NADPH Oxidase Activity Research Articles

A bi-kinase module sensitizes and potentiates plant immune signaling.

Systemic signaling is an essential hallmark of multicellular life. Pathogen encounter occurs locally but triggers organ-scale and organismic immune responses. In plants, elicitor perception provokes systemically expanding Ca2+ and H2O2 signals conferring immunity. Here, we identify a Ca2+ sensing bi-kinase module as becoming super-activated through mutual phosphorylation and as imposing synergistically enhanced NADPH oxidase activation. A combined two-layer bi-kinase/substrate phospho-code allows for sensitized signaling initiation already by near-resting elevations of Ca2+ concentration. Subsequently, it facilitates further signal wave proliferation with minimal Ca2+ amplitude requirement, triggering protective defense responses throughout the plant. Our study reveals how plants build and perpetuate trans-cellular immune signal proliferation while avoiding disturbance of ongoing cellular signaling along the path of response dissemination.

From gut to liver: Exploring the crosstalk between gut-liver axis and oxidative stress in metabolic dysfunction-associated steatotic liver disease.

Non-alcoholic fatty liver disease (NAFLD), now recognized as metabolic dysfunction-associated steatotic liver disease (MASLD), represents a significant and escalating global health challenge. Its prevalence is intricately linked to obesity, insulin resistance, and other components of the metabolic syndrome. As our comprehension of MASLD deepens, it has become evident that this condition extends beyond the liver, embodying a complex, multi-systemic disease with hepatic manifestations that mirror the broader metabolic landscape. This comprehensive review delves into the critical interplay between the gut-liver axis and oxidative stress, elucidating their pivotal roles in the etiology and progression of MASLD. Our analysis reveals several key findings: (1) Bile acid dysregulation can trigger oxidative stress through enhanced ROS production in hepatocytes and Kupffer cells, leading to mitochondrial dysfunction and lipid peroxidation; (2) Gut microbiota dysbiosis disrupts intestinal barrier function, allowing increased translocation of endotoxins like LPS, which activate inflammatory pathways through TLR4 signaling and promote oxidative stress via NADPH oxidase activation; (3) The redox-sensitive transcription factors NF-κB and Nrf2 serve as crucial mediators in the gut-liver axis, with NF-κB regulating inflammatory responses and Nrf2 orchestrating antioxidant defenses; (4) Oxidative stress-induced damage to intestinal barrier function creates a destructive feedback loop, further exacerbating liver inflammation and disease progression. These findings highlight the complex interrelationship between gut-liver axis dysfunction and oxidative stress in MASLD pathogenesis, suggesting potential therapeutic targets for disease management.

Involvement of ADAM17-Klotho Crosstalk in High Glucose-Induced Alterations of Podocyte Function.

Microalbuminuria is the earliest clinical abnormality in diabetic kidney disease. High glucose (HG) concentrations are associated with the induction of oxidative stress in podocytes, leading to disruption of the glomerular filtration barrier. Our recent study revealed a significant decrease in the membrane-bound fraction of Klotho in podocytes that were cultured under HG conditions. Given that disintegrin and metalloproteinase 17 (ADAM17) is responsible for the shedding of Klotho from the cell membrane, the present study investigated the impact of HG on the interplay between ADAM17 and Klotho in human podocytes. We demonstrated that ADAM17 protein levels significantly increased in urine, renal tissue, and glomeruli from diabetic rats, with a concomitant increase in glomerular albumin permeability. High glucose increased ADAM17 extracellular activity, NADPH oxidase activity, and albumin permeability in podocytes. These effects were reversed after treatment with ADAM17 inhibitor, in cells with downregulated ADAM17 expression, or after the addition of Klotho. Additionally, elevations of extracellular ADAM17 activity were observed in podocytes with the downregulation of Klotho expression. Our data indicate a novel mechanism whereby hyperglycemia deteriorates podocyte function via ADAM17 activation. We also demonstrated the ability of Klotho to protect podocyte function under hyperglycemic conditions in an ADAM17-dependent manner.

Effects of the pan-caspase inhibitor Q-VD-OPh on human neutrophil lifespan and function.

