo City, Japan More articles by this author , Satoru KiraSatoru Kira Chuo City, Japan More articles by this author , Hideki KobayashiHideki Kobayashi Chuo City, Japan More articles by this author , Takayuki TsuchidaTakayuki Tsuchida Chuo City, Japan More articles by this author , Masanori KitamuraMasanori Kitamura Chuo City, Japan More articles by this author , and Masayuki TakedaMasayuki Takeda Chuo City, Japan More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.1195AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Urinary tract infection (UTI) is one of the most common infectious diseases in the clinic, which can produce symptoms similar to those experienced with overactive bladder. At present little information is available regarding the mechanisms involved in the urinary inflammation-related symptoms and pathogenesis. Given that intercellular communication via gap junctions (GJ) plays an important role in inflammatory responses and that the enhanced GJ protein connexin43 (Cx43) contributes to myogenic bladder overactivity, we asked whether and how gap junctions in bladder are influenced by inflammatory cytokines. METHODS Bladder smooth muscle cells (BSMCs) isolated from Sprague-Dawley rats were analyzed for Cx43 expression and function after stimulation with inflammatory cytokines, interleukin 1 beta (IL-1β) and tumor necrosis factor alpha (TNFα), by Western blot, immunofluorescent staining and scrape-loading dye-transfer assay. The implicated signal mechanisms were identified by using various kinase inhibitors and specific siRNA. In vivo mouse inflammatory model was induced by intraperitoneal injection of LPS. The expressions of Cx43 in bladder were examined by histocytochemical and western blot analysis. RESULTS (1) Incubation of BSMCs with IL-1β and TNFα resulted in a time-dependent elevation in Cx43 protein levels. This was associated with obviously increased membrane localization of Cx43 protein and GJ function. (2) Increment of Cx43 by the inflammatory cytokines was preceded by an activation of PKA, as revealed by VASP phosphorylation. Blockade of PKA signaling pathway with PKA inhibitor H89, cAMP inhibitor Rp-cAMP or CREB siRNA could largely abolish the effects. (3) The Cx43-elevating effects could also be abolished by NFκb inhibitor SC514 and by iNOS inhibitor L-NAME. Further studies revealed that SC514 abrogated iNOS expression, and exogenous NO donor SNAP, also induced an elevation in Cx43. (4) Intraperitoneal injection of LPS into mouse resulted in an elevated Cx43 expression and membrane localization in bladder. CONCLUSIONS In conclusion, we demonstrated, for the first time, that inflammatory cytokines induced Cx43 expression and function in BSMCs through mechanisms involving NFκB/iNOS/ cAMP signaling cascade. Our finding may open a new window towards our understanding of the molecular mechanisms implicated in inflammatory urinary disorders and provide a possible explanation for urge symptoms seen in bladder infection. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e547 Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Kai Li Chuo City, Japan More articles by this author Jian Yao Chuo City, Japan More articles by this author Norifumi Sawada Chuo City, Japan More articles by this author Hiroshi Nakagomi Chuo City, Japan More articles by this author Tatsuya Miyamoto Chuo City, Japan More articles by this author Satoru Kira Chuo City, Japan More articles by this author Hideki Kobayashi Chuo City, Japan More articles by this author Takayuki Tsuchida Chuo City, Japan More articles by this author Masanori Kitamura Chuo City, Japan More articles by this author Masayuki Takeda Chuo City, Japan More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Activation of GluR6-containing Kainate Receptors Induces Ubiquitin-dependent Bcl-2 Degradation via Denitrosylation in the Rat Hippocampus after Kainate Treatment

