Unlocking the interactive role of GABA and NO in alleviating chlorpyrifos induced toxicity in Solanum melongena L. through modulation of physio-biochemical traits.

Targeted therapeutic delivery for treating bacterial infections remains underutilized in most pharmaceutical interventions. Existing therapeutics (i.e., antibiotics) are often systematically administered despite the presence of localized infection, leading to both off-target toxicity and suboptimal bacterial clearance with limited efficacy against biofilms. The overuse of antibiotics has resulted in increased antimicrobial resistance, creating a need for alternative interventions that are unlikely to confer resistance. Nitric oxide (NO), an endogenous mediator produced by macrophages and other immune cells in response to infection, elicits broad spectrum antibacterial and antibiofilm activity. The use of exogenous NO donors, alone or as conjugated ligands to macromolecular scaffolds, has proven effective in treating anatomical targets, including dermal wounds, dental infections, and pulmonary conditions, in a localized manner. In this perspective, we provide an overview of the recent advancements in NO-releasing biomaterials, highlighting design strategy and antimicrobial action across diverse anatomical sites.

The inhibitory effect of exogenous nitric oxide (NO) on the open state of ATP-sensitive potassium channels (K ATP) and its underlying mechanism remain unclear. In this study, patch-clamp and molecular biology techniques are used to investigate this issue. In acutely isolated rat mesenteric artery smooth muscle cells and human embryonic kidney 293 cells (HEK293) expressing inwardly rectifying potassium channel 6.1 subunit/sulfonylurea receptor 2B subunit (Kir6.1/SUR2B), sodium nitroprusside (SNP) was found to significantly inhibit the activity of open K ATP channels. Detection using biotin-labeled glutathione ethyl ester (BioGEE) combined with Western blotting showed that Kir6.1 subunit glutathionylation level was significantly decreased after SNP treatment. These results indicate that exogenous NO directly inhibits the activity of open K ATP channels by nitrosylating key cysteine residues of the Kir6.1 subunit and competitively inhibiting glutathionylation at this site. This study provides new experimental evidence for the molecular mechanism of NO in vascular regulation.

Iron (Fe) deficiency is a widespread constraint on crop productivity, particularly in calcareous soils where elevated bicarbonate levels impair Fe uptake and utilization. Nitric oxide (NO) is a key regulator of Fe homeostasis; however, the physiological effects of exogenous NO donors such as sodium nitroprusside may reflect combined actions of NO and associated by-products. This study evaluated the effects of SNP on M7 apple plants (Malus domestica) under direct low-iron stress (45 and 2µM Fe) and bicarbonate-induced Fe deficiency in a controlled substrate-based greenhouse system. Low Fe availability significantly reduced growth, chlorophyll content, photosynthetic rate, and leaf active Fe concentration. SNP supplementation, particularly at 200µM, partially alleviated these constraints and enhanced ferric chelate reductase activity together with the expression of Fe-acquisition genes (FIT, FRO2-like, and IRT1). Notably, sustained activation of these genes despite partial recovery of leaf active Fe indicates that NO reinforces Strategy I Fe-deficiency signaling rather than fully restoring Fe nutritional status. Under bicarbonate stress, these responses persisted, suggesting additional physiological limitations beyond rhizosphere alkalinization. Although soils can emit NO through microbial processes, its bioavailability is variable and may not consistently sustain Fe acquisition under severe Fe limitation. Therefore, exogenous NO application under controlled conditions provides mechanistic insight into NO-mediated regulation of Fe homeostasis. These findings represent an integrated physiological response to SNP treatment and require validation across genotypes and field environments to confirm NO-specific effects and practical relevance.