Human neutrophils are abundant, short-lived leukocytes that turn over at a rate of approximately 1011 cells/day via a constitutive apoptosis program. Certain growth factors, inflammatory mediators and infectious agents can delay apoptosis or induce neutrophils to die by other mechanisms. Nonetheless, a large body of data demonstrates that apoptosis of untreated neutrophils typically ensues within 24 hours of cell isolation and in vitro culture. At the molecular level apoptosis is driven by executioner caspase-3, and during this process cell proinflammatory capacity and host defense functions are downregulated. We undertook the current study to determine the extent to which human neutrophil viability and function could be prolonged by treatment with the non-toxic, irreversible, pan-caspase inhibitor Q-VD-OPh. Our data demonstrate that a single 10 μM dose of this drug was sufficient to markedly prolong cell lifespan. Specifically, we show that apoptosis was prevented for at least 5 days as indicated by analysis of nuclear morphology, DNA fragmentation, and phosphatidylserine externalization together with measurements of procaspase-3 processing and caspase activity. Conversely, mitochondrial depolarization declined despite abundant Myeloid Cell Leukemia 1 (MCL-1). At the same time, glutathione levels were maintained and Q-VD-OPh prevented age-associated increases mitochondrial oxidative stress. Regarding functional capacity, we show that phagocytosis, NADPH oxidase activity, chemotaxis, and degranulation were maintained following Q-VD-OPh treatment, albeit to somewhat different extents. Thus, a single 10 μM dose of Q-VD-OPh can sustain human neutrophil viability and function for at least 5 days.

The interactive toxic effect of homocysteine and copper on cardiac microvascular endothelial cells during ischemia-reperfusion injury.

Hyperhomocysteinemia (HHcy) is associated with the development and progression of chronic cardiovascular diseases through the deleterious effects of high levels of homocysteine (Hcy) on the cardiovascular system. However, the exact mechanism of action of Hcy on the acute injury of the cardiovascular system following ischemia/reperfusion (I/R) remains unclear. The present study demonstrated that copper mobilization occurs during cardiac I/R, and the interactive toxic effect of Hcy and mobile Cu2+ during cardiac I/R induces necroptosis of cardiac microvascular endothelial cells (CMECs) and thus enhances cardiac dysfunction. In the present study, we utilized three cardiac I/R model: isolated rat heart, in vivo model as well as cell culture, and demonstrated that copper mobilization occurs during cardiac I/R, and the interactive toxic effect of Hcy and mobile Cu2+ during cardiac I/R induces necroptosis of cardiac microvascular endothelial cells (CMECs) and thus enhances cardiac dysfunction. Furthermore, we proved that the Cu2+ chelator TTM significantly mitigated the deleterious effects of Hcy and Cu2+ on CMECs and cardiac function both in vitro and in vivo. Mechanismly, the combinative effect of Hcy and Cu2+ are associated with the production of reactive oxygen species (ROS) and nitric oxide (NO) by NADPH oxidase (NOX) and endothelial nitric oxide synthase (eNOS), respectively. Subsequently, the overproduction of toxic peroxynitrite (ONOO-) induces CMECs necroptosis. The application of ROS scavengers in CMECs resulted in a notable reduction in necroptosis mediated by Hcy and Cu2+ under hypoxia/reperfusion (H/R) condition. These findings indicate that the mechanism by which Hcy and Cu2+ enhances cardiac dysfunction under I/R condition may be attributed to the stimulation of both NOX and eNOS activity, resulting in the generation of excessive ONOO- and subsequent necroptosis of CMECs.

Legionella pneumophila subverts the antioxidant defenses of its amoeba host Acanthamoeba castellanii.

Legionella pneumophila, the causative agent of Legionnaires' disease, interacts in the environment with free-living amoebae that serve as replicative niches for the bacteria. Among these amoebae, Acanthamoeba castellanii is a natural host in water networks and a model commonly used to study the interaction between L. pneumophila and its host. However, certain crucial aspects of this interaction remain unclear. One such aspect is the role of oxidative stress, with studies focusing on reactive oxygen species (ROS) production by the host and putting less emphasis on the involvement of the host's antioxidant defenses during the infectious process. In this study, we propose to examine the consequences of infection with L. pneumophila wild-type or with an isogenic ΔdotA mutant strain, which is unable to replicate intracellularly, on A. castellanii. For this purpose, we looked at the host ROS levels, host antioxidant defense transcripts, and metabolites linked to the amoeba's antioxidant defenses. It is known that L. pneumophila WT can block the activation of NADPH oxidase as soon as it enters the macrophage and suppress ROS production compared to ΔdotA mutant strain. In addition, it has been shown in macrophages that L. pneumophila WT decreases ROS at 24 h p.i.; here we confirm this result in amoebae and suggest that this decrease could be partly explained by L. pneumophila differentially regulated host antioxidant defense transcripts at 6 h p.i.. We also explored the metabolome of A. castellanii infected or not with L. pneumophila. Among the 617 metabolites identified, four with reduced abundances during infection may be involved in antioxidant responses. This study suggests that L. pneumophila could hijack the host's antioxidant defenses during its replication to maintain a reduced level of ROS.

