Endogenous Nitric Oxide Levels Research Articles

Effect of yeast saccharide treatment on nitric oxide accumulation and chilling injury in cucumber fruit during cold storage

The effects of a saccharide fraction prepared from yeast cell walls on chilling injury and endogenous nitric oxide (NO) generation in cucumbers and the role of NO in yeast saccharide (YS)-induced cold tolerance of the fruit during postharvest storage, were investigated. Chilling injury index, malondialdehyde (MDA) content, and ion leakage of fruit treated with 0.5g/L YS were significantly reduced as compared with those of control fruit stored at 4°C, showing that YS treatment may reduce chilling injury in cucumber. Moreover, treatment with YS triggered a marked increase in endogenous NO levels. An NO scavenger and NO synthase inhibitors not only diminished YS-triggered NO generation but also suppressed YS-enhanced cold tolerance. Together, our results indicate that YS could have potential postharvest application for reducing chilling injury in cucumber fruit. Furthermore, we found that YS-induced cold tolerance is linked with the induction of endogenous NO accumulation.

Effects of nitric oxide fumigation on post-harvest core browning in ‘Yali’ pear (Pyrus bretschneideri Rehd.) during cold storage

SummaryCore browning often occurs as a physiological disorder in ‘Yali’ pear (Pyrus bretschneideri Rehd.) and results in a high losses during storage. In this study, the effects of fumigation with nitric oxide (NO) gas on the incidence of core browning in ‘Yali’ pears during cold storage were investigated. ‘Yali’ pear fruit were treated with 0, 10, 20, or 30 µl l–1 NO at 25º ± 2ºC for 3 h under anaerobic conditions, then stored at 0º ± 1ºC under normal air for up to 120 d. The data showed that fumigation with 20 µl l–1 NO was most effective at suppressing core browning. Thereafter, treatment with 20 µl l–1 NO was used for comparisons with untreated control fruit in experiments to measure changes in total phenolics contents, polyphenol oxidase (PPO) activities, and the contents of glutathione (GSH), ascorbic acid (AsA), and NO in fruit core tissue during storage. The results showed that NO-fumigated fruit had lower PPO activity, but higher GSH and AsA contents in their core tissue compared with untreated control fruit. NO fumigation also maintained higher endogenous NO levels in core tissue after 60 d in storage, while the total phenolics contents of fruit remained at lower levels until day-100 of storage. These results indicate that the inhibitory effect of fumigation with20 µl l–1 NO on core browning was associated with its effects on reducing PPO activity and total phenolics contents, while maintaining the contents of GSH and AsA in core tissue of ‘Yali’ pear during cold storage.

Increasing Nitric Oxide Content in Arabidopsis thaliana by Expressing Rat Neuronal Nitric Oxide Synthase Resulted in Enhanced Stress Tolerance

Nitric oxide (NO) plays essential roles in many physiological and developmental processes in plants, including biotic and abiotic stresses, which have adverse effects on agricultural production. However, due to the lack of findings regarding nitric oxide synthase (NOS), many difficulties arise in investigating the physiological roles of NO in vivo and thus its utilization for genetic engineering. Here, to explore the possibility of manipulating the endogenous NO level, rat neuronal NOS (nNOS) was expressed in Arabidopsis thaliana. The 35S::nNOS plants showed higher NOS activity and accumulation of NO using the fluorescent probe 3-amino, 4-aminomethyl-2', 7'-difluorescein, diacetate (DAF-FM DA) assay and the hemoglobin assay. Compared with the wild type, the 35S::nNOS plants displayed improved salt and drought tolerance, which was further confirmed by changes in physiological parameters including reduced water loss rate, reduced stomatal aperture, and altered proline and malondialdehyde content. Quantitative real-time PCR analyses revealed that the expression of several stress-regulated genes was up-regulated in the transgenic lines. Furthermore, the transgenic lines also showed enhanced disease resistance against Pseudomonas syringae pv. tomato (Pst) DC3000 by activating the expression of defense-related genes. In addition, we found that the 35S::nNOS lines flowered late by regulating the expression of CO, FLC and LFY genes. Together, these results demonstrated that it is a useful strategy to exploit the roles of plant NO in various processes by the expression of rat nNOS. The approach may also be useful for genetic engineering of crops with increased environmental adaptations.

