Preserving Retinal Structure and Function with the Novel Nitroxide Antioxidant, DCTEIO.

Oxidative stress is a major contributor to progressive neurodegenerative disease and may be a key target for the development of novel preventative and therapeutic strategies. Nitroxides have been successfully utilised to study changes in redox status (biological probes) and modulate radical-induced oxidative stress. This study investigates the efficacy of DCTEIO (5,6-dicarboxy-1,1,3,3-tetraethyllisoindolin-2-yloxyl), a stable, kinetically-persistent, nitroxide-based antioxidant, as a retinal neuroprotectant. The preservation of retinal function following an acute ischaemic/reperfusion (I/R) insult in the presence of DCTEIO was quantified by electroretinography (ERG). Inflammatory responses in retinal glia were analysed by GFAP and IBA-1 immunohistochemistry, and retinal integrity assessed by histology. A nitroxide probe combined with flow cytometry provided a rapid technique to assess oxidative stress and the mitigation offered by antioxidant compounds in cultured 661W photoreceptor cells. DCTEIO protected the retina from I/R-induced damage, maintaining retinal function. Histological analysis showed preservation of retinal integrity with reduced disruption and disorganisation of the inner and outer nuclear layers. I/R injury upregulated GFAP expression, indicative of retinal stress, which was significantly blunted by DCTEIO. The number of 'activated' microglia, particularly in the outer retina, in response to cellular stress was also significantly reduced by DCTEIO, potentially suggesting reduced inflammasome activation and cell death. DCTEIO mitigated oxidative stress in 661W retinal cell cultures, in a dose-dependent fashion. Together these findings demonstrate the potential of DCTEIO as a neuroprotective therapeutic for degenerative diseases of the CNS that involve an ROS-mediated component, including those of the retina e.g. age-related macular degeneration and glaucoma.

Two polarity reversal modes lead to different nitrate reduction pathways in bioelectrochemical systems.

Polarity reversal is one of the effective strategies to rapidly start up denitrifying BESs，but the long-term performances of the denitrifying BESs operated under polarity reversal receive little attention. This study investigated the effects of periodic polarity reversal (PPR) and polarity reversal once only (PRO) on the long-term performances of denitrifying BESs. Repeatable oxidative and reductive currents were observed in the BESs obtained by PPR (PPR-BESs). The peak reductive currents of the PPR-BESs reached 0.95 A/m2, and nitrate was mainly removed by dissimilatory nitrate reduction to ammonium pathway with removal rates higher than 95 %. In contrast, the peak reductive currents of the BESs obtained by PRO (PRO-BESs) progressively decreased from 1.01 A/m2 to 0.12 A/m2. The nitrate removal rates of the PRO-BESs were <50 %, and the product of nitrate reduction turned to N2 instead of ammonium. 16S rDNA sequencing and metatranscriptomic analysis revealed that Geobacter capable of bidirectional extracellular electron transfer (EET) and Afipia capable of autotrophic growth were the dominant genera in the two types of BESs. Outer membrane cytochrome c and formate dehydrogenase were potentially involved in the cathodic electron uptake. These findings contribute to a better understanding of the EET mechanisms of electroautotrophic denitrifiers.

Coupling mixotrophic denitrification and electroactive anodic nitrification by nitrate addition for promoting current generation and nitrogen removal.

Nitrate promotes anodic denitrification and fasts organic matter removal in microbial fuel cells (MFCs). However, it suffers from poor total nitrogen (TN) removal and current recovery. In this study, some novel electroactive nitrifying/denitrifying bacteria (ENDB) were introduced in a single chambered air-cathode MFC to investigate the performance of this device and the microbial community shift by adding nitrate. Results showed a similar disturbance in current output by adding nitrate during a short-term operation. However, a stable and reproducible current increase was achieved in the continuous experiment. A maximum current of 0.76 A m-3 and a maximum TN removal of >99 % were accomplished. The corresponding corrected coulombic efficiency was approximately 18 %. Under repeatable batches, a sharp decrease in chemical oxygen demand (COD) with feeding nitrate confirmed the temporary competition on electron donors through heterotrophic denitrification. The later current increase and nitrite detection occurring without metabolized COD could be considered evidence of electroactive anodic nitrification. The ENDB biofilm successfully coupled mixotrophic denitrification and electroactive anodic nitrification. It eventually promoted TN removal. In the process, genera Pseudoxanthomonas, Thauera, and Pseudomonas were enriched in the anodic ENDB biofilms. Cyclic voltammetry data confirmed the promotion of the electron transfer process by biofilms. The bacterial function predication revealed that the genes related to nitrogen removal and electron transfer were upregulated. Therefore, mixotrophic denitrification and electroactive anodic nitrification processes facilitated power recovery with the high efficiency of pollutant removal, finally ensuring water body security.