Isolation, identification of endophytic fungi extracts from Codiaeum variegatum and assessment of their anti-amoebic and anti-inflammatory potential

Intestinal amoebiasis (IA) remains a global public health concern. It is often confused with an inflammatory bowel disease, and is caused by Entamoeba histolytica. Although it is more widespread in tropical countries, IA is increasingly becoming a subject of concern in so-called developed countries. The limits of conventional therapy and the concern to promote phytotherapy in a sustainable and ecological manner inspired this work, during which the anti-amoebic and anti-inflammatory potential of endophyte fungi extracts isolated from Codiaeum variegatum were evaluated. C. variegatum leaves and bark was harvested at the University of Yaounde 1. Those two fragments undergo surface decontamination then were placed in petri dishes containing PDA (Potato Agar Dextrose) supplemented with amoxicillin. The cultures obtained were then subcultured in SDA (Sabouraud Dextrose Agar) and CZA (CZapek dextrose Agar) media. The resulting growth was observed using an optical microscope with an x40 objective for grouping on the basis of macroscopic and microscopic characteristics. Screening of extracts of these fungi by the Trypan blue counting method on clinical isolates of E. histolytica, made it possible to retain 4 extracts (6, 8, 14, 18) of fungi with anti-amoebic activity. Subsequently, the anti-inflammatory potential of these extracts was evaluated on a primary culture of macrophages activated by Saccharomyces cerevisiae through the inhibition of nitric oxide (NO) production, NADPH oxidase activity, and activation of alkaline phosphatase (ALP). From that culture, 28 endophytic fungi were isolated, 13 belonging to 7 genus (Absidia sp, Alternaria sp, Aspergillus sp, Trametes sp, Trichoderma sp, Fusarium sp and Geotrichum sp). Anti-amoebic tests showed that extract 14 was the most active with an inhibitory concentration 50 (IC50) of 289.87±18.57; 207.73±22.36 and 87.79±12.51µg/mL respectively after 24, 48 and 72 hours of incubation. Chemical screening of the extracts revealed the majority presence of flavonoids in the 4 extracts (6, 8, 14, and 18). The anti-inflammatory tests of these 4 extracts revealed better anti-inflammatory activity in extract 6 (IC50= 95.94 ± 21.07 µg/mL for NO; IC50= 11.78±0.27 µg/mL for NADPH oxidase). Extract 14 had the best effect on the inhibition of NO production (IC50 = 8.56±2.13 µg/mL) and on the inhibition of the growth of trophozoites. Endophytic fungi extract from C. variegatum have demonstrated promising anti-amoebic and anti-inflammatory potential and therefore deserve special attention for the development of natural and ecological alternative treatment.

HRAMS Proteomics Insights on the Anti-Filarial Effect of Ocimum sanctum: Implications in Phytochemical-Based Drug-Targeting and Designing.

Lymphatic filariasis (LF) continues to impact 657 million individuals worldwide, resulting in lifelong and chronic impairment. The prevalent anti-filarial medications-DEC, albendazole, and ivermectin-exhibit limited adulticidal efficacy. Despite ongoing LF eradication programs, novel therapeutic strategies are essential for effective control. This study examines the mechanism of action of Ocimum sanctum on the filarial parasites Setaria cervi via a synergistic biochemical and proteomics methodology. The ethanolic extract of Ocimum sanctum (EOS) demonstrated potential anti-filarial action in the MTT reduction experiment, with an LC50 value of 197.24 µg/mL. After EOS treatment, an elevation in lipid peroxidation (51.92%), protein carbonylation (48.99%), and NADPH oxidase (88.88%) activity, along with a reduction in glutathione (GSH) (-39.23%), glutathione reductase (GR) (-60.17%), and glutathione S transferase (GST) (-50.48%) activity, was observed. The 2D gel electrophoresis identified 20 decreased and 11 increased protein spots in the EOS-treated parasites relative to the control group. Additionally, in drug docking analysis, the EOS bioactive substances ursolic acid, rutin, and rosmarinic acid show a significant binding affinity with the principal differentially expressed proteins. This paper demonstrates, for the first time, that the anti-filarial efficacy of EOS is primarily facilitated by its impact on energy metabolism, antioxidant mechanisms, and stress response systems of the parasites.