We previously showed that Bcl-2 (B-cell lymphoma 2) is down-regulated in a kainate (KA)-induced rat epileptic seizure model. The underlying mechanism had remained largely unknown, but we here report for the first time that denitrosylation and ubiquitination are involved. Our results show that the S-nitrosylation levels of Bcl-2 are down-regulated after KA injection and that the GluR6 (glutamate receptor 6) antagonist NS102 can inhibit the denitrosylation of Bcl-2. Moreover, the ubiquitin-dependent degradation of Bcl-2 was found to be promoted after KA treatment, which could be suppressed by the proteasome inhibitor MG132 and the NO donors, sodium nitroprusside and S-nitrosoglutathione. In addition, experiments based on siRNA transfections were performed in the human SH-SY5Y neuroblastoma cell line to verify that the stability of Bcl-2 is causal to neuronal survival. At the same time, it was found that the exogenous NO donor GSNO could protect neurons when Bcl-2 is targeted. Subsequently, these mechanisms were morphologically validated by immunohistochemistry, cresyl violet staining, and in situ TUNEL staining to analyze the expression of Bcl-2 as well as the survival of CA1 and CA3/DG pyramidal neurons. NS102, GSNO, sodium nitroprusside, and MG132 contribute to the survival of CA1 and CA3/DG pyramidal neurons by attenuating Bcl-2 denitrosylation. Taken together, our data reveal that Bcl-2 ubiquitin-dependent degradation is induced by Bcl-2 denitrosylation during neuronal apoptosis after KA treatment.

Pathophysiology of sickle cell disease is mirrored by the red blood cell metabolome

AbstractEmerging metabolomic tools can now be used to establish metabolic signatures of specialized circulating hematopoietic cells in physiologic or pathologic conditions and in human hematologic diseases. To determine metabolomes of normal and sickle cell erythrocytes, we used an extraction method of erythrocytes metabolites coupled with a liquid chromatography-mass spectrometry–based metabolite profiling method. Comparison of these 2 metabolomes identified major changes in metabolites produced by (1) endogenous glycolysis characterized by accumulation of many glycolytic intermediates; (2) endogenous glutathione and ascorbate metabolisms characterized by accumulation of ascorbate metabolism intermediates, such as diketogulonic acid and decreased levels of both glutathione and glutathione disulfide; (3) membrane turnover, such as carnitine, or membrane transport characteristics, such as amino acids; and (4) exogenous arginine and NO metabolisms, such as spermine, spermidine, or citrulline. Finally, metabolomic analysis of young and old normal red blood cells indicates metabolites whose levels are directly related to sickle cell disease. These results show the relevance of metabolic profiling for the follow-up of sickle cell patients or other red blood cell diseases and pinpoint the importance of metabolomics to further depict the pathophysiology of human hematologic diseases.

Endothelium-Dependent Contractions in Hypertension

Most isolated arteries respond to shear stress and several vasodilator substances, as demonstrated first for acetylcholine,1 by releasing endothelium-derived relaxing factor (or nitric oxide [NO]), and various endothelium-derived hyperpolarizing signals.2,–,5 However, in certain blood vessels, when exposed to stretch, agonists such as thrombin, acetylcholine, and adenosine nucleotides (adenosine diphosphate [ADP] and adenosine triphosphate [ATP]) or calcium ionophores, the endothelium produces diffusible cyclooxygenase (COX)-derived vasoconstrictor prostanoids (endothelium-derived contracting factors [EDCF])6,–,11 Endothelial cells also produce vasoconstrictor peptides, in particular, endothelin-1.12 The attribution of a role to endothelin-1 in instantaneous changes in vascular tone has been made difficult by the almost insurmountable nature of the vasoconstriction caused by the peptide that can only be tempered by NO or calcitonin gene-related peptide.13,14 Likewise, the evidence linking acute release of endothelin-1 to constriction of arteries is still limited.15 Therefore, the present article focuses on the mechanisms leading to the production of endothelial COX-derived vasoconstrictors, in particular, in the rat aorta, which has been the standard preparation used by the author and his collaborators for the study of EDCF-mediated responses However, the occurrence of such EDCF-mediated responses can vary widely, depending on the species and the blood vessel studied. For example, they are prominent in canine veins but not arteries.6 In the mouse, endothelium-dependent contractions are more pronounced in the carotid artery than in the aorta.16,17 Further, in any given blood vessel, the production of endothelium-derived vasoconstrictor prostanoids is exacerbated by aging and disease, in particular, hypertension.10,11,18,19 ### The Trigger: Calcium Acetylcholine (which activates endothelial M3-muscarinic receptors)20 and ADP and ATP (which activate endothelial purinoceptors)21,22 evoke endothelium-dependent contractions and release of calcium from the sarcoplasmic reticulum.23 Lowering the …