Maize (Zea mays L.) is one of the major grain crops worldwide that is particularly vulnerable to waterlogging (WL) stress. Glutathione (GSH) and nitric oxide (NO) are known to protect plants from a variety of abiotic stresses; however, their potential for improving WL tolerance in maize remains unexplored. The present study examined the impact of exogenously applied GSH and NO on maize plants exposed to WL-stress. Our findings revealed that GSH + NO-treated waterlogged maize plants grew better and produced more biomass than only WL-stressed plants. The improved performance of GSH + NO-sprayed WL-stressed maize seedlings was supported by the increased root dry and fresh weight, shoot length, shoot dry and fresh weight, chlorophyll a, chlorophyll b, and carotenoid content. Exogenous GSH and NO treatments significantly enhanced the amounts of leaf proline, leaf soluble sugars, and total protein in maize seedlings, suggesting adaptive metabolic reprogramming under stress. The increased malondialdehyde (MDA) levels and accumulation of hydrogen peroxide (H2O2) in maize leaves and roots revealed that WL caused significant oxidative damage. Exogenous GSH, NO individually, and combinedly significantly decreased total H2O2 and MDA contents in both leaves and roots. Exogenous GSH and NO reduced oxidative stress by increasing peroxidase activity, ascorbic acid, and anthocyanin content in maize leaf and root tissues. Our findings emphasize the possible relevance of GSH and NO, simultaneously and individually, in enhancing adaptive mechanisms in maize seedlings for reducing WL-induced damage. Although the GSH + NO-mediated approach shows promise for mitigating WL-stress in maize under controlled conditions, further field-based investigations are required to validate its practical applicability.

Bidirectional coupling of neuronal Ca2+ and nitric oxide signals visualized by a dual biosensor.

Nitric oxide (NO), a gaseous signaling molecule, is increasingly recognized for its central role in the pathophysiology of dental and oral diseases. Dysregulation of nitric oxide synthase (NOS) and consequent impairment of endogenous NO production have been implicated in a range of oral conditions. Accordingly, exogenous NO therapy has garnered significant interest due to its broad-spectrum antibacterial, potent anti-inflammatory, tissue-regenerative, and potential antitumor effects. This review provides a comprehensive overview of therapeutic advances in exogenous NO for dental and oral diseases, focusing on underlying mechanisms, such as modulation of the cyclic di-guanosine monophosphate pathway, mediation of S-nitrosylation, activation of the soluble guanylate cyclase-cyclic guanosine monophosphate-protein kinase G pathway, and regulation of the c-Jun N-terminal kinase/mitogen-activated protein kinase pathway. Collectively, these mechanisms confer antimicrobial efficacy, immunomodulation, enhanced periodontal and pulpal tissue repair, and apoptosis induction in oral cancer cells. Various NO delivery platforms, including nanocarriers, hydrogels, implant coatings, and microneedles, are reviewed for their efficacy and application in managing periodontitis, peri-implantitis, dental caries, endodontic and periapical diseases, oral ulcers, orthodontic tooth movement, and oral cancer. Although most of the current evidence is derived from preclinical studies and substantial challenges remain with respect to dosing accuracy and delivery specificity, the unique multitarget properties of exogenous NO highlight its considerable promise as an adjunctive strategy in dental medicine. This review seeks to provide a comprehensive overview that not only advances fundamental understanding of exogenous NO but also promotes its clinical translation for improved management of dental and oral diseases.

Alfalfa (Medicago sativa L.) is a globally important forage legume and the most widely cultivated sown pasture species in China. Drought, as one of the most common abiotic stresses, limits alfalfa growth and development. Nitric oxide (NO), a key signaling molecule, plays an essential role in plant growth, development, and responses to various abiotic stresses. In this study, exogenous NO was applied to alfalfa seedlings under drought stress, followed by physiological and transcriptomic analyses. The results showed that sodium nitroprusside (SNP)-treated alfalfa seedlings grew better than untreated controls (CK), with improved leaf tissue structure. Meanwhile, SNP treatment increased proline content, reduced malondialdehyde accumulation, and enhanced hydroxyl radical scavenging capacity. Under drought stress, lignin content increased in alfalfa seedlings. Following exogenous NO application, lignin content in alfalfa seedlings further increased. RNA-Seq analysis identified 20,183 differentially expressed genes (DEGs) in alfalfa seedlings treated with PEG, SNP, or PEG + SNP. KEGG enrichment analysis indicated that the DEGs under drought stress were involved in the phenylpropanoid biosynthesis pathway, which regulates lignin biosynthesis and enhances drought tolerance. GO enrichment analysis revealed that these DEGs participated in the response to water deprivation, thereby modulating drought stress tolerance and improving drought resistance. Furthermore, we assessed the transcript-level changes in genes induced by phenylpropanoid biosynthesis in alfalfa. Among them, 124 DEGs were identified as participating in phenylpropanoid biosynthesis, including 10 up-regulated DEGs (three of which encode key enzymes associated with lignin synthesis), while the remaining DEGs were down-regulated. These findings provide new insights into the transcriptomic mechanisms of SNP-mediated drought adaptation in alfalfa and reveal key pathways contributing to drought tolerance in this species.