Cold-induced endogenous nitric oxide generation plays a role in chilling tolerance of loquat fruit during postharvest storage

The effects of low temperature on endogenous nitric oxide (NO) generation and the role of endogenous NO in chilling tolerance of loquat fruit during cold storage were investigated. Low temperature at 1 °C triggered a marked increase in endogenous NO levels in loquat fruit during postharvest storage. Pretreatment of fruit with the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) not only abolished endogenous NO accumulation but also aggravated chilling injury symptoms in the fruit stored at 1 °C and 95% RH. Moreover, the cold-stored fruit in which NO accumulation is abolished by cPTIO exhibited significantly higher membrane permeability, lipid peroxidation, superoxide anion (O 2 −) production rates and hydrogen peroxide (H 2O 2) contents than the control fruit. Furthermore, the results show that abolition of endogenous NO accumulation significantly reduced activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and peroxidase (POD) in the fruit during cold storage. Together, our results suggest that cold-induced endogenous NO generation in loquat fruit during postharvest storage plays a critical role in alleviating chilling injury symptoms by affecting the antioxidative defense systems in the fruit.

Nitric oxide causes root apical meristem defects and growth inhibition while reducing PIN-FORMED 1 (PIN1)-dependent acropetal auxin transport

Nitric oxide (NO) is considered a key regulator of plant developmental processes and defense, although the mechanism and direct targets of NO action remain largely unknown. We used phenotypic, cellular, and genetic analyses in Arabidopsis thaliana to explore the role of NO in regulating primary root growth and auxin transport. Treatment with the NO donors S-nitroso-N-acetylpenicillamine, sodium nitroprusside, and S-nitrosoglutathione reduces cell division, affecting the distribution of mitotic cells and meristem size by reducing cell size and number compared with NO depletion by 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). Interestingly, genetic backgrounds in which the endogenous NO levels are enhanced [chlorophyll a/b binding protein underexpressed 1/NO overproducer 1 (cue1/nox1) mirror this response, together with an increased cell differentiation phenotype. Because of the importance of auxin distribution in regulating primary root growth, we analyzed auxin-dependent response after altering NO levels. Both elevated NO supply and the NO-overproducing Arabidopsis mutant cue1/nox1 exhibit reduced expression of the auxin reporter markers DR5pro:GUS/GFP. These effects were accompanied by a reduction in auxin transport in primary roots. NO application and the cue1/nox1 mutation caused decreased PIN-FORMED 1 (PIN1)-GFP fluorescence in a proteasome-independent manner. Remarkably, the cue1/nox1-mutant root phenotypes resemble those of pin1 mutants. The use of both chemical treatments and mutants with altered NO levels demonstrates that high levels of NO reduce auxin transport and response by a PIN1-dependent mechanism, and root meristem activity is reduced concomitantly.

Ca2+ and CaM are Involved in NO- and H2O2-Induced Adventitious Root Development in Marigold

Our previous results have demonstrated that both nitric oxide (NO) and hydrogen peroxide (H2O2) are involved in the promotion of adventitious root development in marigold (Tagetes erecta L.). However, not much is known about the intricate molecular network of adventitious root development triggered by NO and H2O2. In this study, the involvement of calcium (Ca2+) and calmodulin (CaM) in NO- and H2O2-induced adventitious rooting in marigold was investigated. Exogenous Ca2+ was capable of promoting adventitious rooting, with a maximal biological response at 50 μM CaCl2. Ca2+ chelators and CaM antagonists prevented NO- and H2O2-induced adventitious rooting, indicating that both endogenous Ca2+ and CaM may play crucial roles in the adventitious rooting induced by NO and H2O2. NO and H2O2 treatments increased the endogenous content of Ca2+ and CaM, suggesting that NO and H2O2 enhanced adventitious rooting by stimulating the endogenous Ca2+ and CaM levels. Moreover, treatment with Ca2+ enhanced the endogenous levels of NO and H2O2. Additionally, Ca2+ might be involved as an upstream signaling molecule for CaM during NO- and H2O2-induced rooting. Altogether, the results suggest that both Ca2+ and CaM are two downstream signaling molecules in adventitious rooting induced by NO and H2O2.