C-Peptide Ameliorates Particulate Matter 2.5-Induced Skin Cell Apoptosis by Inhibiting NADPH Oxidation.

Connecting peptide (C-peptide), a byproduct of insulin biosynthesis, has diverse cellular and biological functions. Particulate matter 2.5 (PM2.5) adversely affects human skin, leading to skin thickening, wrinkle formation, skin aging, and inflammation. This study aimed to investigate the protective effects of C-peptide against PM2.5-induced damage to skin cells, focusing on oxidative stress as a key mechanism. C-peptide mitigated NADPH oxidation and intracellular reactive oxygen species (ROS) production induced by PM2.5. It also suppressed PM2.5-induced NADPH oxidase (NOX) activity and alleviated PM2.5-induced NOX1 and NOX4 expression. C-peptide protected against PM2.5-induced DNA damage, lipid peroxidation, and protein carbonylation. Additionally, C-peptide mitigated PM2.5-induced apoptosis by inhibiting intracellular ROS production. In summary, our findings suggest that C-peptide mitigates PM2.5-induced apoptosis in human HaCaT keratinocytes by inhibiting intracellular ROS production and NOX activity.

SnRK1α1-mediated RBOH1 phosphorylation regulates reactive oxygen species to enhance tolerance to low nitrogen in tomato.

Nitrogen is essential for plant growth and development. SNF1-related protein kinase 1 (SnRK1) is an evolutionarily conserved protein kinase pivotal for regulating plant responses to nutrient deficiency. Here, we discovered that the expression and activity of the SnRK1 α-catalytic subunit (SnRK1α1) increased in response to low-nitrogen stress. SnRK1α1 overexpression enhanced seedling tolerance, nitrate uptake capacity, apoplastic reactive oxygen species (ROS) accumulation, and NADPH oxidase activity in tomato (Solanum lycopersicum L.) under low-nitrogen stress compared to wild type plants, while snrk1α1 mutants exhibited the opposite phenotypes. Mutation of the NADPH oxidase gene Respiratory burst oxidase homolog 1 (RBOH1) suppressed numerous nitrate uptake and metabolism genes during low-nitrogen stress. rboh1 mutants displayed lower NADPH oxidase activity, apoplastic ROS production, and seedling tolerance to low nitrogen. Silencing RBOH1 expression also compromised SnRK1α1-mediated seedling tolerance to low-nitrogen stress. SnRK1α1 interacts with and activates RBOH1 through phosphorylation of three N-terminal serine residues, leading to increased apoplastic ROS production and enhanced tolerance to low nitrogen conditions. Furthermore, RBOH1-dependent ROS oxidatively modified the transcription factor TGA4 at residue Cys-334, which increased NRT1.1 and NRT2.1 expression under low-nitrogen stress. These findings reveal a SnRK1α1-mediated signaling pathway and highlight the essential role of RBOH1-dependent ROS production in enhancing plant tolerance to low nitrogen.

Noise causes cardiovascular disease: it's time to act.

Chronic transportation noise is an environmental stressor affecting a substantial portion of the population. The World Health Organization (WHO) and various studies have established associations between transportation noise and cardiovascular disease (CVD), such as myocardial infarction, stroke, heart failure, and arrhythmia. The WHO Environmental Noise Guidelines and recent reviews confirm a heightened risk of cardiovascular incidents with increasing transportation noise levels. We present a narrative review of the evidence from epidemiologic studies and translation studies on the adverse cardiovascular effects of transportation noise. We describe the results of a recent Umbrella+ review that combines the evidence used in the 2018 WHO Environmental Noise Guidelines with more recent (post-2015) high-quality systematic reviews of original studies. High-quality systematic reviews were included based on the quality of literature search, risk of bias assessment, and meta-analysis methodology using AMSTAR 2. Epidemiologic studies show that exposure to high levels of road traffic noise for several years lead to numerous adverse health outcomes, including premature deaths, ischemic heart disease (IHD), chronic sleep disturbances, and increased annoyance. Mechanistically, noise exposure triggers oxidative stress, inflammation, endothelial dysfunction, and circadian rhythm disruptions. These processes involve the activation of NADPH oxidase, mitochondrial dysfunction, and nitric oxide synthase uncoupling, leading to vascular and cardiac damage. Studies indicate that chronic noise exposure does not result in habituation, and susceptible individuals, such as those with pre-existing CVD, are particularly vulnerable.

