High Levels Of Nitric Oxide Research Articles

Oxidants-induced high levels of nitric oxide impair the antioxidative property of molybdenum nanoparticles in HUVE cells

In recent years, there has been significant interest in the biomedical potential of redox-active molybdenum nanoparticles (Mo NPs) due to their varied responses from oxidative to antioxidative. Our knowledge of the bio-response of Mo NPs in endothelial cells is lacking. We, therefore, are prompted to examine the biocompatibility of well-characterized Mo NPs in human endothelial (HUVE) cells and their potential antioxidative response against standard oxidants- tert-butyl hydroperoxide (t-BHP) and hydrogen peroxide (H2O2). The study found that Mo NPs were highly biocompatible in HUVE cells and enhanced cellular antioxidant glutathione (GSH), significantly protecting cells against exogenous oxidants. Moreover, Mo NPs significantly restored the loss of mitochondrial membrane potential (MMP) determined by the Rh123 probe. They decreased reactive oxygen species (ROS) levels as measured by DHE and DCFH-DA probes. In light of Mo involvement in the nitric oxide (NO) metabolism and dependency of HUVE cells on NO signaling, intracellular NO was determined using DAR-2 fluorescent dye and the Griess assay. NO was not produced significantly by Mo NPs alone or t-BHP or H2O2. However, NO generation was significantly high when HUVE cells were co-exposed with Mo NPs and exogenous oxidants. Although the exact mechanism is unclear to us, our study concludes that the enhanced generation of NO under the co-exposure of oxidants with Mo NPs can impair the potential antioxidative property of Mo NPs, especially in endothelial cells. The study also suggests that NO modulatory strategies can improve and broaden the antioxidative properties of Mo-based nanoparticles.

Effect of low-molecular-weight heparin calcium combined with magnesium sulfate and labetalol on coagulation, vascular endothelial function and pregnancy outcome in early-onset severe preeclampsia.

This paper was aimed at unveiling the effect of low-molecular-weight heparin calcium (LMWH) combined with magnesium sulfate and labetalol on coagulation, vascular endothelial function, and pregnancy outcome in early-onset severe preeclampsia (EOSP). Pregnant women with EOSP were divided into the control group and the study group, each with 62 cases. Patients in the control group were treated with labetalol and magnesium sulfate, and those in the study group were treated with LMWH in combination with the control grou Blood pressure (systolic blood pressure [SBP] and diastolic blood pressure [DBP]), 24-h urine protein, coagulation indices [D-dimer (D-D), plasma fibrinogen (Fg), prothrombin time (PT), activated partial thromboplastin time (APTT), and prothrombin time (TT)], endothelial function [endothelin (ET-1) and nitric oxide (NO)], oxidative stress indices [oxidized low-density lipoproteins (ox-LDL), lipid peroxidation (LPO), superoxide dismutase (SOD), and malondialdehyde (MDA)], pregnancy outcome, and adverse effects occurred in the two groups were compared. After treatment, lower SBP, DBP, and 24-h urine protein levels; lower Fg and D-D levels; higher PT, APPT, and TT levels; higher NO levels; lower ET-1 levels; lower ox-LDL, MDA, and LPO levels; higher SOD levels; and lower incidence of adverse pregnancy and adverse reactions were noted in the study group in contrast to the control group. EOSP patients given with LMWH combined with magnesium sulfate and labetalol can effectively reduce the patient's blood pressure and urinary protein level; improve coagulation function, oxidative stress, and vascular endothelial function indices; reduce the adverse pregnancy outcomes; and improve the safety of treatment.

Regulatory effects of vitamin E nano-emulsion on blood metabolites, immunological variables, testicular architecture, and sperm ultrastructure of heat-stressed V-line rabbit bucks