Nitric oxide is involved in dehydration/drought tolerance in Poncirus trifoliata seedlings through regulation of antioxidant systems and stomatal response

Nitric oxide (NO) is a component of the repertoire of signals implicated in plant responses to environmental stimuli. In the present study, we investigated the effects of exogenous application of NO-releasing donor sodium nitroprusside (SNP) and nitric oxide synthase inhibitor N(G)-nitro-L-arginine-methyl ester (L-NAME) on dehydration and drought tolerance of Poncirus trifoliata. The endogenous NO level was enhanced by SNP pretreatment, but decreased by L-NAME, in the hydroponic or potted plants with or without stresses. Under dehydration, leaves from the SNP-treated hydroponic seedlings displayed less water loss, lower electrolyte leakage and reactive oxygen species accumulation, higher antioxidant enzyme activities and smaller stomatal apertures as compared with the control (treated with water). In addition, pretreatment of the potted plants with SNP resulted in lower electrolyte leakage, higher chlorophyll content, smaller stomatal conductance and larger photosynthetic rate relative to the control. By contrast, the inhibitor treatment changed these physiological attributes or phenotypes in an opposite way. These results indicate that NO in the form of SNP enhanced dehydration and drought tolerance, whereas the inhibitor makes the leaves or plants more sensitive to the stresses. The stress tolerance by NO might be ascribed to a combinatory effect of modulation of stomatal response and activation of the antioxidant enzymes. Taken together, NO is involved in dehydration and drought tolerance of P. trifoliata, implying that manipulation of this signal molecule may provide a practical approach to combat the environmental stresses.

Effects of Nitric Oxide on the Survival and Neuritogenesis of Cerebellar Purkinje Neurons

Nitric oxide has been investigated widely both during neurodevelopment and in neurological diseases. However, whilst it has been established that nitric oxide-producing enzymes of nitric oxide synthase family are expressed in cerebellar Purkinje neurons, the effects of nitric oxide on the viability and morphology of these neurons remain unknown. Here, we have demonstrated that the activity of neuronal nitric oxide synthase, but not the inducible or endothelial forms of this enzyme, is required to support the survival of a proportion of cerebellar Purkinje neurons in vitro. We discovered that donation of high concentrations of exogenous nitric oxide reduces Purkinje neuron survival in culture and that peroxynitrite is also toxic to these cells. Finally, we demonstrated that exogenous nitric oxide and peroxynitrite reduce both the magnitude and the complexity of the neurite arbour extended by cerebellar Purkinje neurons. Taken together, these findings reveal that whilst a low level of endogenous nitric oxide, released by the activity of neuronal nitric oxide synthase, is beneficial to cerebellar Purkinje neurons in vitro, high levels of exogenous nitric oxide and peroxynitrite are detrimental to both the survival of these neurons and to their ability to extend processes and form functional neural networks.

Chronic treatment with angiotensin‐(1‐7) improves renal endothelial dysfunction in apolipoproteinE‐deficient mice

ApolipoproteinE-deficient [apoE (-/-)] mice, a model of human atherosclerosis, develop endothelial dysfunction caused by decreased levels of nitric oxide (NO). The endogenous peptide, angiotensin-(1-7) [Ang-(1-7)], acting through its specific GPCR, the Mas receptor, has endothelium-dependent vasodilator properties. Here we have investigated if chronic treatment with Ang-(1-7) improved endothelial dysfunction in apoE (-/-) mice. ApoE (-/-) mice fed on a lipid-rich Western diet were divided into three groups and treated via osmotic minipumps with either saline, Ang-(1-7) (82 µg·kg(-1) ·h(-1) ) or the same dose of Ang-(1-7) together with D-Ala-Ang-(1-7) (125 µg·kg(-1) ·h(-1) ) for 6 weeks. Renal vascular function was assessed in isolated perfused kidneys. Ang-(1-7)-treated apoE (-/-) mice showed improved renal endothelium-dependent vasorelaxation induced by carbachol and increased renal basal cGMP production, compared with untreated apoE (-/-) mice. Tempol, a reactive oxygen species (ROS) scavenger, improved endothelium-dependent vasorelaxation in kidneys of saline-treated apoE (-/-) mice whereas no effect was observed in Ang-(1-7)-treated mice. Chronic treatment with D-Ala-Ang-(1-7), a specific Mas receptor antagonist, abolished the beneficial effects of Ang-(1-7) on endothelium-dependent vasorelaxation. Renal endothelium-independent vasorelaxation showed no differences between treated and untreated mice. ROS production and expression levels of the NAD(P)H oxidase subunits gp91phox and p47phox were reduced in isolated preglomerular arterioles of Ang-(1-7)-treated mice, compared with untreated mice, whereas eNOS expression was increased. Chronic infusion of Ang-(1-7) improved renal endothelial function via Mas receptors, in an experimental model of human cardiovascular disease, by increasing levels of endogenous NO.

