Elevated Nitric Oxide Production Research Articles

Potential Role and Mechanism of Mulberry Extract in Immune Modulation: Focus on Chemical Compositions, Mechanistic Insights, and Extraction Techniques.

Mulberry is a rapidly growing plant that thrives in diverse climatic, topographical, and soil types, spanning temperature and temperate countries. Mulberry plants are valued as functional foods for their abundant chemical composition, serving as a significant reservoir of bioactive compounds like proteins, polysaccharides, phenolics, and flavonoids. Moreover, these compounds displayed potent antioxidant activity by scavenging free radicals, inhibiting reactive oxygen species generation, and restoring elevated nitric oxide production induced by LPS stimulation through the downregulation of inducible NO synthase expression. Active components like oxyresveratrol found in Morus demonstrated anti-inflammatory effects by inhibiting leukocyte migration through the MEK/ERK signaling pathway. Gallic and chlorogenic acids in mulberry leaves (ML) powder-modulated TNF, IL-6, and IRS1 proteins, improving various inflammatory conditions by immune system modulation. As we delve deeper into understanding its anti-inflammatory potential and how it works therapeutically, it is crucial to refine the extraction process to enhance the effectiveness of its bioactive elements. Recent advancements in extraction techniques, such as solid-liquid extraction, pressurized liquid extraction, superficial fluid extraction, microwave-assisted extraction, and ultrasonic-assisted extraction, are being explored. Among the extraction methods tested, including Soxhlet extraction, maceration, and ultrasound-assisted extraction (UAE), UAE demonstrated superior efficiency in extracting bioactive compounds from mulberry leaves. Overall, this comprehensive review sheds light on the potential of mulberry as a natural immunomodulatory agent and provides insights into its mechanisms of action for future research and therapeutic applications.

Selection of goat β-casein derived ACE-inhibitory peptide SQPK and insights into its effect and regulatory mechanism on the function of endothelial cells

The angiotensin I-converting enzyme (ACE)-inhibitory peptide SQPK was selected by in silico digestion and virtual screening from goat β-casein, and its effect and regulatory mechanism on function of endothelial cells was further evaluated. The results showed that SQPK exhibited relatively good ACE inhibition capacity (IC50 = 452.7 μg/mL). Treatment with 25 μg/mL SQPK for 12 h significantly elevated nitric oxide (NO) production, stimulated eNOS expression (p < 0.05) and affected the transcriptomic profiling of EA. Hy926 cells. In particular, SQPK stimulated the expression of genes encoding inflammatory cytokines (CXCL1/2 and IL6) but depressed encoding mesenchymal markers (FN1 and CNN3). Furthermore, SQPK modified the expression of genes involved in endothelial-to-mesenchymal transition (EndMT). Therefore, the selected peptide SQPK may exert potential protective effects on the function of endothelial cells by inhibiting the EndMT.

Counteracting Age-Related Netrin-1 Signaling Insufficiency Ameliorates Endothelial Cell Senescence and Angiogenesis Impairment.

Senescent cells that accumulate are regarded as promising therapeutic targets. However, senolytic therapy failed to achieve satisfactory results. We previously discovered that young human plasma improved vascular endothelial cell senescence, and UNC5B might be a novel intervention target. Netrin-1, as a natural ligand of UNC5B, plays roles in multiple age-related vascular disorders, but its involvement in aging is still unclear. Here, we observed a significant decrease in plasma Netrin-1 levels in old healthy subjects compared to the young. In vivo, adeno-associated-virus-mediated delivery of Netrin-1 into aged mice significantly improved functional recovery in a model of hindlimb ischemia, promoted angiogenesis in ischemic tissues, and activated the endothelial nitric oxide synthase. Furthermore, we revealed that low-dose Netrin-1 recombinant protein significantly reduced senescence-associated-β-galactosidase-positive cells, inhibited the P53 pathway, promoted cell migration, increased tubule formation, and elevated nitric oxide production in senescent endothelial cells. However, UNC5B inhibition blocked the pro-angiogenesis effect of low-dose Netrin-1 on senescent cells or aortic rings. In summary, this study depicts that modulating Netrin-1 signaling can result in improved vascular health and Netrin-1 may have therapeutic potential for age-related ischemic diseases.

Determining the potent immunostimulation potential arising from the heteropolysaccharide structure of a novel fucoidan, derived from Sargassum Zhangii.