The study investigated the beneficial effects of vitamin E-loaded nano-emulsion (NEVE) on hemato-biochemical changes, oxidative stress, inflammatory responses, immunological variables, and semen quality in heat-stressed rabbit bucks. Forty adult V-line rabbit bucks were randomly assigned to four groups and fed diets containing NEVE at 0 (NEVE0), 50 (NEVE50), 100 (NEVE100), and 200 (NEVE200) mg/kg for 12 weeks. The NEVE had a mean particle diameter of 32.6 nm, a polydispersity index of 0.6, and a zeta potential of −31.1 mV. The dietary NEVE significantly improved the hematological, blood protein, lipid profile, and liver functions of rabbit bucks (p < 0.05). The NEVE200 treated group showed higher levels of nitric oxide, TAC, and SOD in blood serum and seminal plasma compared to the control group (p < 0.05). Meanwhile, levels of MDA, IL-4, and TNF-α were significantly shrunken in response to the dietary treatment (p < 0.0001), while the IgA and IgG were improved in the NEVE100 and NEVE200 groups. The NEVE200 exhibited higher progressive motility, ejaculate volume, viability, and better libido (lower reaction time) than the untreated group (p < 0.05). TEM images illustrated that NEVE200 exhibited higher sperm cells with a normal structure, intact plasma membranes, and acrosomes than the other groups. Moreover, testicular architecture was improved with NEVE supplementation. Overall, the dietary addition of NEVE at 100 or 200 mg/kg could be an effective strategy to improve the health and semen quality of rabbit bucks during hot climates.

Renin-angiotensin system-mediated nitric oxide signaling in podocytes.

Nitric oxide (NO) is widely recognized for its role in regulating renal function and blood pressure. However, the precise mechanisms by which NO affects renal epithelial cells remain understudied. Our previous research has shown that NO signaling in glomerular podocytes can be initiated by Angiotensin II (ANG II) but not by ATP. This study aims to elucidate the crucial interplay between the renin-angiotensin system (RAS) and NO production in podocytes. To conduct our research, we used cultured human podocytes and freshly isolated rat glomeruli. A variety of RAS peptides were used, alongside confocal microscopy, to detect NO production and NO/Ca2+ cross talk. Dynamic changes in the podocyte cytoskeleton, mediated by RAS-NO intracellular signaling, were observed using fluorescent labeling for F-actin and scanning probe microscopy. The experiments demonstrated that ANG II and ANG III generated high levels of NO by activating the angiotensin II type 2 receptor (AT2R). We did not detect functional MAS receptor presence in podocytes, and the moderate NO response to ANG 1-7 was also mediated through AT2R. Furthermore, NO production impacted intracellular Ca2+ signaling and correlated with an increase in podocyte volume and growth. Scanning probe experiments revealed that AT2R activation and the corresponding NO generation are responsible for the protrusion of podocyte lamellipodia. Taken together, our data indicate that AT2R activation enhances NO production in podocytes and subsequently mediates changes in Ca2+ signaling and podocyte volume dynamics. These mechanisms may play a significant role in both physiological and pathophysiological interactions between the RAS and podocytes.NEW & NOTEWORTHY The renin-angiotensin system plays a crucial role in the production of intracellular nitric oxide within podocytes. This mechanism operates through the activation of the angiotensin II type 2 receptor, leading to dynamic modifications in intracellular calcium levels and the actin filament network. This intricate process is vital for linking the activity of angiotensin receptors to podocyte function.

Association between Angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism on the susceptibility to psoriasis and oxidative stress (OS) in a cohort of pediatric psoriatic patients in Sri Lanka: A cross sectional study.

Pediatric psoriasis accounts for nearly one-third of the global psoriasis burden. Multiple lines of evidence have shown the relationship between Angiotensin-converting enzyme (ACE) Insertion (I)/deletion(D) polymorphism with psoriasis susceptibility, and oxidative stress (OS) in psoriatic patients. However, such studies, particularly on pediatric psoriasis, are scarce in the local setting. Our study investigated the prevalence of ACE I/D polymorphism and its associations with oxidative stress in pediatric psoriasis patients in Sri Lanka. Thirty patients were recruited for this study after obtaining ethical clearance. The polymerase chain reaction was used to explore the ACE I/D polymorphism. Serum Nitric Oxide (NO) levels and the Total Antioxidant Capacity (TAC) were measured using the Griess assay and the FRAP assay. Clinical details were obtained from the clinic reports. Female predominance (76.67%) in pediatric psoriasis was reported, while Plaque psoriasis (66.67%) was found to be the most prevalent form. I/D was reported as the predominant genotype (66.67%) while I/I and D/D genotypes were recorded in 23.33% and 10% of patients, respectively. Significantly higher NO levels were observed in I/D patients than in I/I patients but not among other groups. No differences in TAC among ACE genotypes were reported. This pilot study revealed female gender and I/D genotype with increased NO levels as risk factors for pediatric psoriasis in Sri Lanka. However, it is prudent to increase the sample size to further validate the results.