Identification of a Lactate-Quinone Oxidoreductase in Staphylococcus aureus that is Essential for Virulence

Staphylococcus aureus is an important human pathogen commonly infecting nearly every host tissue. The ability of S. aureus to resist innate immunity is critical to its success as a pathogen, including its propensity to grow in the presence of host nitric oxide (NO·). Upon exogenous NO· exposure, S. aureus immediately excretes copious amounts of L-lactate to maintain redox balance. However, after prolonged NO·-exposure, S. aureus reassimilates L-lactate specifically and in this work, we identify the enzyme responsible for this L-lactate-consumption as a L-lactate-quinone oxidoreductase (Lqo, SACOL2623). Originally annotated as Mqo2 and thought to oxidize malate, we show that this enzyme exhibits no affinity for malate but reacts specifically with L-lactate (KM = ∼330 μM). In addition to its requirement for reassimilation of L-lactate during NO·-stress, Lqo is also critical to respiratory growth on L-lactate as a sole carbon source. Moreover, Δlqo mutants exhibit attenuation in a murine model of sepsis, particularly in their ability to cause myocarditis. Interestingly, this cardiac-specific attenuation is completely abrogated in mice unable to synthesize inflammatory NO· (iNOS−/−). We demonstrate that S. aureus NO·-resistance is highly dependent on the availability of a glycolytic carbon sources. However, S. aureus can utilize the combination of peptides and L-lactate as carbon sources during NO·-stress in an Lqo-dependent fashion. Murine cardiac tissue has markedly high levels of L-lactate in comparison to renal or hepatic tissue consistent with the NO·-dependent requirement for Lqo in S. aureus myocarditis. Thus, Lqo provides S. aureus with yet another means of replicating in the presence of host NO·.

NO signaling in exercise training-induced anti-apoptotic effects in human neutrophils

Short-lived neutrophils play a predominant role in innate immunity, the effects of exercise training on neutrophil survival is unclear. In this study, we investigated the underlying mechanisms of training effects on human neutrophil apoptosis. Healthy male subjects were trained on a cycling ergometer for 8 weeks and followed by 4 weeks of detraining. Blood neutrophils were collected before exercise, after training, and after detraining. Comparing with pre-exercise specimens, neutrophils collected after training showed reduced apoptosis rate, which partially returned after detraining. Various intracellular proteins, including iNOS, Mcl-1, A1, Grp78, and IL-8, were upregulated by training, and they remained high after detraining. Upregulated iNOS was closely correlated with these anti-apoptotic molecules in neutrophils. Furthermore, the possible mechanism by which iNOS suppressed apoptosis was explored. Neutrophil apoptosis was accelerated by blocking and retarded by stimulating the endogenous iNOS activity. As an anti-apoptosis mediator of NO signaling, the Mcl-1 level dropped by depletion of the major NO downstream molecule cGMP and such loss of Mcl-1 was avoidable when supplying exogenous NO. Upon activation of NO-cGMP signaling, neutrophils held increased Mcl-1 expression and delayed apoptosis. Collectively, our results suggested that exercise training may retard neutrophil apoptosis by upregulating the iNOS-NO-cGMP-Mcl-1 pathway.