A preliminary study was conducted of the chemical, structural properties and immunomodulatory activities of fucoidan isolated from Sargassum Zhangii (SZ). Sargassum Zhangii fucoidan (SZF) was determined to have a sulfate content of 19.74±0.01% (w/w) and an average molecular weight of 111.28kDa. SZF possessed a backbone structure of (1,4)-α-d-linked-galactose, (3,4)-α-l-fucose, (1,3)-α-d-linked-xylose, β-d-linked-mannose and a terminal (1,4)-α-d-linked-glucose. The main monosaccharide composition was determined as (w/w) 36.10% galactose, 20.13% fucose, 8.86% xylose, 7.36% glucose, 5.62% mannose, and 18.07% uronic acids, respectively. An immunostimulatory assay showed that SZF, compared to commercial fucoidans (Undaria pitnnaifida and Fucus vesiculosus sources), significantly elevated nitric oxide production via up-regulation of cyclooxygenase-2 and inducible nitric oxide synthase at both gene and protein levels. These results suggest that SZ has the potential to be a source of fucoidan with enhanced properties that may act as a useful ingredient for functional foods, nutritional supplements, and immune enhancers.

Obesity- and Sex-Related Disturbances in Arginine and Nitric Oxide Kinetics

Abstract Objectives Arginine (Arg) is considered a conditionally essential amino acid (AA) in the state of inflammation because of its role in nitric oxide (NO) production. NO is emerging as a regulator of energy metabolism and body composition. Although a growing interest exists in supplementation of Arg and citrulline (Cit) because of their potential anti-obesity effects, there is no consensus on the physiological effects of obesity on Arg metabolism and the role of sex. Methods 163 subjects were stratified based on their BMI (20–30 vs &amp;gt; 30 kg/m2) into normal-to-overweight (n = 83, age 49.5 [44.4, 54.5] y, BMI = 25.2 [24.36, 25.8] kg/m2) and obese (n = 80, age: 54.0 [5.07, 57.3] y, BMI = 38.2 [36.9, 39.6] kg/m2) groups. In the overnight fasted state, we collected from each subject arterialized blood for analysis of concentrations of plasma AA, C-reactive protein (CRP as a marker of systemic inflammation), and glucose. We subsequently administered a pulse of AA stable tracers containing Arg, Cit, and ornithine (Orn) and measured their whole-body productions (WBP), calculated their clearance, and the conversions of Arg to Cit (a marker of NO synthesis), Cit to Arg (de novo Arg production), and Arg to Orn (a surrogate of urea production). We measured body composition by DXA. Statistics by ANCOVA adjusted for age, sex, and lean body mass. Data are mean [95% CI]. Significance at P &amp;lt; 0.05. Results The obese subjects were characterized by higher CRP (P &amp;lt; 0.0001) but comparable fasting glucose levels (P = 0.0884) as the normal-to-overweight subjects. We found reduced plasma Cit concentration (P &amp;lt; 0.0001) in the obese group and upregulated whole-body clearance of Arg (P = 0.0002), Cit (P &amp;lt; 0.0001), and Orn (P = 0.0002) but no differences in their WBP. We observed elevated production of NO (P &amp;lt; 0.0001) in the obese but reduced de novo Arg synthesis (P = 0.0007). Females had lower plasma Arg and Orn concentrations, WBP, and clearance of Arg, Cit, and Orn than the males (p &amp;lt; 0.05) but comparable Cit concentration and NO production. Obese females had 3.5-fold higher NO production (P = 0.0020) than obese males concomitant with a 40% decrease in de novo Arg production (P = 0.0059). Conclusions Plasma Cit is lower in obesity despite enhanced NO production and is related to reduced de novo Arg synthesis. Arg might be considered an essential amino acid in obese women. Funding Sources None.