Analysis of Risk Factors and The Impact of Nitric Oxide (NO) and Carbon Monoxide (CO) Levels on Lung Function: A Comparative Study of Palm Oil Factory Workers and the Community in Kuantan Mudik District, Riau, Indonesia

Background: Air pollution, particularly from industrial sources like palm oil factories, poses a significant risk to respiratory health. This study aimed to investigate the impact of nitric oxide (NO) and carbon monoxide (CO) exposure on lung function in palm oil factory workers and compare it to the lung function of individuals in the surrounding community. Methods: A cross-sectional study was conducted in Kuantan Mudik District, Riau, Indonesia. One hundred palm oil factory workers with a minimum of two years of employment and 100 community members not occupationally exposed to NO and CO were recruited. Lung function was assessed using spirometry, and NO and CO levels were measured using specialized analyzers. Participants completed the European Community Respiratory Health Survey (ECRHS) questionnaire, and demographic and clinical data were collected. Statistical analyses included univariate, bivariate, and multivariate logistic regression. Results: Factory workers exhibited significantly higher levels of NO (p=0.000) and a higher prevalence of restrictive lung function (17%) compared to the community members (3%). Multivariate analysis identified NO exposure as the most significant risk factor for restrictive lung function (p=0.005). Each unit increase in NO was associated with a 1.12-fold increased risk of restrictive lung function. CO exposure, age, gender, smoking history, and the use of personal protective equipment (PPE) were not significantly associated with lung function impairment in this study. Conclusion: Occupational exposure to NO in palm oil factories is a significant risk factor for restrictive lung function impairment. Regular monitoring of NO levels and the implementation of effective control measures are crucial to protect the respiratory health of palm oil factory workers.

Arsenic Trioxide Suppresses Angiogenesis in Non-small Cell Lung Cancer via the Nrf2-IL-33 Signaling Pathway.

Non-Small Cell Lung Cancer (NSCLC) ranks as a leading cause of cancer-related mortality, necessitating the urgent search for cost-effective and efficient anti-NSCLC drugs. Our preliminary research has demonstrated that arsenic trioxide (ATO) significantly inhibits NSCLC angiogenesis, exerting anti-tumor effects. In conjunction with existing literature reports, the Nrf2-IL-33 pathway is emerging as a novel mechanism in NSCLC angiogenesis. This study aimed to elucidate whether ATO can inhibit NSCLC angiogenesis through the Nrf2-IL-33 pathway. Immunohistochemistry was employed to assess the expression of Nrf2, IL-33, and CD31 in tumor tissues from patients with NSCLC. DETA-NONOate was used as a nitric oxide (NO) donor to mimic high levels of NO in the tumor microenvironment. Western blot, quantitative real-time PCR, and enzyme-linked immunosorbent assay were utilized to evaluate the expression of Nrf2 and IL-33 in the NCI-H1299 cell line. Subcutaneous xenograft models were established in nude mice by implanting NCI-H1299 cells to assess the anti-tumor efficacy of ATO. High expression levels of Nrf2 and IL-33 were observed in tumor samples from patients with NSCLC, and Nrf2 expression positively correlated with microvascular density in NSCLC. In vitro, NO (released from 1mM DETA-NONOate) promoted activation of the Nrf2-IL-33 signaling pathway in NCI-H1299 cells, which was reversed by ATO. Additionally, both Nrf2 deficiency and ATO treatment significantly attenuated NOinduced IL-33 expression. In vivo, both ATO and the Nrf2 inhibitor ML385 demonstrated significant inhibitory effects on angiogenesis tumor growth. Nrf2-IL-33 signaling is usually activated in NSCLC and positively correlates with tumor angiogenesis. ATO effectively disrupts the activation of the Nrf2-IL-33 pathway in NSCLC and thus inhibits angiogenesis, suggesting its potential as an anti-angiogenic agent for use in the treatment of NSCLC.

Blends of Organic Acids Are Weaponizing the Host iNOS and Nitric Oxide to Reduce Infection of Piscirickettsia salmonis in vitro.