Fungal endophyte-induced volatile oil accumulation in Atractylodes lancea plantlets is mediated by nitric oxide, salicylic acid and hydrogen peroxide

Inoculation with endophytic fungus Gilmaniella sp. induced multiple responses in host plantlets of Atractylodes lancea, including the burst of NO, SA, H2O2, and the accumulation of volatile oil. To investigate the pathways that the three signal molecules affect the volatile oil production, the plantlets were treated with NO specific scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide potassium salt (cPTIO), SA synthesis inhibitor trans-cinnamic acid (trans-CA), membrane NADPH oxidase inhibitor diphenylene iodonium (DPI) and catalase (CAT). The results showed pretreatment of plantlets with cPTIO, DPI/CAT and trans-CA inhibited the burst of signal molecules and volatile oil accumulation induced by the fungus. Exogenous NO donor SNP could promote SA and H2O2 production as well as volatile oil accumulation. Combined application of trans-CA and DPI/CAT could inhibit the promotion effects of exogenous NO. Exogenous SA and H2O2 had no effects on NO production, but they could reverse the total inhibitory effects of cPTIO, CAT/DPI and trans-CA toward volatile oil accumulation. Exogenous H2O2 had promotion effect on SA production, yet CAT/DPI could not significantly prevent SA synthesis. These data indicated NO mediates violate oil accumulation induced by the fungus through SA and H2O2 dependent pathways. H2O2 can regulate SA production, but does not act as upstream signal molecule.

Effect of exogenous Ca2+ and NO donor SNP on seed germination and antioxidase activities of Perilla frutescens seedlings under NaCl stress

In order to find a method for improving the salt resistance of seeds and seedlings for Perilla frutescens under NaCl stress, seed germination and physiological characteristics of P. frutescens seedlings were studied. Several physiological indexes of P. frutescens seeds treated by Ca2+ and sodium nitroprusside (SNP) under NaCl stress like the germination vigor, germination rate, germination index and vigor index were measured. And other indexes like the biomass of the seedlings, the content of malondialdehyde (MDA) in leaves, the activities of superoxide (SOD), peroxidase (POD) and catalase (CAT) were also measured. The germination of P. frutescens seeds under NaCl stress was inhibited obviously. But after the treatment with Ca2+ and SNP, all of the germination indexes increased. And the seeds that treated with SNP + Ca2+ had the most significantly increase in all indexes. The germination vigor was 65.1%, the germination rate was 89.3%, the germination index and vigor index were 13.9 and 0.1109, respectively. The content of MDA decreased after the treatment. The activities of three enzymes include SOD, POD and CAT were increased by the treatment and get the maximin 0.84, 5.71 and 4.92 U x mg(-1) respectively. And the EGTA showed an obvious inhibition to the effect of SNP on P. frutescens. SNP (0.1 mmol x L(-1)) and Ca2+ (10 mmol x L(-1)) could significantly alleviate the damages to the seeds and seedlings of P. frutescens under NaCl stress, and promote the salt resistance of the seeds and seedlings. These results might suggest that exogenous NO might enhance P. frutescens salt resistance and alleviate salt injury possible by enhancing Ca2+ influx by activating Ca2+ channels and improving concentration of Ca2+ of P. frutescens seedlings.

English

  Exogenous sodium nitroprusside (SNP), a NO donor, was applied in this study to investigate the potential role of NO in photosynthetic performance of tomato (Lycopersicon esculentum L. cv. Hufan2560) seedlings under salt-stressed conditions. Exogenous NO alleviated the decrease in dry mass of shoot and root caused by salt stress. In parallel, NO application in salt-stressed plants attenuated the decrease in the photosynthetic parameters such as leaf chlorophyll, net photosynthetic rate (PN), stomatal conductance (gs), transpiration rate (E), the ratio of variable to maximum fluorescence (Fv/Fm), electron transport rate (ETR), the efficiency of excitation energy capture by open photosystem II (PSII) reaction centers (Fv´/Fm´), and the photochemical quenching coefficient (qP), and counteracted the increase in on-photochemical quenching coefficient (qN). Furthermore, the changes as mentioned above reversed by NO treatment are specific to salt stress since application of NO alone to tomato seedlings without salt stress had slight effects on the tested parameters. The results obtained here demonstrated that the photosynthetic performance was improved by NO application in salt-stressed plants and such an improvement was associated with an enhancement of gas-exchange and the actual PSII efficiency, which revealed an important role of NO in enhancing resistance of plants to salt stress.   Key words: Nitric oxide, photosynthesis, salt stress, Lycopersicon esculentum L., stomatal conductance, transpiration rate.