Attenuation of hyperglycemia-associated dyslipidemic, oxidative, cognitive, and inflammatory crises via modulation of neuronal ChEs/NF-κB/COX-2/NOx, and hepatorenal functional deficits by the Tridax procumbens extract.

Tridax procumbens (cotton buttons) is a flowering plant with a medicinal reputation for treating infections, wounds, diabetes, and liver and kidney diseases. The present research was conducted to evaluate the possible protective effects of the T. procumbens methanolic extract (TPME) on an experimentally induced type 2 diabetes rat model. Wistar rats with streptozotocin (STZ)-induced diabetes were randomly allocated into five groups of five animals each, viz., a normal glycemic group (I), diabetic rats receiving distilled water group (II), diabetic rats with 150 (III) and 300 mg/kg of TPME (IV) groups, and diabetic rats with 100 mg/kg metformin group (V). All treatments were administered for 21 consecutive days through oral gavage. Results: Administration of the T. procumbens extract to diabetic rats significantly restored alterations in levels of fasting blood glucose (FBG), body weight loss, serum and pancreatic insulin levels, and pancreatic histology. Furthermore, T. procumbens significantly attenuated the dyslipidemia (increased cholesterol, low-density lipoprotein-cholesterol (LDL-C), triglycerides, and high-density lipoprotein (HDL) in diabetic rats), serum biochemical alterations (alanine transaminase (ALT), aspartate transaminase (AST), alanine phosphatase (ALP), blood urea nitrogen (BUN), creatinine, uric acid, and urea) and full blood count distortion in rats with STZ-induced diabetes. The TPME also improved the antioxidant status as evidenced by increased superoxide dismutase (SOD), catalase (CAT), glutathione (GSH), and decreased malondialdehyde (MDA); and decreased levels of cholinesterases (acetylcholinesterase (AChE) and butyrylcholinesterase (BChE)), and proinflammatory mediators including nuclear factor (NF)-κB, cyclooxygenase (COX)- 2, and nitrogen oxide (NOx) in the brain of rats with STZ-induced diabetes compared to rats with STZ-induced diabetes that received distilled water. However, TPME treatment failed to attenuate the elevated monoamine oxidases and decreased dopamine levels in the brain of rats with STZ-induced diabetes. Extract characterization by liquid chromatography mass spectrometry (LC-MS) identified isorhamnetin (retention time (RT)= 3.69 min, 8.8%), bixin (RT: 25.06 min, 4.72%), and lupeol (RT: 25.25 min, 2.88%) as the three most abundant bioactive compounds that could be responsible for the bioactivity of the plant. In conclusion, the TPME can be considered a promising alternative therapeutic option for managing diabetic complications owing to its antidiabetic, antihyperlipidemic, antioxidant, and anti-inflammatory effects in rats with STZ-prompted diabetes.

Effects of magnetite on microbially driven nitrate reduction processes in groundwater.

Nitrate is a common pollutant in the aquatic environment. Denitrification and dissimilatory nitrate reduction to ammonium (DNRA) are the main reduction processes of nitrate. In the relatively closed sediment environment, the competitive interaction of these two nitrate reduction determines whether the ecosystem removes or retains nitrogen. In the process of NO3--N bioreduction, Magnetite, which is a common mineral present in soil and other sediments can play a crucial role. However, it is still not clear whether magnetite promotes or inhibits NO3--N bioreduction. In this paper, the effect of magnetite on NO3--N bioreduction was studied by batch experiments. The results show that magnetite can increase the NO3--N reduction rate by 1.48 %, and can inhibit the DNRA process at the beginning of the reaction and then promote the DNRA process. Magnetite changed the microbial community structure in our experiment systems. The relative abundance of Sphingomonas, which mainly exists in a high carbon and low nitrogen environment, increased under sufficient carbon source conditions. The relative abundance of Fe-oxidizing and NO3--N reducing bacteria, such as Flavobacterium, increased in the absence of carbon sources but in the presence of magnetite. In addition, magnetite can significantly increase activity of the microbial electron transport system (ETS). the added microbial electronic activity of magnetite increased nearly two-fold under the same experiment conditions. The acid produced by the metabolisms of Pseudomonas and Acinetobacter further promotes the dissolution of magnetite, thus increasing the concentration of Fe (II) in the system, which is beneficial to autotrophic denitrifying bacteria and promote the reduction of NO3--N. These findings can enhance our understanding of the interaction mechanism between iron minerals and nitrate reducing bacteria during nitrate reduction under natural conditions.