Nitric Oxide Regulates DELLA Content and PIF Expression to Promote Photomorphogenesis in Arabidopsis

The transition from etiolated to green seedlings involves a shift from hypocotyl growth-promoting conditions to growth restraint. These changes occur through a complex light-driven process involving multiple and tightly coordinated hormonal signaling pathways. Nitric oxide (NO) has been lately characterized as a regulator of plant development interacting with hormone signaling. Here, we show that Arabidopsis (Arabidopsis thaliana) NO-deficient mutant hypocotyls are longer than those from wild-type seedlings under red light but not under blue or far-red light. Accordingly, exogenous treatment with the NO donor sodium nitroprusside and mutant plants with increased endogenous NO levels resulted in reduced hypocotyl length. In addition to increased hypocotyl elongation, NO deficiency led to increased anthocyanin levels and reduced PHYB content under red light, all processes governed by phytochrome-interacting factors (PIFs). NO-deficient plants accordingly showed an enhanced expression of PIF3, PIF1, and PIF4. Moreover, exogenous NO increased the levels of the gibberellin (GA)-regulated DELLA proteins and shortened hypocotyls, likely through the negative regulation of the GA Insensitive Dwarf1 (GID1)-Sleepy1 (SLY1) module. Consequently, NO-deficient seedlings displayed up-regulation of SLY1, defective DELLA accumulation, and altered GA sensitivity, thus resulting in defective deetiolation under red light. Accumulation of NO in wild-type seedlings undergoing red light-triggered deetiolation and elevated levels of NO in the GA-deficient ga1-3 mutant in darkness suggest a mutual NO-GA antagonism in controlling photomorphogenesis. PHYB-dependent NO production promotes photomorphogenesis by a GID1-GA-SLY1-mediated mechanism based on the coordinated repression of growth-promoting PIF genes and the increase in the content of DELLA proteins.

Seed-specific elevation of non-symbiotic hemoglobin AtHb1: beneficial effects and underlying molecular networks in Arabidopsis thaliana.

BackgroundSeed metabolism is dynamically adjusted to oxygen availability. Processes underlying this auto-regulatory mechanism control the metabolic efficiency under changing environmental conditions/stress and thus, are of relevance for biotechnology. Non-symbiotic hemoglobins have been shown to be involved in scavenging of nitric oxide (NO) molecules, which play a key role in oxygen sensing/balancing in plants and animals. Steady state levels of NO are suggested to act as an integrator of energy and carbon metabolism and subsequently, influence energy-demanding growth processes in plants.ResultsWe aimed to manipulate oxygen stress perception in Arabidopsis seeds by overexpression of the non-symbiotic hemoglobin AtHb1 under the control of the seed-specific LeB4 promoter. Seeds of transgenic AtHb1 plants did not accumulate NO under transient hypoxic stress treatment, showed higher respiratory activity and energy status compared to the wild type. Global transcript profiling of seeds/siliques from wild type and transgenic plants under transient hypoxic and standard conditions using Affymetrix ATH1 chips revealed a rearrangement of transcriptional networks by AtHb1 overexpression under non-stress conditions, which included the induction of transcripts related to ABA synthesis and signaling, receptor-like kinase- and MAP kinase-mediated signaling pathways, WRKY transcription factors and ROS metabolism. Overexpression of AtHb1 shifted seed metabolism to an energy-saving mode with the most prominent alterations occurring in cell wall metabolism. In combination with metabolite and physiological measurements, these data demonstrate that AtHb1 overexpression improves oxidative stress tolerance compared to the wild type where a strong transcriptional and metabolic reconfiguration was observed in the hypoxic response.ConclusionsAtHb1 overexpression mediates a pre-adaptation to hypoxic stress. Under transient stress conditions transgenic seeds were able to keep low levels of endogenous NO and to maintain a high energy status, in contrast to wild type. Higher weight of mature transgenic seeds demonstrated the beneficial effects of seed-specific overexpression of AtHb1.