Influence of exogenous NO donator and variations in the Ca2+ content on transport activity related to tonoplast proton pumps in ontogenesis and under hyperosmotic stress

The changes in transport activity of tonoplast proton pumps under the influence of exogenous NO donator and modulation of Ca2+ concentration jointly and separately were investigated at different stages of ontogenesis and under hyperosmotic stress. The results suggest that both exogenous NO donator and Ca2+ ions can influence the activity of transport processes related to tonoplast and this influence is especially evident in the period of growth and accumulation of metabolites. Under hyperosmotic stress, H+-pyrophosphatase plays a more important role than H+-ATPase: the activity of the former increases 2.3-fold compared to the control osmotic conditions, whereas the activity of H+-ATPase is practically unchanged. H+-pyrophosphatase was more responsive to the presence of exogenous NO donator and to variations in Ca2+ concentration. The effects of exogenous NO donator on tonoplast proton pumps depended on the concentration of Ca2+, which apparently can mediate NO action.

Dual role of nNOS in ischemic injury and preconditioning

BackgroundNitric oxide (NO) is cardioprotective and a mediator of ischemic preconditioning (IP). Endothelial nitric oxide synthase (eNOS) is protective against myocardial ischemic injury and a component of IP but the role and location of neuronal nitric oxide synthase (nNOS) remains unclear. Therefore, the aims of these studies were to: (i) investigate the role of nNOS in ischemia/reoxygenation-induced injury and IP, (ii) determine whether its effect is species-dependent, and (iii) elucidate the relationship of nNOS with mitoKATP channels and p38MAPK, two key components of IP transduction pathway.ResultsVentricular myocardial slices from rats and wild and nNOS knockout mice, and right atrial myocardial slices from human were subjected to 90 min ischemia and 120 min reoxygenation (37°C). Specimens were randomized to receive various treatments (n = 6/group). Both the provision of exogenous NO and the inhibition of endogenous NO production significantly reduced tissue injury (creatine kinase release, cell necrosis and apoptosis), an effect that was species-independent. The cardioprotection seen with nNOS inhibition was as potent as that of IP, however, in nNOS knockout mice the cardioprotective effect of non-selective NOS (L-NAME) and selective nNOS inhibition and also that of IP was blocked while the benefit of exogenous NO remained intact. Additional studies revealed that the cardioprotection afforded by exogenous NO and by inhibition of nNOS were unaffected by the mitoKATP channel blocker 5-HD, although it was abrogated by p38MAPK blocker SB203580.ConclusionsnNOS plays a dual role in ischemia/reoxygenation in that its presence is necessary to afford cardioprotection by IP and its inhibition reduces myocardial ischemic injury. The role of nNOS is species-independent and exerted downstream of the mitoKATP channels and upstream of p38MAPK.