For the last 30 years, Piscirickettsia salmonis has caused major economic losses to the aquaculture industry as the aetiological agent for the piscirickettsiosis disease. Replacing the current interventions, based on antibiotics, with natural alternatives (e.g., organic acids) represents a priority. With this study, we aimed to better understand their biological mechanism of action in an in vitro model of infection with salmon epithelial cells (CHSE-214). Our first observation revealed that at the sub-inhibitory concentration of 0.5%, the organic acid blend (Aq) protected epithelial cell integrity and significantly reduced P. salmonis invasion. The MIC was established at 1% Aq and the MBC at 2% against P. salmonis. The sub-inhibitory concentration significantly increased the expression of the antimicrobial peptides Cath2 and Hepcidin1, and stimulated the activity of the innate immune effector iNOS. The increase in iNOS activity also led to higher levels of nitric oxide (NO) being released in the extracellular space. The exposure of P. salmonis to the endogenous NO caused an increase in bacterial lipid peroxidation levels, a damaging effect which can ultimately reduce the pathogen's ability to attach or multiply intracellularly. We also demonstrate that the increased NO release by the host CHSE-214 cells is a consequence of direct exposure to Aq and is not dependent on P. salmonis infection. Additionally, the presence of Aq during P. salmonis infection of CHSE-214 cells significantly mitigated the expression of the pro-inflammatory cytokines IL-1β, IL-8, IL-12, and IFNγ. Taken together, these results indicate that, unlike antibiotics, natural antimicrobials can weaponize the iNOS pathway and secreted nitric oxide to reduce infection and inflammation in a Piscirickettsia salmonis in vitro model of infection.

Determination of Malondialdehyde, Nitric Oxide, Reduced Glutathione, Sialic Acid and Ceruloplasmin Levels in Sheep Liver Tissue With Hydatic Cyst

Cystic echinococcosis is a prevalent helminth-zoonosis that poses a significant threat to human and animal health worldwide. Although it typically shows an asymptomatic clinical course, it has been reported to cause many damages and biochemical changes in tissues and organism. The aim of this study was to investigate the concentrations of nitric oxide (NO), reduced glutathione (GSH), malondialdehyde (MDA), ceruloplasmin (Cp), and total sialic acid (TSA) in sheep liver tissue affected by cystic echinococcosis. The study was carried out on sheep between 4-5 years of age brought to the slaughterhouse in the Igdir territory. The livers were examined post-mortem for cystic echinococcosis and cystic structures. Sheep liver tissues that tested positive for protoscolex were designated as the ‘infected group’, while healthy sheep liver tissues without lesions were assigned to the ‘control group’. The results showed significantly higher levels of NO, MDA, Cp and TSA in the infected group compared to the control group (p&amp;lt;0.05), while GSH levels were significantly lower (p&amp;lt;0.05). These findings indicate that cystic echinococcosis in sheep is closely associated with mechanisms of inflammation, oxidative stress, and tissue damage. Moreover, our study provides insights into the oxidative response of cystic echinococcosis in liver tissue and enhances our understanding of the disease’s pathogenesis.

Regional variations in allergen-induced airway inflammation correspond to changes in soluble guanylyl cyclase heme and expression of heme oxygenase-1.

Asthma is characterized by airway remodeling and hyperreactivity. Our earlier studies determined that the nitric oxide (NO)-soluble guanylyl cyclase (sGC)-cGMP pathway plays a significant role in human lung bronchodilation. However, this bronchodilation is dysfunctional in asthma due to high NO levels, which cause sGC to become heme-free and desensitized to its natural activator, NO. In order to determine how asthma impacts the various lung segments/lobes, we mapped the inflammatory regions of lungs to determine whether such regions coincided with molecular signatures of sGC dysfunction. We demonstrate using murine models of asthma (OVA and CFA/HDM) that the inflamed segments of these murine lungs can be tracked by upregulated expression of HO1 and these regions in turn overlap with regions of heme-free sGC as evidenced by a decreased sGC-α1β1 heterodimer and an increased response to heme-independent sGC activator, BAY 60-2770, relative to naïve uninflamed regions. We also find that NO generated from iNOS upregulation in the inflamed segments has a higher impact on developing heme-free sGC as increasing iNOS activity correlates linearly with elevated heme-independent sGC activation. This excess NO works by affecting the epithelial lung hemoglobin (Hb) to become heme-free in asthma, thereby causing the Hb to lose its NO scavenging function and exposing the underlying smooth muscle sGC to excess NO, which in turn becomes heme-free. Recognition of these specific lung segments enhances our understanding of the inflamed lungs in asthma with the ultimate aim to evaluate potential therapies and suggest that regional and not global inflammation impacts lung function in asthma.