DNIC is a Structure Providing Specificity of Physiological Effects of NO.

Metabolism of nitric oxide (NO) donors: dinitrosyl iron complexes (DNIC), nitrosothiols (RSNO), and nitroprusside was studied on a chick embryo model. The obtained results give reason to assume that DNIC constituting the main pool of nitroso compounds in the vast majority of tissues are NO donors immediately interacting with the physiological target of NO, and other NO donors can perform this function after their transformation into DNIC. NO is released from DNIC not spontaneously, but under a joint influence of a factor destroying the complex and a target having chemical affinity for NO. A similar mechanism is apparently implicated in NO passage through the cell membrane.

Effects of Combined Inorganic Nitrate and Nitrite Supplementation on Cardiorespiratory Fitness and Skeletal Muscle Oxidative Capacity in Type 2 Diabetes: A Pilot Randomized Controlled Trial.

Nitric oxide (NO) stimulates mitochondrial biogenesis in skeletal muscle. However, NO metabolism is disrupted in individuals with type 2 diabetes mellitus (T2DM) potentially contributing to their decreased cardiorespiratory fitness (i.e., VO2max) and skeletal muscle oxidative capacity. We used a randomized, double-blind, placebo-controlled, 8-week trial with beetroot juice containing nitrate (NO3−) and nitrite (NO2−) (250 mg and 20 mg/day) to test potential benefits on VO2max and skeletal muscle oxidative capacity in T2DM. T2DM (N = 36, Age = 59 ± 9 years; BMI = 31.9 ± 5.0 kg/m2) and age- and BMI-matched non-diabetic controls (N = 15, Age = 60 ± 9 years; BMI = 29.5 ± 4.6 kg/m2) were studied. Mitochondrial respiratory capacity was assessed in muscle biopsies from a subgroup of T2DM and controls (N = 19 and N = 10, respectively). At baseline, T2DM had higher plasma NO3− (100%; p < 0.001) and lower plasma NO2− levels (−46.8%; p < 0.0001) than controls. VO2max was lower in T2DM (−26.4%; p < 0.001), as was maximal carbohydrate- and fatty acid-supported oxygen consumption in permeabilized muscle fibers (−26.1% and −25.5%, respectively; p < 0.05). NO3−/NO2− supplementation increased VO2max (5.3%; p < 0.01). Further, circulating NO2−, but not NO3−, positively correlated with VO2max after supplementation (R2= 0.40; p < 0.05). Within the NO3−/NO2− group, 42% of subjects presented improvements in both carbohydrate- and fatty acid-supported oxygen consumption in skeletal muscle (vs. 0% in placebo; p < 0.05). VO2max improvements in these individuals tended to be larger than in the rest of the NO3−/NO2− group (1.21 ± 0.51 mL/(kg*min) vs. 0.31 ± 0.10 mL/(kg*min); p = 0.09). NO3−/NO2− supplementation increases VO2max in T2DM individuals and improvements in skeletal muscle oxidative capacity appear to occur in those with more pronounced increases in VO2max.

Qingda granule alleviate angiotensin â ±-induced hypertensive renal injury by suppressing oxidative stress and inflammation through NOX1 and NF-κB pathways.