The effect of nitric oxide and hydrogen peroxide in the activation of the systemic immune response of Anopheles albimanus infected with Plasmodium berghei

The expression of genes encoding the antimicrobial peptides (AMPs) attacin, cecropin and gambicin, as well as the effects of NO and H 2O 2 on their expression was investigated in midguts and fat bodies of Anopheles albimanus during the midgut infection with Plasmodium berghei. Midgut infection induced an increase in the expression of the three AMPs in both tissues; while NO and H 2O 2 were present in haemolymph. Treatment with L-NAME and vitamin C reduced the effect of P. berghei infection on the AMP's expression, and exogenous NO and H 2O 2 induced their expression in the mosquito fat body. The induction of AMPs in abdominal tissues, while the malaria parasites are in the mosquito midgut, suggests communication between the midgut epithelial cells and the abdominal tissue which has not yet had direct contact with the parasites. Free radical production in mosquito midgut and haemolymph during Plasmodium infection and their inductive effect on AMPs in abdominal tissues indicates the possible participation of these radicals in mediating a systemic immune response in this mosquito.

Trigonal Bipyramidal {Fe(NO)}7 Complex [(NO)Fe(SC9H6N)2] Containing an Equatorial Nitrosyl Ligand: The Critical Role of Chelating Ligands in Regulating the Geometry and Transformation of Mononitrosyl Iron Complex (MNIC)

AbstractTrigonal bipyramidal {Fe(NO)}7 complex [(NO)Fe(SC9H6N)2] (2) with NO ligand occupying equatorial position, obtained from nitrosylation of complex [(Cl)Fe(SC9H6N)2]− (1) accompanied by elimination of [Cl]−, was characterized by IR, UV‐Vis, EPR, SQUID and single‐crystal X‐ray diffraction. Compared to the S = 3/2 ground state displayed by the trigonal bipyramidal complex [Et4N][(ON)Fe(N‐(CH2CH2S)3)], the chelating ligands ([SC9H6N]−) play a key role in modulating the position of exogenous NO ligand and controlling the spin state of complex 2 (S = 1/2). In contrast to nitrosylation of [Fe(NO)(SPh)3]− MNIC yielding [Fe(NO)2(SPh)2]− DNIC, the inertness of complex 2 toward NO(g) was observed. Reaction of [NO][BF4] and complex 2 in a 1:1 stoichiometry led to the formation of homodinuclear {Fe(NO)}7‐{Fe(NO)}9 complex [(ON)Fe(‐SC9H6N)2Fe(NO)2][BF4] (3) characterized by IR, UV‐vis, 1H NMR and single‐crystal X‐ray diffraction. Antiferromagnetic coupling between the S = 1/2 {Fe(NO)}7 [S2Fe(NO)2] core and the S = 1/2 {Fe(NO)}7 [(NO)FeS2N2] core may account for the diamagnetism and the EPR silence of complex 3. This study provides the evidence that the geometric and electronic structures of {Fe(NO)}7 MNICs modulated by the coordination environment of iron center determines the nitrosylation products.

Nitric Oxide Plays a Key Role in Myelination in the Developing Brain

Inhaled nitric oxide (iNO) is one of the most promising therapies used in neonates, but there is little information available about its effect on the developing brain. We explored the effects of both iNO and endogenous NO on developing white matter in rodents. Rat or mouse pups and their mothers were placed in a chamber containing 5 to 20 ppm of NO for 7 days after birth. Neonatal exposure to iNO was associated with a transient increase in central nervous system myelination in rats and C57BL/6 mice without any deleterious effects at low doses (5 ppm) or behavioral consequences in adulthood. Exposure to iNO was associated with a proliferative effect on immature oligodendrocytes and a subsequent promaturational effect. The role of endogenous NO in myelination was investigated in animals treated with the nitric oxides synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME) in the neonatal period; this led to protracted myelination defects and subsequent behavioral deficits in adulthood. These effects were reversed by rescuing L-NAME-treated animals with iNO. Thus, we demonstrate considerable effect of both exogenous and endogenous NO on myelination in rodents. These data point to potential new avenues for neuroprotection in human perinatal brain damage.