Propionyl-l-carnitine mitigates ischemia-reperfusion injury in rat epigastric island flaps

BackgroundIschemia-reperfusion injury presents a substantial concern in various medical scenarios, notably in reconstructive surgery involving tissue flaps. Despite reports on the protective benefits of Propionyl-l-carnitine against ischemia-reperfusion injury, a thorough assessment of its efficacy in epigastric island flap models is currently lacking. MethodsSixteen male Sprague-Dawley rats underwent epigastric island flap surgery and were divided into two groups: a Propionyl-l-carnitine group that received intraperitoneal Propionyl-l-carnitine prior to ischemia induction and a sham group that received saline treatment. A comprehensive evaluation was performed including macroscopic, biochemical and histological assessments encompassing measurements of flap survival areas, lipid peroxidation (malondialdehyde), glutathione, myeloperoxidase, nitric oxide and peripheral neutrophil counts. ResultsThe Propionyl-l-carnitine group demonstrated significantly increased flap survival areas when compared to the sham group. Administration of Propionyl-l-carnitine led to reduced malondialdehyde levels and elevated glutathione levels indicating a reduction in oxidative stress. Furthermore, the Propionyl-l-carnitine group exhibited lower myeloperoxidase levels, higher nitric oxide levels and reduced peripheral neutrophil counts, suggesting a decrease in the inflammatory response. Histopathological analysis revealed decreased levels of inflammation, necrosis, polymorphonuclear leukocyte infiltration and edema in the Propionyl-l-carnitine group. Additionally, vascularity was enhanced in the Propionyl-l-carnitine group. ConclusionThis study provides compelling evidence that Propionyl-l-carnitine administration effectively mitigates the deleterious effects of ischemia-reperfusion injury in epigastric island flaps. This is substantiated by the improved flap survival, diminished oxidative stress and inflammation, as well as the enhanced vascularity observed. Propionyl-l-carnitine emerges as a promising therapeutic intervention to enhance tissue flap survival in reconstructive surgery, warranting further exploration through larger-scale investigations.

Regulation of nitric oxide by phytoglobins in Lotus japonicus is involved in mycorrhizal symbiosis with Rhizophagus irregularis

Various reactive molecular species are generated in plant–microbe interactions, and these species participate in defense and symbiotic responses. Leguminous plants successfully establish symbiosis by maintaining an appropriate level of nitric oxide (NO), which is generated in the roots and nodules during root nodule symbiosis. Phytoglobin (plant hemoglobin) controls NO levels in plants. In this study, we investigated mycorrhizal symbiosis, which occurs in more than 80% of land plants, between Rhizophagus irregularis and Lotus japonicus to clarify the involvement of phytoglobin-mediated NO regulation. The mycorrhizae of L. japonicus exhibited higher NO levels in the presence of R. irregularis than in its absence, especially at the infection site. LjGlb1–1, a phytoglobin that regulates NO level in L. japonicus, was upregulated during symbiosis with R. irregularis. In transformed hairy roots carrying the ProLjGlb1–1:GUS construct, LjGlb1–1 expression was observed at the R. irregularis infection site. We further examined the symbiotic phenotypes of L. japonicus lines with high and low LjGlb1–1 expression with R. irregularis. During mycorrhizal symbiosis, the high LjGlb1–1 expression line exhibited better growth than the wild-type, whereas the low expression line exhibited poor growth. In addition, the expression of LjPT4, a phosphate transporter specific to mycorrhizal symbiosis, was higher in the high LjGlb1–1 expression line, whereas that of the tubulin gene of R. irregularis was lower in the low LjGlb1–1 expression line than in the wild-type. These results confirm that NO regulation by LjGlb1–1 is involved in mycorrhizal symbiosis in L. japonicus, as it is reportedly in nitrogen-fixing symbiosis.

Adrenoreceptor phylogeny and novel functions of nitric oxide in ascidian immune cells

Nitric oxide (NO) is a simple molecule involved in many biological processes and functions in the cardiovascular, neural, and immune systems. In recent years, NO has also been recognized as a crucial messenger in communication between the nervous and immune systems. Together with NO, catecholamines are the main group of neurotransmitters involved in cross-talk between the nervous and immune systems. Catecholamines such as noradrenaline, can act on immune cells through adrenoreceptors (ARs) present on the cell surface, and NO can cross the cell membrane and interact with secondary messengers, modulating catecholamine production. Here, we analyzed the mutual modulation by noradrenaline and NO in Phallusia nigra immune cells for specific subtypes of ARs. We also investigated the involvement of protein kinases A and C as secondary messengers to these specific subtypes of ARs in the adrenergic signaling pathway that culminates in NO modulation, and the phylogenetic distribution of ARs in deuterostome genomes. This analysis provided evidence for single-copy orthologs of α1, α2 and β-AR in ascidian genomes, suggesting that NO and NA act on a less diverse set of ARs in urochordates. Pharmacological assays showed that high levels of NO can induce ascidian immune cells to produce catecholamines. We also observed that protein kinases A and C are the secondary messengers involved in downstream modulation of NO production through an ancestral β-AR. Taken together, these results provide new information on NO as a modulator of immune cells, and reveal the molecules involved in the signaling pathway of ARs. The results also indicate that ARs may participate in NO modulation. Finally, our results suggest that the common ancestor of urochordates possessed a less complex system of ARs required for immune action and diverse pharmacological responses, since the α-ARs are phylogenetically more related to D1-receptors than are the β-ARs.