Myriscagayanone C, a new compound from the fruit of myristica cagayanensis, inhibits fMLP-induced respiratory bursts by specifically preventing Akt translocation in human neutrophils.

Neutrophils that are overactivated can cause inflammatory diseases. Neutrophils possess various surface receptors, including G-protein-coupled chemoattractant receptors, which assist in recognizing pathogen attacks and the inflammatory environment. Therefore, targeting G-protein-coupled chemoattractant receptors and their downstream molecules is important for preventing abnormal neutrophil activation. This study examines the effects and underlying mechanism of myriscagayanone C, a new compound obtained from the fruit of myristica cagayanensis, on neutrophil respiratory burst induced by fMLP. The immunoblotting assay was conducted to assess the mechanisms by which myriscagayanone C inhibits fMLP-induced respiratory burst by disrupting the translocation of Akt to the cellular membrane. Briefly, myriscagayanone C suppressed the production of superoxide anions induced by fMLP on human neutrophils in a concentration-dependent manner (IC50: 4.73±0.68 μM). Myriscagayanone C blocked fMLP-induced Akt translocation to the cell membrane, inhibiting AktT308 and AktS473 phosphorylation by PDK1Y373/376 and mTORS2481, respectively. Myriscagayanone C inhibited fMLP-induced p47phox phosphorylation and translocation. Myriscagayanone C did not inhibit the activity of PI3K, the amount of phosphatidylinositol (3, 4, 5)-trisphosphate, or the translocation of phosphorylated-PDK1Y373/376 and -mTORS2481 to the membrane. Myriscagayanone C did not inhibit fMLP-induced PKC, Src, ERK1/2, p38 phosphorylation, and intracellular calcium mobilization. Myriscagayanone C did not inhibit the chemotaxis and CD11b expression induced by fMLP. Myriscagayanone C did not inhibit PMA-induced superoxide anion production and neutrophil extracellular trap formation. According to this data, myriscagayanone C inhibits fMLP-induced neutrophil superoxide anion production by interrupting the translocation of Akt to the plasma membrane, which affects the NADPH oxidase activity by preventing p47phox phosphorylation and translocation.

Comparative effects of mesenchymal stromal cells of various origins and sources on biochemical parameters in the hippocampus of rats during cerebral ischemia-reperfusion

In neurodegenerative processes of the brain, the hippocampus is primarily damaged with subsequent cognitive impairments as a result of increased generation of reactive oxygen species and reactive nitrogen species, which lead to apoptosis and necrosis of neurons. Cell therapy with the use of mesenchymal stromal cells (MSCs) has shown encouraging results regarding endogenous mechanisms of neuroregeneration in response to ischemic injury of brain structures. The work aimed to study the therapeutic potential of MSCs of various origins and sources, MSCs lysate and reference drug citicoline on the energy component of neuronal metabolism, oxidative and nitrosative stress in the rat hippocampus under conditions of cerebral ischemia-reperfusion (IR). Material and methods. The experiment was carried out on 126 Wistar rats with modeled pathology (20-minute IR of the internal carotid arteries), which were injected with MSCs derived from Wharton’s jelly of the human umbilical cord, MSCs derived from human and rat adipose tissues, rat embryonic fibroblasts, MSCs lysate and citicoline immediately after removal of the ligatures. Biochemical parameters of carbohydrate metabolism (glucose, lactate), oxidative (NADPH oxidase activity) and nitrosative stress (NO synthase activity) in the rat hippocampus were determined on days 7 and 14 after IR under the conditions of cerebral IR and on the background of its correction. Results. It was established that during IR in the rat hippocampus of the brain, the content of glucose and lactate increases, the process of aerobic oxidation of glucose is inhibited, anaerobic glycolysis increases, lactic acidosis develops in hippocampal cells, the level of NADPH oxidase activity decreases, and an imbalance occurs in the functioning of the nitrogen monoxide system. A positive effect of human umbilical cord Wharton’s jelly-derived MSC and rat embryonic fibroblasts transplantation, and the use of citicoline on the stabilization of glucose, lactate, NADPH-oxidase and nitric oxide levels was found. Transplantation of human and rat adipose-derived MSCs was significantly less effective than citicoline and demonstrated no statistically significant modulatory effect on biochemical parameters in the hippocampus of experimental animals with IR. Conclusions. Transplantation of the human umbilical cord Wharton’s jelly-derived MSCs was not inferior to the citicoline and, better than other tested MSCs and their lysate, contributed to the recovery of disturbed energy processes (glucose level) and eliminated metabolic acidosis (lactate level) in the hippocampus of rats, i.e. had a positive modulating effect on the oxidant-antioxidant balance (NADPH oxidase activity level).