Involvement of nitric oxide synthase-dependent nitric oxide and exogenous nitric oxide in alleviating NaCl induced osmotic and oxidative stress in Arabidopsis thaliana

To elucidate the roles of endogenous nitric oxide on Arabidopsis thaliana tolerance to salt stress, a moderate concentration of NaCl was applied to wild-type (WT) and mutant (Atnoa1) plants which have an impaired in vivo nitric oxide synthase and reduced endogenous nitric oxide content due to T-DNA insertion in the first exon of the NOA1 gene. The exhibited greater inhibition of root growth, higher leaf water loss (LWL), lower contents of chlorophyll, soluble protein, proline, higher activities of peroxidase (POD), ascorbate peroxide (APX), and gluthinone reductase (GR), it also showed lower activities of superoxide dismutase (SOD) and catalase (CAT) than wild-type plants under NaCl stress. The nitric oxide synthase (NOS) inhibitor NG-nitro-L-Arg (L-NNA) enhanced NaCl induced growth inhibition, osmotic stress, and oxidative stress in wild-type plants. Meanwhile the NO donor, sodium nitroprusside (SNP), alleviated the NaCl induced damages in Atnoa1 plants. These results indicate that both NOS-dependent endogenous NO and exogenous NO were involved in salt resistance in A. thaliana.

Protein Tyrosine Nitration of Aldolase in Mast Cells: A Plausible Pathway in Nitric Oxide-Mediated Regulation of Mast Cell Function

NO is a short-lived free radical that plays a critical role in the regulation of cellular signaling. Mast cell (MC)-derived NO and exogenous NO regulate MC activities, including the inhibition of MC degranulation. At a molecular level, NO acts to modify protein structure and function through several mechanisms, including protein tyrosine nitration. To begin to elucidate the molecular mechanisms underlying the effects of NO in MCs, we investigated protein tyrosine nitration in human MC lines HMC-1 and LAD2 treated with the NO donor S-nitrosoglutathione. Using two-dimensional gel Western blot analysis with an anti-nitrotyrosine Ab, together with mass spectrometry, we identified aldolase A, an enzyme of the glycolytic pathway, as a target for tyrosine nitration in MCs. The nitration of aldolase A was associated with a reduction in the maximum velocity of aldolase in HMC-1 and LAD2. Nuclear magnetic resonance analysis showed that despite these changes in the activity of a critical enzyme in glycolysis, there was no significant change in total cellular ATP content, although the AMP/ATP ratio was altered. Elevated levels of lactate and pyruvate suggested that S-nitrosoglutathione treatment enhanced glycolysis. Reduced aldolase activity was associated with increased intracellular levels of its substrate, fructose 1,6-bisphosphate. Interestingly, fructose 1,6-bisphosphate inhibited IgE-mediated MC degranulation in LAD2 cells. Thus, for the first time we report evidence of protein tyrosine nitration in human MC lines and identify aldolase A as a prominent target. This posttranslational nitration of aldolase A may be an important pathway that regulates MC phenotype and function.

Nitric oxide is not involved in Neisseria gonorrhoeae-induced cellular damage of human Fallopian tubes in vitro.

In the present study, we investigated whether cellular damage, as demonstrated by lactate dehydrogenase (LDH) release in the human fallopian tube (FT) infected by Neisseria gonorrhoeae (Ngo), correlated with high levels of nitric oxide synthase (NOS) mRNA and enzyme activity. Infection with Ngo induced a significant increase (~35-fold) in mRNA transcripts of the inducible isoform of NOS. Paradoxically, a reduction in NOS enzyme activity was observed in infected cultures, suggesting that gonococcal infection possibly influences translation of iNOS mRNA to the enzyme. In addition, treatment with the NOS inhibitor TRIM did not prevent gonococcal-induced cellular damage. In contrast, the addition of the inhibitor L-NAME induced a 40% reduction in LDH release, which correlated with a ~50% reduction in gonococcal numbers. Moreover, treatment of normal FT explants with an exogenous NO donor, SNAP, did not induce significant cellular damage. Taken together, our data suggest that NO does not contribute to cellular damage during infection of the human FT with Neisseria gonorrhoeae.