Nitric oxide production is associated with response to brown planthopper infestation in rice

Nilaparvata lugens Stål, the brown planthopper (BPH), is one of the most destructive phloem-feeding insects of rice ( Oryza sativa L.) throughout Asia. Here, we show that BPH feeding increases the level of endogenous nitric oxide (NO) in the leaf and sheath tissue of both resistant and susceptible rice cultivars. However, in the roots, the NO level increased in the resistant cultivar, but decreased in the susceptible one. A burst of NO production occurred in the sheath within 1 h of infestation with BPH. The production of NO in response to BPH feeding appears to be dependent primarily on the activity of nitric oxide synthase. The application of exogenous NO reduced plant water loss by its effect on both stomatal opening and root architecture. It also stimulated the expression of certain drought stress-related genes, reduced plant height and delayed leaf senescence. Over the short term, NO supplementation reduced the seedling mortality caused by BPH feeding. This suggests that NO signaling plays a role in the rice tolerance response to BPH feeding.

Nitric oxide and hydrogen peroxide are involved in indole-3-butyric acid-induced adventitious root development in marigold

SummaryPrevious results have shown that both nitric oxide (NO) and hydrogen peroxide (H2O2) play crucial roles in the promotion of adventitious root development in explants of marigold (Tagetes erecta L.). In this study, the involvement of NO and H2O2 in indole-3-butyric acid (IBA)-induced adventitious root development in marigold explants, and the order of action of these molecules in the signal transduction pathway were investigated. The results indicated that, like NO and H2O2, IBA-induced adventitious rooting occurred in a dose-dependent manner. Two parameters of root growth in explants treated simultaneously with IBA and H2O2 were significantly higher than those in explants treated with IBA or H2O2 alone, suggesting that IBA and H2O2 may act synergistically to mediate adventitious rooting. NO and H2O2 were pre-requisites for the adventitious rooting induced by IBA. Moreover, treatment with IBA enhanced the endogenous levels of NO and H2O2 in explants, indicating that the induction of adventitious roots by IBA occurred through enhancing the levels of NO and H2O2. NO and H2O2 may be downstream signal molecules in the auxin signalling cascade, and NO may be involved as an upstream signalling molecule for H2O2 production.

Reduction of Nω-hydroxy-L-arginine by the mitochondrial amidoxime reducing component (mARC)

NOSs (nitric oxide synthases) catalyse the oxidation of L-arginine to L-citrulline and nitric oxide via the intermediate NOHA (N(ω)-hydroxy-L-arginine). This intermediate is rapidly converted further, but to a small extent can also be liberated from the active site of NOSs and act as a transportable precursor of nitric oxide or potent physiological inhibitor of arginases. Thus its formation is of enormous importance for the nitric-oxide-generating system. It has also been shown that NOHA is reduced by microsomes and mitochondria to L-arginine. In the present study, we show for the first time that both human isoforms of the newly identified mARC (mitochondrial amidoxime reducing component) enhance the rate of reduction of NOHA, in the presence of NADH cytochrome b₅ reductase and cytochrome b₅, by more than 500-fold. Consequently, these results provide the first hints that mARC might be involved in mitochondrial NOHA reduction and could be of physiological significance in affecting endogenous nitric oxide levels. Possibly, this reduction represents another regulative mechanism in the complex regulation of nitric oxide biosynthesis, considering a mitochondrial NOS has been identified. Moreover, this reduction is not restricted to NOHA since the analogous arginase inhibitor NHAM (N(ω)-hydroxy-N(δ)-methyl-L-arginine) is also reduced by this system.