Comparative effects of mesenchymal stromal cells of various origins and sources on biochemical parameters in the hippocampus of rats during cerebral ischemia-reperfusion

In neurodegenerative processes of the brain, the hippocampus is primarily damaged with subsequent cognitive impairments as a result of increased generation of reactive oxygen species and reactive nitrogen species, which lead to apoptosis and necrosis of neurons. Cell therapy with the use of mesenchymal stromal cells (MSCs) has shown encouraging results regarding endogenous mechanisms of neuroregeneration in response to ischemic injury of brain structures. The work aimed to study the therapeutic potential of MSCs of various origins and sources, MSCs lysate and reference drug citicoline on the energy component of neuronal metabolism, oxidative and nitrosative stress in the rat hippocampus under conditions of cerebral ischemia-reperfusion (IR). Material and methods. The experiment was carried out on 126 Wistar rats with modeled pathology (20-minute IR of the internal carotid arteries), which were injected with MSCs derived from Wharton’s jelly of the human umbilical cord, MSCs derived from human and rat adipose tissues, rat embryonic fibroblasts, MSCs lysate and citicoline immediately after removal of the ligatures. Biochemical parameters of carbohydrate metabolism (glucose, lactate), oxidative (NADPH oxidase activity) and nitrosative stress (NO synthase activity) in the rat hippocampus were determined on days 7 and 14 after IR under the conditions of cerebral IR and on the background of its correction. Results. It was established that during IR in the rat hippocampus of the brain, the content of glucose and lactate increases, the process of aerobic oxidation of glucose is inhibited, anaerobic glycolysis increases, lactic acidosis develops in hippocampal cells, the level of NADPH oxidase activity decreases, and an imbalance occurs in the functioning of the nitrogen monoxide system. A positive effect of human umbilical cord Wharton’s jelly-derived MSC and rat embryonic fibroblasts transplantation, and the use of citicoline on the stabilization of glucose, lactate, NADPH-oxidase and nitric oxide levels was found. Transplantation of human and rat adipose-derived MSCs was significantly less effective than citicoline and demonstrated no statistically significant modulatory effect on biochemical parameters in the hippocampus of experimental animals with IR. Conclusions. Transplantation of the human umbilical cord Wharton’s jelly-derived MSCs was not inferior to the citicoline and, better than other tested MSCs and their lysate, contributed to the recovery of disturbed energy processes (glucose level) and eliminated metabolic acidosis (lactate level) in the hippocampus of rats, i.e. had a positive modulating effect on the oxidant-antioxidant balance (NADPH oxidase activity level).

A transient blood IL-17 increase triggers neuroinflammation in cerebellum and motor incoordination in hyperammonemic rats

Patients with liver cirrhosis may show minimal hepatic encephalopathy (MHE) with motor incoordination which is reproduced in hyperammonemic rats. Hyperammonemia induces peripheral inflammation which triggers neuroinflammation and enhanced GABAergic neurotransmission in cerebellum and motor incoordination. The mechanisms involved remain unknown. The aims were to assess if the early increase of peripheral IL-17 triggers motor incoordination in hyperammonemic rats and to identify some underlying mechanisms. We assessed if blocking peripheral IL-17 with anti-IL-17 at 2–4 days of hyperammonemia prevents motor incoordination and analyzed underlying mechanisms. Hyperammonemia induces a transient blood IL-17 increase at days 3–4. This is associated with increased IL-17 receptor membrane expression and activation in cerebellum, leading to NADPH oxidase activation, increased superoxide production and MLCK that induce blood–brain barrier (BBB) permeabilization by reducing occludin and ZO-1. BBB permeabilization facilitates the entry of IL-17, which increases in cerebellum and activates microglia. This increases TNFα and the TNFR1-S1PR2-CCL2-BDNF-TrkB pathway. This enhances GABAergic neurotransmission which impairs motor coordination. Blocking peripheral IL-17 with anti-IL-17 prevents all the above process and prevents motor incoordination. Early treatment to reduce blood IL-17 may be a useful treatment to reverse motor incoordination in patients with MHE.Graphical abstract