Nitric Oxide and Hypochlorite Assessment and Screening of some β-Lactamase Encoding Genes among Gram Negative Bacteria in Patients with Acute Leukemia

Objective: The serologic levels of both nitric oxide (NO) and hypochlorite (ClO-) were assessed in this study for Iraqi acute leukemic patients infected with Gram negative bacteria (GNB). Alongside, the phylogenetic analysis for both blaSHV and blaTEM genes of GNB was performed. Methods: The clinical samples were recovered from acute leukemic patients at hematology center in Baghdad from January 2021 to December 2021. The identification of bacterial isolates and susceptibility test were performed using Vitek 2 Compact System. Serum levels of NO and ClO- were assayed by Enzyme Linked Immunosorbent assay and colorimetric method, respectively. Gene screening was done utilizing polymerase chain reaction. DNA sequencing, GenBank accession numbers submission, and phylogenetic analysis were also conducted. Results: From 260 patients with acute leukemia, 485 clinical samples were collected from different sites. Of this total, 70 (15%) isolates of GNB were obtained, distributing as Klebsiella pneumoniae 23 (33%), Escherichia coli 21 (30%), Pseudomonas aeruginosa 18 (26%), and Acinetobacter baumannii 8 (11%). These isolates were mainly collected from urine 39 (55.71%), followed by blood 23 (32.85%), and the least from swabs 8 (11.42%). The infections of GNB were higher among acute myeloblastic leukemia (AML) patients 40 (57.14%) than these with acute lymphoblastic leukemia (ALL) 30 (42.85%). The levels of NO were higher among groups of patients than control groups. Additionally, ClO-levels were observed to be slightly increased in patients above these of controls. Most isolates of K. pneumoniae and E. coli showed high resistance rates for penicillins (ampicillin and ticarcillin), cephalosporins (ceftazidime, cefotaxime, ceftriaxone, and cefepime), followed by gentamicin and ciprofloxacin. Cefoxitin, aztreonam, imipenem and meropenem were nearly more effective against GNB isolates. On the other hand, about half isolates P. aeruginosa and all A. baumannii were resistant to the tested antibiotics. Extended Spectrum β- Lactamases (ESBLs) were released in 49 (70%) of the tested isolates of GNB. The multidrug resistant (MDR) pattern was noticed among 58 (82.85%) of GNB. Interestingly, ESBLs producing MDR isolates were determined in 34 (58.62%) of the studied GNB. Genotypic screening revealed that blaSHV and blaTEM genes were characterized in 20 (28.57%) and 32 (45.71%) of GNB isolates, respectively, while all GNB were negative for blaCTX-M. DNA sequencing of blaSHV amplicons revealed the occurrence of one point mutation (111T&gt;A) in six isolates of K. pneumoniae with a missense effect (p.31Q&gt;L) on the encoded protein. Forty-five GenBank accession numbers were recorded in NCBI to represent the studied variants of ESBLs. Phylogenetic analysis of both blaSHV and blaTEM displayed the possible epidemiologic routes of the tested GNB by detecting the origin host, source of collection, and geographic spread as compared with the reference sequences. Conclusion: high serum levels of both NO and ClO- indicated the potential role of these microbicidal agents to predict the serious infections of GNB in acute leukemia population. Phylogenetic analysis represented an important tool, possibly aiding in early control and prevention of pathogenic GNB.