Hypertension has become one of the important diseases harmful to human health. In China, Qingda granule (QDG) has been used to treat hypertension for decades. Previous studies by our team have shown that oxidative stress may be one of the pathways through which QDG inhibits hypertension-induced organs injury. However, the specific molecular mechanism of its anti-hypotension and renal oxidative stress response were unclearly. This study investigated QDG's potential protective mechanism against hypertension-induced renal injury. Mice were infused with Angiotensin Ⅱ (Ang Ⅱ, 500 ng/kg/min) or equivalent saline solution (Control) and administered oral QDG (1.145 g/kg/day) or saline for four weeks. QDG treatment mitigated the elevated blood pressure and reduced renal pathological changes induced by Ang Ⅱ. As per the RNA sequencing results, QDG affects oxidative stress signaling. In agreement with these findings, QDG significantly attenuated the Ang Ⅱ-induced increase in Nitrogen oxides 1 (NOX1) and reactive oxygen species and the decrease in superoxide dismutase in renal tissue. Additionally, QDG significantly inhibited Interleukin 6 (IL-6), Tumor necrosis factor α (TNF-α), and Interleukin 1ß (IL-1ß) expression in renal tissues and blocked the phosphorylation of P65 (NF-κB subunit) and IκB. These results were confirmed in vitro. Overall, QDG reduced Ang Ⅱ-induced elevated blood pressure and renal injury by inhibiting oxidative stress and inflammation caused by NOX1 and NF-κB pathways. The results of this study provide an experimental basis for the clinical application of QDG, and to open up a new direction for the clinical treatment of hypertension.

Nitric Oxide, Nitric Oxide Formers and Their Physiological Impacts in Bacteria.

Nitric oxide (NO) is an active and critical nitrogen oxide in the microbe-driven nitrogen biogeochemical cycle, and is of great interest to medicine and the biological sciences. As a gas molecule prior to oxygen, NO respiration represents an early form of energy generation via various reactions in prokaryotes. Major enzymes for endogenous NO formation known to date include two types of nitrite reductases in denitrification, hydroxylamine oxidoreductase in ammonia oxidation, and NO synthases (NOSs). While the former two play critical roles in shaping electron transport pathways in bacteria, NOSs are intracellular enzymes catalyzing metabolism of certain amino acids and have been extensively studied in mammals. NO interacts with numerous cellular targets, most of which are redox-active proteins. Doing so, NO plays harmful and beneficial roles by affecting diverse biological processes within bacterial physiology. Here, we discuss recent advances in the field, including NO-forming enzymes, the molecular mechanisms by which these enzymes function, physiological roles of bacterial NOSs, and regulation of NO homeostasis in bacteria.

Mass transfer characteristics and effect of flue gas used in microalgae culture.

Flue gas not only contains carbon dioxide (CO2) but also air pollutants (sulfur oxides (SOx) and nitrogen oxides (NOx)). The effective utilization of flue gas could help us to reduce the cost of microalgal biomass production. This study assessed and explored the utilization of flue gas for the absorption characteristics of different components and their biological effect in microalgal culture systems. In abiotic absorption experiments, the absorptivity of CO2 was reduced by a maximum of 3.1%, and the concentration of the available carbon source in the culture medium was decreased by 6.7% when sulfur dioxide (SO2, at 100 mg/m3) was presented in the flue gas. Meanwhile, the presence of oxygen (O2, at 4%) in the flue gas improved the absorptivity of nitric oxide (NO). When Scenedesmus dimorphus was cultured using bisulfites and nitrites (at 10 mmol/L and 8 mmol/L, respectively) as the sulfur and nitrogen sources, SOx and NOx in the flue gas did not significantly affect growth of microalgal cells and the carbohydrate, lipid, and protein content. The consumption rates of nutrient elements were calculated, which could provide an adjustment strategy for the initial gas source when culturing microalgae with the flue gas. This study indicates that the flue gas used for microalgal culture should be partially desulfurized, so that the SOx and CO2 concentrations can optimize growth of microalgal cells, while the denitrification might not be needed since the flue gas can be oxidized to utilize the NO. KEY POINTS: • The concentration of the available carbon source in the culture medium was decreased when SO2 was presented in the flue gas, and the presence of O2 in the flue gas improved the absorptivity of NO. • An adjustment strategy for the initial gas source when culturing microalgae with the flue gas was firstly proposed. • For flue gas containing 10% CO2 and 60 mg/m3 of SO2, growth of Scenedesmus dimorphus showed no difference in cell growth in normal culture conditions.

Ethylene Acts as a Local and Systemic Signal to Mediate UV-B-Induced Nitrate Reallocation to Arabidopsis Leaves and Roots via Regulating the ERFs-NRT1.8 Signaling Module.