In cardiac pathologies, levels of G protein-coupled receptor kinase 2 (GRK2)-which is involved in receptor desensitization and internalization-are elevated. Beyond these receptor-mediated effects, GRK2 also localizes to mitochondria, where it contributes to pathology. GRK2's activity can be inhibited via S-nitrosylation at Cysteine 340, a post-translational modification mediated by both endogenous and exogenous nitric oxide. Thus, S-nitrosylation is considered as an endogenous brake on GRK2's catalytic activity, counteracting its hyperactivity observed in disease states. However, it remains unclear whether S-nitrosylation also regulates GRK2's influence on mitochondrial function. This study aims to investigate how S-nitrosylation regulates mitochondrial localization and function of GRK2 under hypoxia/reoxygenation stress. To prevent S-nitrosylation at Cys340, we infected AC16 cardiac cells with adenoviruses carrying a GRK2 C340S (Ser) mutation. Our results indicate that inhibiting S-nitrosylation enhances mitochondrial localization of GRK2, especially in response to pathological stimuli. Additionally, mitochondrial function was impaired, as measured by oxygen consumption rates at ATP production. Furthermore, alterations in mitochondrial dynamics and mitophagy led to adverse outcomes when GRK2 was not subject to S-nitrosylation, presumably due to increased catalytic activity. Our findings underscore the importance of GRK2 regulation in cardiac pathologies and suggest that targeting GRK2 or its post-translational modifications may provide therapeutic benefits.

A chemiresistive nitric oxide (NO) gas sensor based on Pt/WO3 co-decorated carbon nanofibers (CNFs) was fabricated using a simple and scalable electrospinning process. This sensor demonstrates high-ppb-level NO detection at room temperature (25 °C), with an experimentally demonstrated detection limit of 100 ppb. It exhibits rapid response, good signal repeatability, excellent batch-to-batch reproducibility, and high selectivity toward NO. Compared with previously reported NO sensors, this work highlights the integration of Pt and WO3 within a conductive CNF network, enabling room-temperature NO detection down to 100 ppb using a simple chemiresistive architecture. In addition, preliminary sensing tests were conducted using dried simulated breath samples prepared by introducing exogenous NO into exhaled breath from healthy volunteers, demonstrating the sensor’s capability to resolve different NO levels in a complex breath-related background. Owing to its reliable performance and cost-effective fabrication, the sensor holds potential as a NO sensing platform, providing a materials-level basis for future breath NO analysis and other related applications.

Over 50% of global arable soils are acidic; acidic soil-induced aluminum (Al) phytotoxicity primarily inhibits root elongation, thereby reducing the plant's absorption of water and nutrients, which suppresses crop yield. Gibberellic acid (GA), a class of critical plant hormones, acts as a core regulator of plant development and growth mechanisms and contributes to the physiological adaptation of plants under stress conditions. In this study, we selected the rice variety Nipponbare (Nip) to investigate whether GA exerts an effect on alleviating Al toxicity and to explore the underlying mechanisms in rice. This study shows that Al stress quickly elevated the endogenous GA content in rice root tissues, consequently alleviating Al-induced suppression of root development. Exogenous GA application increased the expression of the Oryza sativa Cysteine-rich Peptide (Peptide with Cysteine-rich TDIF motif) 3 (OsCDT3) and Oryza sativa Ferric Reductase Defective Like 4 (OsFRDL4) genes, which reduce the toxicity of Al to plants and conversely decreased the expression of the Oryza sativa Natural Resistance-Associated Macrophage Protein 1 for Aluminum Transport (OsNRAT1) gene, which transports Al ions from the extracellular environment to the intracellular space. Furthermore, exogenous GA treatment modified the hemicellulose and pectin levels, therefore decreasing the absorption of Al. Further research shows that GA reduced the endogenous nitric oxide (NO) levels; nevertheless, the application of an exogenous nitric oxide donor sodium nitroprusside (SNP) offset the alleviatory role of GA. In conclusion, GA accelerated the cell wall Al exclusion mechanism, probably improving rice tolerance to Al toxicity via regulating the accumulation of NO.