NO production of granulocytes associated with antibacterial immune response in Tegillarca granosa

Nitric oxide (NO) is a signaling molecule and immune effector produced by the nitric oxide synthases (NOS), which involved to various physiological processes of animals. In marine bivalves, hemocytes play important roles in antimicrobial innate immune response. Although hemocyte-derived NO has been detected in several bivalves, the immune function of hemocyte-derived NO is not well understood. Here, we investigated the antibacterial response of hemocyte-derived NO in the blood clam Tegillarca granosa. Two types of hemocytes including erythrocytes and granulocytes were isolated by Percoll density gradient centrifugation, their NO production and TgNOS expression level were analyzed. The results showed that NO was mainly produced in granulocytes and almost no detected in erythrocytes. The granulocytes showed significantly higher NO level and TgNOS expression level than the erythrocytes. And the TgNOS expression level was significantly increased in granulocytes after Vibro parahemolyticus challenge. In addition, the NO donor sodium nitroprusside (SNP) significantly increased the NO production of hemocytes to kill pathogenic bacteria. In summary, the results revealed that granulocytes-derived NO play vital roles in the antimicrobial immune response of the blood clam.

Probing the mechanism of the dedicated NO sensor [4Fe-4S] NsrR: the effect of cluster ligand environment

NsrR from Streptomyces coelicolor is a bacterial nitric oxide (NO) sensor/nitrosative stress regulator as its primary function, and has been shown to have differential response at low, mid, and high levels of NO. These must correspond to discrete structural changes at the protein-bound [4Fe-4S] cluster in response to stepwise nitrosylation of the cluster. We have investigated the effect of the monohapto carboxylate ligand in the site differentiated [4Fe-4S] cluster cofactor of the protein NsrR on modulating its reactivity to NO with a focus on indentifying mechanistic intermediates. We have prepared a synthetic model [4Fe-4S] cluster complex with tripodal ligand and one single site differentiated site occupied by either thiolate or carboxylate ligand. We report here the mechanistic details of sequential steps of nitrosylation as observed by ESI MS and IR spectroscopy. Parallel non-denaturing mass spectrometry analyses were performed using site-differentiated variants of NsrR with the native aspartic acid, cysteine, or alanine in the position of the forth ligand to the cluster. A mono-nitrosylated synthetic [4Fe-4S] cluster was observed for the first time in a biologically-relevant thiolate-based coordination environment. Combined synthetic and protein data give unprecedented clarity in the modulation of nitrosylation of a [4Fe-4S] cluster.

Nitric oxide-induced endoplasmic reticulum stress of Schistosoma japonicum inhibits the worm development in rats

Schistosomiasis, caused by Schistosoma spp., is a zoonotic parasitic disease affecting human health. Rattus norvegicus (rats) are a non-permissive host of Schistosoma, in which the worms cannot mature and cause typical egg granuloma. We previously demonstrated that inherent high levels of nitric oxide (NO), produced by inducible NO synthase (iNOS), is a key molecule in blocking the development of S. japonicum in rats. To further explore the mechanism of NO inhibiting S. japonicum development in rats, we performed S-nitrosocysteine proteomics of S. japonicum collected from infected rats and mice. The results suggested that S. japonicum in rats may have undergone endoplasmic reticulum (ER) stress. Interestingly, we found that the ER of S. japonicum in rats showed marked damage, while the ER of the worm in iNOS−/− rats and mice were relatively normal. Moreover, the expression of ER stress markers in S. japonicum from WT rats was significantly increased, compared with S. japonicum from iNOS−/− rats and mice. Using the NO donor sodium nitroprusside in vitro, we demonstrated that NO could induce ER stress in S. japonicum in a dose-dependent manner, and the NO-induced ER stress in S. japonicum could be inhibited by ER stress inhibitor 4-Phenyl butyric acid. We further verified that inhibiting ER stress of S. japonicum in rats promoted parasite development and survival. Furthermore, we demonstrated that NO-induced ER stress of S. japonicum was related to the efflux of Ca2+ from ER and the impairment of mitochondrial function. Collectively, these findings show that high levels of NO in rats could induce ER stress in S. japonicum by promoting the efflux of Ca2+ from ER and damaging the mitochondrial function, which block the worm development. Thus, this study further clarifies the mechanism of anti-schistosome in rats and provides potential strategies for drug development against schistosomiasis and other parasitosis.

Association between Plasmodium Infection and Nitric Oxide Levels: A Systematic Review and Meta-Analysis.