Nitrate is the preferred nitrogen source for plants and plays an important role in plant growth and development. Under various soil stresses, plants reallocate nitrate to roots to promote stress tolerance through the ethylene-ethylene response factors (ERFs)-nitrate transporter (NRT) signaling module. As a light signal, ultraviolet B (UV-B) also stimulates the production of ethylene. However, whether UV-B regulates nitrate reallocation in plants via ethylene remains unknown. Here, we found that UV-B-induced expression of ERF1B, ORA59, ERF104, and NRT1.8 in both Arabidopsis shoots and roots as well as nitrate reallocation from hypocotyls to leaves and roots were impaired in ethylene signaling mutants for Ethylene Insensitive2 (EIN2) and EIN3. UV-B-induced NRT1.8 expression and nitrate reallocation to leaves and roots were also inhibited in the triple mutants for ERF1B, ORA59, and ERF104. Deletion of NRT1.8 impaired UV-B-induced nitrate reallocation to both leaves and roots. Furthermore, UV-B promoted ethylene release in both shoots and roots by enhancing the gene expression and enzymatic activities of ethylene biosynthetic enzymes only in shoots. These results show that ethylene acts as a local and systemic signal to mediate UV-B-induced nitrate reallocation from Arabidopsis hypocotyls to both leaves and roots via regulating the gene expression of the ERFs-NRT1.8 signaling module.

Organic carbon quantity and composition jointly modulate the differentiation of nitrate reduction pathways in sediments of the Chinese eutrophic lake, Lake Chaohu.

Twelve sampling sites from two basins of Lake Chaohu were studied seasonally from June 2020 to April 2021 in Hefei City (China) to better understand the effect of organic carbon (C) quantity and composition on nitrate (NO3--N) reduction pathways. Serious algal bloom in the west basin of Lake Chaohu (WLC) resulted in higher organic C accumulation and NO3--N deficiency in interstitial water compared to the east basin of Lake Chaohu (ELC), jointly leading to a high C/NO3--N ratio. This triggered dissimilatory nitrate reduction to ammonium (DNRA) over denitrification in terms of higher DNRA rate, nitrogen retaining index (NRI), and nrfA gene abundance mediating DNRA. Furthermore, high oxygen-alkyl C and abundance of functional genes mediating labile organic C decomposition and DNRA suggested that the alkyl carbon-oxygen bond was responsible for DNRA induction. Different bacterial community composition and diversity involved in C and nitrogen (N) metabolism in two basins indicated that bacteria in sediments of WLC were more active in NO3--N reduction. Spearman correlation analysis showed that the less represented genera, such as Thiobacillus and Clostridium, were positively correlated with both organic C and NO3--N reduction rates, respectively. Hence, organic C composition could affect NO3--N reduction function by shaping the specific bacterial community.

The oral microbiome, nitric oxide and exercise performance.

The human microbiome comprises ∼1013-1014 microbial cells which form a symbiotic relationship with the host and play a critical role in the regulation of human metabolism. In the oral cavity, several species of bacteria are capable of reducing nitrate to nitrite; a key precursor of the signaling molecule nitric oxide. Nitric oxide has myriad physiological functions, which include the maintenance of cardiovascular homeostasis and the regulation of acute and chronic responses to exercise. This article provides a brief narrative review of the research that has explored how diversity and plasticity of the oral microbiome influences nitric oxide bioavailability and related physiological outcomes. There is unequivocal evidence that dysbiosis (e.g. through disease) or disruption (e.g. by use of antiseptic mouthwash or antibiotics) of the oral microbiota will suppress nitric oxide production via the nitrate-nitrite-nitric oxide pathway and negatively impact blood pressure. Conversely, there is preliminary evidence to suggest that proliferation of nitrate-reducing bacteria via the diet or targeted probiotics can augment nitric oxide production and improve markers of oral health. Despite this, it is yet to be established whether purposefully altering the oral microbiome can have a meaningful impact on exercise performance. Future research should determine whether alterations to the composition and metabolic activity of bacteria in the mouth influence the acute responses to exercise and the physiological adaptations to exercise training.

Electrodialysis ion-exchange membrane bioreactor (EDIMB) to remove nitrate from water: Optimization of operating conditions and kinetics analysis.