Construction of a biocompatible supramolecular sensor for fluorescence imaging of Nitric oxide in plant tissues.

Introduction: The risk of cerebral vascular disease increases with age and after menopause in women. Although estrogen has positive cardiovascular effects, hormone therapy increases the risk of stroke. Importantly, our mechanistic understanding of the cerebrovascular connections between age, menopause, and estrogen replacement remains incomplete. The purpose of this study was to determine the effects of age and estrogen status on endothelium-dependent and -independent vasodilation in cerebral resistance arteries. Methods: Posterior communicating arteries (PCoA) were isolated from young, middle-aged, and old female Fisher-344 rats which were either gonadally intact, ovariectomized (OVX), or ovariectomized with estrogen replacement. Endothelium-dependent vasodilation was evaluated by assessing flow-induced vasodilation (FID) under normal, nitric oxide synthase (NOS) blockade (L-NAME), and cyclooxygenase activity inhibition conditions. Endothelium-independent vasodilation was evaluated by assessing responsiveness to an exogenous nitric oxide donor. Results: In intact rats, FID was decreased in PCoAs from old rats in comparison to those from young and middle-aged rats. This age-related deficit in FID was not influenced by OVX or L-NAME, but was further reduced when either estrogen therapy or L-NAME was given after OVX in old rats. FID was reduced by L-NAME in PCoAs from young intact rats. OVX also reduced FID in PCoAs from young rats, but L-NAME had no effect on FID after OVX. Estrogen therapy partially rescued FID in PCoA of young OVX rats only while NOS was active. In middle-aged rats, FID was reduced by OVX and not rescued by estrogen therapy. Similarly, FID in PCoAs from both intact and OXV middle-aged rats was reduced by L-NAME and not rescued by estrogen therapy. COX-mediated endothelium-dependent vasodilation and endothelium-independent vasodilation were not different in PCoAs across age and estrogen status conditions. Conclusions: We report age-specific effects of OVX and estrogen replacement on cerebrovascular function. NOS contributed to endothelium-dependent FID in PCoAs from both young and middle-aged, but not old rats. Estrogen therapy reversed OVX-induced reductions in a mechanism dependent on NOS in young rats. In old OVX rats, estrogen replacement exacerbated the age-related impairment of FID. These findings give mechanistic insight into the risk of stroke in women imposed by older age, menopause, and hormone replacement.

Peripheral artery disease (PAD) remains a great threat to the health of older people globally. Nitric oxide (NO), as an important signaling molecule, is integral to processes such as angiogenesis, inflammation, and tissue regeneration, making it a potential candidate for PAD treatment. Nevertheless, NO-based therapies are frequently limited in clinical utility, primarily due to the lack of effective strategies for fine-tuning the release of exogenous NO. In this study, we developed an enzyme-prodrug pair based on endocellulase (Cel5A-h38), which ensured complete bioorthogonality, thus avoiding interference with endogenous enzymes and eliciting an inflammatory response. This delivery system enables localized and controlled NO release, thus preventing side effects induced by systemic exposure. The therapeutic efficacy of the NO delivery system was systematically evaluated in a porcine model of hindlimb ischemia. Our results confirmed the benefits of targeted NO delivery in hindlimb ischemia, which include enhanced neovascularization and tissue perfusion, reduced inflammation, and alleviated muscle fibrosis, demonstrating its optimal translational potential.