Nitric oxide increases mitochondrial respiration in a cGMP-dependent manner in the callus from Arabidopsis thaliana

Nitric oxide (NO) acts as a key molecule in many physiological processes in plants. In this study, the roles of NO in mitochondrial respiration were investigated in the calli from wild-type Arabidopsis and NO associated 1 mutant ( Atnoa1) which has a reduced endogenous NO level. Long-term exposure of wild-type Arabidopsis callus to sodium nitroprusside (SNP) increased mitochondrial respiration in both cytochrome and alternative pathways. In Atnoa1 callus, the capacity of both the cytochrome pathway and the alternative pathway was lower than that in wild-type callus. Further study indicated that NO enhanced the transcript abundance of genes encoding mitochondrial respiration-chain proteins as well as the protein expression of the NADH-ubiquinone reductase 75 kDa subunit and the alternative oxidase 1/2 in wild-type and Atnoa1 calli. 2-Phenyl-4,4,5,5-tetremethy-limidazolinone-1-oxyl-3-oxide (PTIO), a NO scavenger, inhibited the effects of NO in both calli. Co-incubation of callus with 1 H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), a guanylate cyclase inhibitor, also abolished NO effects. The membrane-permeable cGMP analog 8Br-cGMP mimicked NO effects. Moreover, the alternative pathway showed a higher sensitivity to the cellular cGMP changes than the cytochrome pathway did in gene transcription, protein expression and O 2 consumption. Taken together, NO could enhance mitochondrial respiration in both cytochrome and alternative pathways in a cGMP-dependent manner in Arabidopsis.

Nitric Oxide Functions as a Signal and Acts Upstream of AtCaM3 in Thermotolerance in Arabidopsis Seedlings

To characterize the role of nitric oxide (NO) in the tolerance of Arabidopsis (Arabidopsis thaliana) to heat shock (HS), we investigated the effects of heat on three types of Arabidopsis seedlings: wild type, noa1(rif1) (for nitric oxide associated1/resistant to inhibition by fosmidomycin1) and nia1nia2 (for nitrate reductase [NR]-defective double mutant), which both exhibit reduced endogenous NO levels, and a rescued line of noa1(rif1). After HS treatment, the survival ratios of the mutant seedlings were lower than those of wild type; however, they were partially restored in the rescued line. Treatment of the seedlings with sodium nitroprusside or S-nitroso-N-acetylpenicillamine revealed that internal NO affects heat sensitivity in a concentration-dependent manner. Calmodulin 3 (CaM3) is a key component of HS signaling in Arabidopsis. Real-time reverse transcription-polymerase chain reaction analysis after HS treatment revealed that the AtCaM3 mRNA level was regulated by the internal NO level. Sodium nitroprusside enhanced the survival of the wild-type and noa1(rif1) seedlings; however, no obvious effects were observed for cam3 single or cam3noa1(rif1) double mutant seedlings, suggesting that AtCaM3 is involved in NO signal transduction as a downstream factor. This point was verified by phenotypic analysis and thermotolerance testing using seedlings of three AtCaM3-overexpressing transgenic lines in an noa1(rif1) background. Electrophoretic mobility-shift and western-blot analyses demonstrated that after HS treatment, NO stimulated the DNA-binding activity of HS transcription factors and the accumulation of heat shock protein 18.2 (HSP18.2) through AtCaM3. These data indicate that NO functions in signaling and acts upstream of AtCaM3 in thermotolerance, which is dependent on increased HS transcription factor DNA-binding activity and HSP accumulation.

English

Nitric oxide (NO) is a key molecule involved in many physiology processes. The effects of NO on alleviating arsenic-induced oxidative damage in tall fescue leaves were investigated. Arsenic (25 μM) treatment induced significantly accumulation of reactive oxygen species (ROS) and led to serious lipid peroxidation in tall fescue leaves and the application of 100 μM SNP before arsenic stress resulted in alleviated arsenic-induced electrolyte leakage and malondiadehyde (MDA) content in tall fescue leaves, the levels of hydrogen peroxide (H2O2) and superoxide radical (QUOTE  ) were reduced as well. Moreover, the activities of superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX) increased in tall fescue leaves in presence of SNP under arsenic stress. This pattern was reversed by application of NO scavenger, 2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethy-limidazoline-1-oxyl-3-oxide (PTIO) before arsenic treatment. Pronounced increases in endogenous NO production was found in plants after exposure to arsenic stress. The results suggested that arsenic stress elevated endogenous NO level and that NO might act as a signaling molecule to enhance antioxidant enzyme activities, further protecting against injuries caused by arsenic toxicity.   Key words: Antioxidant enzymes, arsenic stress, nitric oxide, oxidative stress, tall fescue.