Nitrate pollution has become a worldwide problem. In this study, we remove nitrate from water by electrodialysis ion-exchange membrane bioreactor (EDIMB) and enabling simultaneous nitrate enrichment and denitrification. In this reactor, nitrate migrated from the water chamber to the biological chamber via electrodialysis and was degraded by microorganisms. The effects of voltage and biomass concentration on the reactor performance were examined, and the kinetics data of the water chamber and biological chamber were fitted. The experimental results showed that the migration of nitrate in the water chamber conformed to the first-order model, and the constructed zero-Michaelis-Menten model described changes in nitrate concentration in the biological chamber. Furthermore, when the inflow nitrate concentration was 40 mg N/L, 5 V was the best voltage, and 3.00 g VSS/L was the best biomass concentration. The nitrate removal rate in the water chamber was 98.94%, and there was no accumulation of nitrate or nitrite in the biological chamber. Compared with traditional ED processes, the nitrate removal efficiency was 8.86% higher, and the current efficiency was 22.14% higher. The total organic carbon (TOC) of the water chamber was only 1.43 mg C/L, which proves that the structure of the EDIMB confined the denitrifying bacteria and organic carbon donors in the biological chamber and avoided secondary pollution in the water chamber. Microbial community analysis showed that Thauera (66.06%) was the dominant bacterium in the EDIMB system, and Azoarcus (9.81%) was a minor denitrifying genus.

Nitrate respiration occurs throughout the depth of mucoid and non-mucoid Pseudomonas aeruginosa submerged agar colony biofilms including the oxic zone.

Pseudomonas aeruginosa is an opportunistic pathogen and well characterized biofilm former. P. aeruginosa forms strong oxygen gradients inside biofilms due to rapid oxygen respiration in the top layers and the poor solubility of oxygen coupled with diffusion limited transport. Transcriptomic evidence from in vitro and ex vivo sampling suggests that denitrification is occurring in biofilms in ostensibly oxic environments. It is hypothesized that in the presence of nitrate there is stratification with aerobic respiration occurring in the outer oxic layer and denitrification in the lower anoxic zone. We used submerged agar colony biofilms grown from mucoid (FRD1) and non-mucoid (PAO1) strains to simultaneously measure depth microprofiles of oxygen and nitrous oxide in the same colony with microelectrodes. Oxygen respiration occurred at the top of the colony as expected but denitrification occurred throughout the entire depth, even in the oxic region. Local denitrification rates were highly variable suggesting heterogenous metabolic activity within the colony. We also assessed the short-term influence of tobramycin on aerobic respiration within a PAO1 colony. Although there was an immediate reduction in respiration it was never completely arrested over a 2 h period. On tobramycin removal the oxygen gradient steadily reestablished, demonstrating immediate recovery of metabolic activity.

The mechanism by which Enteromorpha Linza polysaccharide promotes Bacillus subtilis growth and nitrate removal.

In this study, we discussed the relationship between Entermorpha linza polysaccharide (EP) and Bacillus subtilis, which can transform nitrate. A sole carbon source experiment showed that Bacillus subtilis could utilize EP, and the bacterial density was maximally increased by 54.43% in the EP groups. The results of reducing sugar determination proved the secretion of polysaccharide-degrading enzymes. Scanning electron microscopy (SEM) showed that the EP groups had fewer spores and shrunken bacteria, indicating that EP could improve the growth environment and maintain bacterial integrity. Additionally, the ratios of periplasmic nitrate reductase (NAP), nitrite reductase (NIR), and dissimilatory nitrate reductase (D-NRase) in the EP groups were maximally increased by 107.22%, 84.70% and 36.10%, respectively. Transcriptome analysis further confirmed the above mentioned results. For example, the high expression of quorum sensing genes indicated that EP groups had higher bacterial density. Moreover, the high expression of antioxidant genes in the EP groups may be related to morphological integrity. Our study provides a basis for further discussion of the mechanism.

Effect of Copper on Expression of Functional Genes and Proteins Associated with Bradyrhizobium diazoefficiens Denitrification.