Abstract Elucidating the physiological and biochemical responses of ornamental plants to drought stress and nitric oxide-mediated amelioration strategies is essential for developing sustainable production systems under climate change scenarios. However, comprehensive assessments of sodium nitroprusside (SNP) effects on morpho-physiological traits, oxidative stress responses, and metabolic pathways in drought-stressed ornamentals remain limited. We evaluated the effects of foliar SNP applications at 0, 50, 100, and 150 µM on Calendula officinalis L. grown under three irrigation regimes: 100%, 70%, and 40% of field capacity (FC), followed by assessment of morphological parameters, water relations, photosynthetic pigments, oxidative stress markers, antioxidant enzyme activities, osmoprotectant accumulation, and GABA metabolism. Stress treatment and SNP concentration significantly influenced all measured parameters ( p ≤ 0.01). Particularly, 100 µM SNP consistently demonstrated superior ameliorative effects, maintaining higher plant height, shoot biomass, leaf area, and flower production under both well-watered and drought conditions. Under severe drought (40% FC), 150 µM SNP reduced RWC decline from 24.6% to 11.2% and effectively mitigated reductions in total chlorophyll (63.1%) and carotenoids (55.3%) compared to untreated controls. Moreover, SNP enhanced proline accumulation up to 3-fold and substantially reduced oxidative stress markers (H₂O₂, MDA, and EL) by up to 50%. Although drought stress upregulated antioxidant enzymes (CAT, APX, POD, and SOD) by 2.2–4.3 times, SNP application moderated these elevations, suggesting reduced oxidative burden. GABA metabolism also responded markedly, with SNP enhancing glutamate decarboxylase (GAD) activity and GABA content to 0.68 U mg⁻¹ protein and 2.47 µmol g⁻¹ FW, respectively, while downregulating GABA transaminase (GABA-T) activity to 0.13 U mg⁻¹ protein. Principal component analysis revealed that 96% of total trait variance was explained by the first two components, clearly separating treatments by stress level and SNP concentration. We conclude that SNP significantly enhances drought tolerance in pot marigold through coordinated modulation of water relations, antioxidant systems, and GABA shunt pathway activation. The consistent superiority of specific SNP concentrations across multiple functional traits provides insights for optimizing foliar bio-stimulant applications, while the quantitative response patterns identified offer valuable parameters for developing stress-resilient ornamental production systems under changing climatic conditions.

Preparation of a Golgi-targetable and photo-triggered NO donor and its application in living cells and zebrafish imaging.

Radix Scutellariae, a crucial botanical drug widely used in Asian countries, is derived from the roots of Scutellaria baicalensis Georgi and exhibits diverse pharmacological activities, including antipyretic, analgesic, anti-inflammatory, antibacterial, antiviral, sedative, hypotensive, hypolipidemic, hepatoprotective, and choleretic effects. Owing to excessive market demand, the wild resources are insufficient, and cultivated Radix Scutellariae has thus become the primary source; however, this shift has resulted in a decline in medicinal quality. To address this quality issue, the present study proposes the hypothesis that exogenous nitric oxide (NO) enhances secondary metabolism in fresh roots of S. baicalensis Georgi via reactive oxygen species (ROS)-mediated pathways, thereby improving its medicinal quality. Fresh roots of S. baicalensis Georgi were treated with sodium nitroprusside (SNP) at concentrations of 0.0, 7.5, and 20 mmol/L to induce ROS bursts. At the optimal concentration of 20 mmol/L SNP, the levels of superoxide anion radicals (O 2 ·⁻), hydrogen peroxide (H 2 O 2 ), and malondialdehyde (MDA) increased by 95.1%, 134.5%, and 211.5%, respectively ( P < 0.01). Correspondingly, the activities of antioxidant enzymes—including superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD)—rose by 87.2%, 47.5%, and 59.4%, respectively ( P < 0.01). The activity of phenylalanine ammonia-lyase (PAL), a key enzyme in flavonoid biosynthesis, increased by 70% ( P < 0.01). While the contents of major secondary metabolites (baicalin and wogonoside) remained stable, the levels of their high-activity derivatives (baicalein and wogonin) increased dramatically by 1603.2% and 1256.1%, respectively ( P < 0.01). Pharmacological assays revealed that 20 mmol/L SNP treatment enhanced pharmacological activities: the inhibition rate of body temperature elevation in rats, the inhibition rate of writhing response in mice, and the inhibition rate of auricular swelling in mice increased by 11.7%, 14.9%, and 27.4% ( P < 0.01), respectively. This treatment also reduced serum interleukin-6 (IL-6) and tumor necrosis factor- α (TNF- α ) levels in mice by 13.5% and 16.8% ( P < 0.01), respectively. Collectively, this study confirms that exogenous NO induces ROS bursts in fresh S. baicalensis Georgi roots and improves the quality of cultivated Radix Scutellariae via ROS-mediated enhancement of secondary metabolism, and clarifies the NO-ROS-secondary metabolism regulatory axis, providing valuable insights for quality improvement strategies of other medicinal plants.