Effects of sucrose and 1-MCP on enzymatic and nonenzymatic antioxidants in postharvest Gynura bicolor DC

After harvesting, Gynura bicolor DC (G. bicolor) undergoes rapid quality deterioration, including decay, nutrient loss, and reactive oxygen species (ROS) burst, greatly limiting its shelf life. This study was performed to evaluate the effects of treatment with sucrose and 1-methylcyclopropene (1-MCP) on indices of quality deterioration, ROS metabolism, and phenylpropanoid metabolism, with the goals of resisting oxidative stress and improving the postharvest quality of G. bicolor. Sucrose treatment activated phenylpropanoid metabolism, increased phenylalanine ammonia lyase (PAL), cinnamate-4-hydroxylase (C4H), and 4-coumarate:CoA ligase (4CL) activities, and promoted the accumulation of phenolics. The increases in nonenzymatic antioxidants decreased O2·−, ·OH, and H2O2 contents. Conversely, 1-MCP treatment increased NADPH oxidase (NOX) activity and induced transient oxidative stress, which significantly activated enzymatic antioxidants, including superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX), thus maintaining a lower ROS level. Treatment with both sucrose and 1-MCP exhibited a synergistic effect on alleviating oxidative stress, downregulating GbSAG101 and GbATG expression. Compared with sucrose treatment, 1-MCP treatment showed a greater inhibitory effect on ROS burst. Phenolic substance contents were similar in plants treated with sucrose and 1-MCP at the end of the storage period. Therefore, we speculated that higher levels of antioxidant enzyme activity may decrease the consumption of nonenzymatic antioxidants. Our experimental results indicated the involvement of different pathways in the effects of sucrose treatment, 1-MCP treatment, and combined treatment on delaying ROS burst, and suggested potential alternative methods for the preservation of vegetables with stems during the postharvest storage.

Effect of elevated ammonium on biotic and abiotic stress defense responses and expression of related genes in cucumber (Cucumis sativus L.) plants

Ammonium (NH4+) enhances plant defense mechanisms but can be phytotoxic as the sole nitrogen source. To investigate the impact of a balanced NH4+ and NO3− ratio on plant defense parameters without adverse effects, cucumber plants (Cucumis sativus L.) were grown under control (14 mM NO3− + 2 mM NH4+) and elevated level of NH4+ (eNH4+, 8 mM NO3−+ 8 mM NH4+). Plants subjected to eNH4+ showed significantly increased shoot and root biomass by about 41% and 47%, respectively. Among the antioxidant enzymes studied, ascorbate peroxidase (EC 1.11.1.11) activity was increased up to 3.3 fold in eNH4+ compared with control plants, which was associated with enhanced resistance to paraquat. Upregulation of PATHOGENESIS RELATED PROTEIN 4 (PR4) and LIPOXYGENASE 1 (LOX1), accompanied by increased concentrations of salicylic acid and nitric oxide, conferred more excellent resistance of eNH4+ plants to powdery mildew infection. However, the expression levels of ACC OXIDASE 1 (ACO1) and RESPIRATORY BURST OXIDASE HOMOLOGS B (RBOHB) were lower in eNH4+ plants, which was consistent with decreased NADPH oxidase activity and lower leaf H2O2 levels. The biosynthesis of phenolics was enhanced, whereas the activities of polymerizing enzymes and lignin deposition were reduced by half in eNH4+ plants. Besides, a significant effect on plant biomass under salt or drought stress has not been observed between control and eNH4+ plants. These results showed that different defense pathways are distinctively affected by eNH4+ treatment, and the NH4+ to NO3− ratio may play a role in fine-tuning the plant defense response.

Angiotensin type-1 receptor autoantibodies promote alpha-synuclein aggregation in dopaminergic neurons.

Angiotensin, through its type-1 receptor (AT1), is a major inducer of inflammation and oxidative stress, contributing to several diseases. Autoimmune processes have been involved in neurodegeneration, including Parkinson's disease (PD). AT1 autoantibodies (AT1-AA) enhance neurodegeneration and PD, which was related to increased neuronal oxidative stress and neuroinflammation. However, the effect of AT1-AA on α-synuclein aggregation, a major factor in PD progression, has not been studied. In cultures of dopaminergic neurons, we observed that AT1-AA promote aggregation of α-synuclein, as AT1-AA upregulated major mechanisms involved in the α-synuclein aggregation process such as NADPH-oxidase activation and intracellular calcium raising. The results further support the role of AT1 receptors in dopaminergic neuron degeneration, and several recent clinical studies observing the neuroprotective effects of AT1 receptor blockers.