Nitrous oxide (N2O) is a powerful greenhouse gas that contributes to climate change. Denitrification is one of the largest sources of N2O in soils. The soybean endosymbiont Bradyrhizobium diazoefficiens is a model for rhizobial denitrification studies since, in addition to fixing N2, it has the ability to grow anaerobically under free-living conditions by reducing nitrate from the medium through the complete denitrification pathway. This bacterium contains a periplasmic nitrate reductase (Nap), a copper (Cu)-containing nitrite reductase (NirK), a c-type nitric oxide reductase (cNor), and a Cu-dependent nitrous oxide reductase (Nos) encoded by the napEDABC, nirK, norCBQD and nosRZDFYLX genes, respectively. In this work, an integrated study of the role of Cu in B. diazoefficiens denitrification has been performed. A notable reduction in nirK, nor, and nos gene expression observed under Cu limitation was correlated with a significant decrease in NirK, NorC and NosZ protein levels and activities. Meanwhile, nap expression was not affected by Cu, but a remarkable depletion in Nap activity was found, presumably due to an inhibitory effect of nitrite accumulated under Cu-limiting conditions. Interestingly, a post-transcriptional regulation by increasing Nap and NirK activities, as well as NorC and NosZ protein levels, was observed in response to high Cu. Our results demonstrate, for the first time, the role of Cu in transcriptional and post-transcriptional control of B. diazoefficiens denitrification. Thus, this study will contribute by proposing useful strategies for reducing N2O emissions from agricultural soils.

Root system size and root hair length are key phenes for nitrate acquisition and biomass production across natural variation in Arabidopsis.

The role of root phenes in nitrogen (N) acquisition and biomass production was evaluated in 10 contrasting natural accessions of Arabidopsis thaliana L. Seedlings were grown on vertical agar plates with two different nitrate supplies. The low N treatment increased the root to shoot biomass ratio and promoted the proliferation of lateral roots and root hairs. The cost of a larger root system did not impact shoot biomass. Greater biomass production could be achieved through increased root length or through specific root hair characteristics. A greater number of root hairs may provide a low-resistance pathway under elevated N conditions, while root hair length may enhance root zone exploration under low N conditions. The variability of N uptake and the expression levels of genes encoding nitrate transporters were measured. A positive correlation was found between root system size and high-affinity nitrate uptake, emphasizing the benefits of an exploratory root organ in N acquisition. The expression levels of NRT1.2/NPF4.6, NRT2.2, and NRT1.5/NPF7.3 negatively correlated with some root morphological traits. Such basic knowledge in Arabidopsis demonstrates the importance of root phenes to improve N acquisition and paves the way to design eudicot ideotypes.

Photolytic Measurement of Tissue S-Nitrosothiols in Rats and Humans In Vivo.

S-nitrosothiols are labile thiol-NO adducts formed in vivo primarily by metalloproteins such as NO synthase, ceruloplasmin, and hemoglobin. Abnormal S-nitrosothiol synthesis and catabolism contribute to many diseases, ranging from asthma to septic shock. Current methods for quantifying S-nitrosothiols in vivo are suboptimal. Samples need to be removed from the body for analysis, and the S-nitrosothiols can be broken down during ex vivo processing. Here, we have developed a noninvasive device to measure mammalian tissue S-nitrosothiols in situ non-invasively using ultraviolet (UV) light, which causes NO release in proportion to the S-nitrosothiol concentration. We validated the assay in vitro; then, we applied it to measure S-nitrosothiols in vivo in rats and in humans. The method was sensitive to 0.5 µM, specific (did not detect other nitrogen oxides), and was reproducible in rats and in humans. This noninvasive approach to S-nitrosothiol measurements may be applicable for use in human diseases.

Reactions of Recombinant Neuronal Nitric Oxide Synthase with Redox Cycling Xenobiotics: A Mechanistic Study.

Neuronal nitric oxide synthase (nNOS) catalyzes single-electron reduction of quinones (Q), nitroaromatic compounds (ArNO2) and aromatic N-oxides (ArN → O), and is partly responsible for their oxidative stress-type cytotoxicity. In order to expand a limited knowledge on the enzymatic mechanisms of these processes, we aimed to disclose the specific features of nNOS in the reduction of such xenobiotics. In the absence or presence of calmodulin (CAM), the reactivity of Q and ArN → O increases with their single-electron reduction midpoint potential (E17). ArNO2 form a series with lower reactivity. The calculations according to an "outer-sphere" electron transfer model show that the binding of CAM decreases the electron transfer distance from FMNH2 to quinone by 1-2 Å. The effects of ionic strength point to the interaction of oxidants with a negatively charged protein domain close to FMN, and to an increase in accessibility of the active center induced by high ionic strength. The multiple turnover experiments of nNOS show that, in parallel with reduced FAD-FMN, duroquinone reoxidizes the reduced heme, in particular its Fe2+-NO form. This finding may help to design the heme-targeted bioreductively activated agents and contribute to the understanding of the role of P-450-type heme proteins in the bioreduction of quinones and other prooxidant xenobiotics.