Nitric oxide (NO) is an important biological signaling molecule. S-nitrosoglutathione reductase (GSNOR), a master regulator of NO signaling, regulates various biological processes. However, little is known about the role of GSNOR in Aspergillus flavus. Here, we identified a gene encoding GSNOR in this aflatoxigenic fungus and demonstrated that GSNOR shows activity during the critical life cycle stages, including spore germination, hyphal growth, and conidiogenesis. We found that GSNOR plays a crucial role in NO homeostasis, as GSNOR deletion resulted in significantly elevated NO levels and heightened sensitivity to exogenous NO stress. GSNOR also participated in multiple biological processes in A. flavus; for that, GSNOR deletion impaired conidia germination, reduced growth, decreased conidiogenesis and sclerotial development, attenuated virulence on kernels, and notably decreased aflatoxin production. Furthermore, we demonstrated that GSNOR is important for reactive oxygen species (ROS) balance, as its deletion significantly elevated mycelial ROS levels and made the strain more sensitive to oxidative stress.IMPORTANCEAspergillus flavus is a notorious saprophytic filamentous fungus, with its production of carcinogenic aflatoxins posing serious threats to food safety and human health. Aflatoxin contamination prevention and control have long been a global challenge. In previous studies, we observed that nitric oxide (NO) significantly inhibits the aflatoxin production by A. flavus. This study further investigated the role of the key regulatory enzyme S-nitrosoglutathione reductase (GSNOR) in the NO signaling pathway. Our findings indicate that GSNOR is crucial for maintaining both NO homeostasis and reactive oxygen species (ROS) balance and plays an essential role in fungal development, pathogenicity, and aflatoxin biosynthesis. These results highlight the potential of targeting components in the NO signaling pathway, such as GSNOR, as a novel strategy for the early prevention of aflatoxin contamination in food.

Salinity stress severely hinders global agricultural productivity, and the issue will only increase under current climate scenarios. Due to complexity of salinity tolerance traits, crop breeding for salt tolerance remains a highly challenging task. The exogenous application of growth regulators, such as nitric oxide (NO), is considered a viable practical alternative to boost crop yield and quality under conditions of soil salinity. Numerous papers reported beneficial role of exogenous NO application on plant growth under salt stress, but very few explored the mechanistic basis of this process. Here, we investigated the effects of NO (generated by 0.1 mM NO donor sodium nitroprusside) on ionic homeostasis in pea mesophyll cells in response to 100 mM NaCl and 10 mM H2O2 treatments. Membrane potential (MP) and fluxes of Na+, K+, and Ca2+ were measured using the Microelectrode Ion Flux Estimation (MIFE) technique. Application of NO reduced Na+ accumulation and salt-induced K+ loss from leaf mesophyll, thus improving cell viability and leaf photochemistry (SPAD and Fv/Fm characteristics). These ameliorating effects were attributed to NO's ability to restore (otherwise depolarized) MP, enhance Na+ efflux from the cytosol, and alter sensitivity of reactive oxygen species (ROS)-inducible Ca2+- and K+-permeable ion channels. Pharmacological experiments indicated that the Na+ efflux was attributed to Na+/H+ exchanger activity. Altogether, this study demonstrated, for the first time, a direct control of plasma membrane ion transporters in leaf mesophyll cells by NO, thereby affecting NaCl-induced Ca2+ signaling and intracellular Na+ and K+ homeostasis, thus conferring salt tolerance in pea mesophyll cells. These findings expanded our understanding of the role of NO in enhancing salinity stress tolerance in plants.

Nitric oxide-modulating biomaterials for therapeutic Immunoengineering.