Accumulation of Advanced Oxidation Protein Products Promotes Age-Related Decline of Type H Vessels in Bone.

Type H vessels have been proven to couple angiogenesis and osteogenesis. The decline of type H vessels contributes to bone loss in the aging process. Aging is accompanied by the accumulation of advanced oxidation protein products (AOPPs). However, whether AOPP accumulation is involved in age-related decline of type H vessels is unclear. Here, we show that the increase of AOPP levels in plasma and bone was correlated with the decline of type H vessels and loss of bone mass in old mice. Exposure of microvascular endothelial cells to AOPPs significantly inhibited cell proliferation, migration, and tube formation; increased NADPH oxidase activity and excessive reactive oxygen species generation; upregulated the expression of vascular cell adhesion molecule-1 and intercellular cell adhesion molecule-1; and eventually impaired angiogenesis, which was alleviated by redox modulator N-acetylcysteine and NADPH oxidase inhibitor apocynin. Furthermore, reduced AOPP accumulation by NAC treatment was able to alleviate significantly the decline of type H vessels, bone mass loss, and deterioration of bone microstructure in old mice. Collectively, these findings suggest that AOPPs accumulation contributes to the decline of type H vessels in the aging process, and illuminate a novel potential mechanism underlying age-related bone loss.

β3-adrenergic agonist counters oxidative stress and Na+-K+ pump inhibitory S-glutathionylation of placental cells: implications for preeclampsia.

Oxidative stress from placental ischemia/reperfusion and hypoxia/reoxygenation (H/R) in preeclampsia is accompanied by Na+-K+ pump inhibition and S-glutathionylation of its β1 subunit (GSS-β1), a modification that inhibits the pump. β3-adrenergic receptor (β3-AR) agonists can reverse GSS-β1. We examined the effects of the agonist CL316,243 on GSS-β1 and sources of H/R-induced oxidative stress in immortalized first-trimester human trophoblast (HTR-8/SVneo) and freshly isolated placental explants from normal-term pregnancies. H/R increased GSS-β1 and, reflecting compromised α1/β1 subunit interaction, reduced α1/β1 pump subunit coimmunoprecipitation. H/R increased p47phox/p22phox NADPH oxidase subunit coimmunoprecipitation, reflecting membrane translocation of cytosolic p47phox that is needed to activate NADPH oxidase. Fluorescence of O2•--sensitive dihydroethidium increased in parallel. H/R increased S-glutathionylation of endothelial nitric oxide synthase (GSS-eNOS) that uncouples nitric oxide synthesis toward the synthesis of O2•- and reduced trophoblast migration. Oxidative stress induced by tumor necrosis factor α increased soluble fms-like tyrosine kinase receptor 1 (sFlt-1) trophoblast release, a marker of preeclampsia, and reduced trophoblast integration into endothelial cellular networks. CL316,243 eliminated H/R-induced GSS-β1 and decreases of α1/β1 subunit coimmunoprecipitation, eliminated NADPH oxidase activation and increases in GSS-eNOS, restored trophoblast migration, eliminated increased sFlt-1 release, and restored trophoblast integration in endothelial cell networks. H/R-induced GSS-β1, α1/β1 subunit coimmunoprecipitation, and NADPH oxidase activation of placental explants reflected effects of H/R for trophoblasts and CL316,243 eliminated these changes. We conclude a β3-AR agonist counters key pathophysiological features of preeclampsia in vitro. β3 agonists already in human use for another purpose are potential candidates for repurposing to treat preeclampsia.NEW & NOTEWORTHY H/R-induced oxidative stress and deficient NO-dependent placentation are features of preeclampsia, yet nonspecific antioxidants and NO donors are ineffective. Here, activation of the microdomain-confined signaling pathway with an agonist for the eNOS-coupled β3-AR eliminates inhibitory glutathionylation of the Na+-K+ pump's β1 subunit, uncoupling of eNOS, and activation of NADPH oxidase that are sources of H/R-induced oxidative stress. The agonist also eliminates H/R-induced inhibition of trophoblast migration and their integration into an endothelial network.