Nitrosyl ethylenediaminetetraacetate ruthenium(II) complex promotes cellular growth and could be used as nitric oxide donor in plants

Over recent years nitric oxide (NO) not only has appeared as an important endogenous signaling molecule in plants and as a mediator in many developmental and physiological processes, but has also received recognition as a plant hormone. The impressive recent achievements in elucidating the role of NO in plants have come about by the application of NO donors. The aim herein was to study the effects of the different NO donors, sodium nitroprusside (SNP) and the nitrosyl ethylenediaminetetraacetate ruthenium(II) ([Ru(NO)(Hedta)]) complex on cellular growth in embryogenic suspension cultures of Araucaria angustifolia. Appraisal of our data revealed that [Ru(NO)(Hedta)] stimulated about 60% of cellular growth in embryogenic suspension cultures of A. angustifolia, with results similar to those observed with the SNP donor. Nevertheless, application of the NO scavenger PTIO (2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide) inhibited this cellular growth in both. Cellular growth was correlated with an increase in endogenous NO levels after 21 days of culture, especially in treatments with NO donors. Our results demonstrated that the [Ru(NO)Hedta] complex could possibly be used as a NO donor in plants.

TRPV1 and TRPA1 Mediate Peripheral Nitric Oxide-Induced Nociception in Mice

Nitric oxide (NO) can induce acute pain in humans and plays an important role in pain sensitization caused by inflammation and injury in animal models. There is evidence that NO acts both in the central nervous system via a cyclic GMP pathway and in the periphery on sensory neurons through unknown mechanisms. It has recently been suggested that TRPV1 and TRPA1, two polymodal ion channels that sense noxious stimuli impinging on peripheral nociceptors, are activated by NO in heterologous systems. Here, we investigate the relevance of this activation. We demonstrate that NO donors directly activate TRPV1 and TRPA1 in isolated inside-out patch recordings. Cultured primary sensory neurons display both TRPV1- and TRPA1-dependent responses to NO donors. BH4, an essential co-factor for NO production, causes activation of a subset of DRG neurons as assayed by calcium imaging, and this activation is at least partly dependent on nitric oxide synthase activity. We show that BH4-induced calcium influx is ablated in DRG neurons from TRPA1/TRPV1 double knockout mice, suggesting that production of endogenous levels of NO can activate these ion channels. In behavioral assays, peripheral NO-induced nociception is compromised when TRPV1 and TRPA1 are both ablated. These results provide genetic evidence that the peripheral nociceptive action of NO is mediated by both TRPV1 and TRPA1.

Synergistic anti-inflammatory effects of low doses of curcumin in combination with polyunsaturated fatty acids: Docosahexaenoic acid or eicosapentaenoic acid

Inflammatory response plays an important role not only in the normal physiology but also in the pathology such as cancers. As chronic inflammations are associated with malignancies, it is important to prevent inflammation-mediated neoplastic formation, promotion and/or progression. One possible intervention will be using cancer chemopreventive agents such as curcumin (CUR), a potent anti-inflammatory and anti-oxidative stress compound. Polyunsaturated fatty acids (PUFA) such as docosahexaenoic acid (DHA) or eicosapentaenoic acid (EPA) are potent anti-inflammatory agents by decreasing the production of inflammatory eicosanoids, cytokines, and reactive oxygen species (ROS). The present study aims at examining whether CUR with DHA or EPA would have synergistic anti-inflammatory effects in RAW 264.7 cells. Non-toxic concentrations of single and combination of the compounds were investigated at 6, 12 and 24h. The nitric oxide (NO) suppression effects were most prominent at 24h. All the combinations of CUR and DHA or EPA with lower concentrations of CUR 5μM and 25μM of DHA or EPA were found to have synergistic effects in suppressing LPS-stimulated NO and endogenous NO levels. Importantly, very low doses of CUR 2.5μM and DHA or EPA of 0.78μM could synergistically suppress the LPS-induced prostaglandin E2 (PGE2). The combinations were also found to suppress iNOS, COX-2, 5-lipoxygenase (5-LOX) and cPLA2 but induce HO-1. Taken together, the present study clearly shows the synergistic anti-inflammatory as well as anti-oxidative stress effects of CUR and PUFA.