Nitric oxide (NO) has been implicated in the pathology of malaria. This systematic review and meta-analysis describe the association between NO levels and malaria. Embase, Ovid, PubMed, Scopus, and Google Scholar were searched to identify studies evaluating NO levels in malaria patients and uninfected controls. Meta-regression and subgroup analyses were conducted to discern differences in NO levels between the groups. Of the 4517 records identified, 21 studies were included in the systematic review and meta-analysis. The findings illustrated significant disparities in NO levels based on geographic location and study time frames. Despite the fluctuations, such as higher NO levels in adults compared to children, no significant differences in mean NO levels between patients and uninfected controls (p = 0.25, Hedge's g: 0.35, 95% confidence interval (CI): -0.25-0.96, I2: 97.39%) or between severe and non-severe malaria cases (p = 0.09, Hedge's g: 0.71, 95% CI: -0.11-1.54, I2: 96.07%) were detected. The systematic review and meta-analysis highlighted inconsistencies in NO levels in malaria patients. Given the high heterogeneity of the results, further studies using standardized metrics for NO measurements and focusing on biochemical pathways dictating NO responses in malaria are imperative to understand the association between NO and malaria.

Nitric Oxide Detection Using a Chemical Trap Method for Applications in Bacterial Systems.

Plant growth-promoting bacteria (PGPB) can be incorporated in biofertilizer formulations, which promote plant growth in different ways, such as fixing nitrogen and producing phytohormones and nitric oxide (NO). NO is a free radical involved in the growth and defense responses of plants and bacteria. NO detection is vital for further investigation in different agronomically important bacteria. NO production in the presence of KNO3 was evaluated over 1-3 days using eight bacterial strains, quantified by the usual Griess reaction, and monitored by 2,3-diaminonaphthalene (DAN), yielding 2,3-naphthotriazole (NAT), as analyzed by fluorescence spectroscopy, gas chromatography-mass spectrometry, and high-performance liquid chromatography. The Greiss and trapping reaction results showed that Azospirillum brasilense (HM053 and FP2), Rhizobium tropici (Br322), and Gluconacetobacter diazotrophicus (Pal 5) produced the highest NO levels 24 h after inoculation, whereas Nitrospirillum amazonense (Y2) and Herbaspirillum seropedicae (SmR1) showed no NO production. In contrast to the literature, in NFbHP-NH4Cl-lactate culture medium with KNO3, NO trapping led to the recovery of a product with a molecular mass ion of 182 Da, namely, 1,2,3,4-naphthotetrazole (NTT), which contained one more nitrogen atom than the usual NAT product with 169 Da. This strategy allows monitoring and tracking NO production in potential biofertilizing bacteria, providing future opportunities to better understand the mechanisms of bacteria-plant interaction and also to manipulate the amount of NO that will sustain the PGPB.

Triterpenoid and Coumarin Isolated from Astilbe grandis with Anti-Inflammatory Effects through Inhibiting the NF-κB Pathway in LPS-Induced RAW264.7 Cells.

The roots of Astilbe grandis, known as "Ma sang gou bang", are used as a Miao traditional medicine with anti-inflammatory and analgesic properties. However, the active components and mechanism of action of this plant remain mostly uncharacterized. The aim of this study was to identify its active components and verify their pharmacological activity. The extract of A. grandis root was separated using various chromatographic methods. As a result, we obtained one novel triterpenoid, named astigranlactone (1), which has an unusual lactone moiety formed between C-7 and C-27. Additionally, a known coumarin compound, 11-O-galloyl bergenin (2) was isolated from this plant. The structures of these two compounds were elucidated by extensive NMR experiments in conjunction with HR-ESI-MS data. To the best of our knowledge, both compounds were isolated from this species for the first time. Moreover, we tested the anti-inflammation effect of the two compounds by establishing a cellular inflammation model induced by LPS in RAW264.7 cells. The effect of different concentrations of these compounds on the activity of RAW264.7 cells was assessed using a CCK8 assay. The levels of nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) in the supernatant of each group were evaluated using the Griess method and an enzyme-linked immunosorbent assay (ELISA). Western blot and quantitative real-time PCR (qRT-RCR) were used to measure the levels of cyclooxygenase 2 (COX-2) and nitric oxide synthase (iNOS) gene expression. Our findings revealed that these two compounds inhibited the high levels of NO, TNF-α, IL-6, IL-1β, COX-2, and iNOS (induced by LPS). Mechanistic studies demonstrated that these two compounds reduced the activation of the nuclear transcription factor-B (NF-κB) signaling pathway by inhibiting the phosphorylation of p65. Therefore, our study indicates that compounds 1 and 2 can exert a definite anti-inflammatory effect by inhibiting the NF-κB signaling pathway.