ALDH2 mutation results in excessive basal nitric oxide production and a delayed response to nitroglycerin

Abstract Background and purpose: Nitroglycerin tolerance is a common phenomenon in patients with coronary artery disease (CAD). Aldehyde dehydrogenase 2 (ALDH2) is the enzyme that metabolizes nitroglycerin to its active form nitric oxide (NO). Previous studies showed altered nitroglycerin in subjects with ALDH2 mutation, but the functional impact on endothelial cells is not fully understood. Methods: In the first step of in vitro experiments, we examined functional properties of induced pluripotent stem cell-derived CD144+ endothelial cells (iPSC-ECs) that expressed wildtype (WT) vs ALDH2+/− variant. In the second step of human studies, diameter of the left anterior descending (LAD) coronary artery was determined using angiography in 151 adult volunteers (111 with WT ALDH2, 32 with ALDH2+/− and 8 with ALDH2−/− genotype) prior to as well as after intracoronary injection of 200-μg nitroglycerin. Results: Briefly, the ALDH2+/− iPSC-ECs demonstrated impaired low-density lipoprotein (LDL) uptake, proliferation, migration, tube formation, oxidative stress resistance, and viability. In comparison to the WT control, the ALDH2+/− iPSC-ECs had elevated NO production under baseline conditions, but exhibited a delayed NO release after nitroglycerin treatment. Exposure to 10-μg/mL nitroglycerin for 2 h increased NO production in WT iPSC-ECs but 4-h exposure was required to stimulate NO production in the ALDH2+/− iPSC-ECs. In comparison to the WT control, the subjects carrying the ALDH2+/− variants had seemingly larger LAD coronary artery diameter (3.5 and 3.8 mm vs 3.4 mm in the WT control), but attenuated vasodilatory response to nitroglycerin (ALDH2MUT group vs the WT control, 7.1 ± 0.6% vs 10.1 ± 0.8%, P = 0.024). Conclusion: These findings indicated elevated NO production by endothelia cells under basal conditions but attenuated response to nitroglycerin upon ALDH2 mutation.

Arginine deprivation alters microglial polarity and synergizes with radiation to eradicate non-arginine-auxotrophic glioblastoma tumors.

New approaches for the management of glioblastoma (GBM) are an urgent and unmet clinical need. Here, we illustrate that the efficacy of radiotherapy for GBM is strikingly potentiated by concomitant therapy with the arginine-depleting agent ADI-PEG20 in a non-arginine-auxotrophic cellular background (argininosuccinate synthetase 1 positive). Moreover, this combination led to durable and complete radiological and pathological response, with extended disease-free survival in an orthotopic immune-competent model of GBM, with no significant toxicity. ADI-PEG20 not only enhanced the cellular sensitivity of argininosuccinate synthetase 1–positive GBM to ionizing radiation by elevated production of nitric oxide (˙NO) and hence generation of cytotoxic peroxynitrites, but also promoted glioma-associated macrophage/microglial infiltration into tumors and turned their classical antiinflammatory (protumor) phenotype into a proinflammatory (antitumor) phenotype. Our results provide an effective, well-tolerated, and simple strategy to improve GBM treatment that merits consideration for early evaluation in clinical trials.

Nitric Oxide on Pathophysiology of SARS-CoV 19: Toward Possible Role of Acupuncture Treatment

The ongoing outbreak of COVID-19 has quickly become a daunting challenge to global health. In the absence of satisfied therapy, effective treatment interventions are urgently needed. Previous studies have demonstrated that acupuncture is effective at relieving common symptoms of COVID-19 including breathlessness, nausea, insomnia, leukopenia, fatigue, vomiting, and abdominal pain. Experiments have shown that nitric oxide (NO) inhibits the replication cycle of severe acute respiratory syndrome (SARS) coronavirus with similar structures of COVID-19. Increase in level of NO by using NO gas inhalation has been shown to restore lung function by reducing airway resistance and improving virus-induced lung infections in SARS patients. Recent case report showed that a medical acupuncturist with symptoms consistent with severe COVID pneumonia achieved full recovery by self-administered medical acupuncture and cupping therapy at home. Clinical features and pathophysiology demonstrated that NO deficiency and endothelial dysfunction contribute to the development of COVID-19. Several studies from different groups consistently demonstrated that acupuncture increases NO synthase expression and induces an elevation of NO production and release in plasma and the local skin regions in both animals and humans. It is suggested that exogenous NO supplies or interventions that induce increasing levels of NO can play an important role in protective effects against inflammation and acute lung injury. This article reviews the rationale for mechanisms of NO induction induced by acupuncture in the possible treatment of COVID-19 and highlights its potential for contributing to better clinical outcomes and improving future clinical studies of acupuncture on treatment of COVID-19.

Nitric Oxide Dependent Metabolism Regulates the Proliferation and Differentiation of Stress Erythroid Progenitors

Infection and tissue damage induce inflammation, which increases myelopoiesis at the expense of steady state erythropoiesis. Stress erythropoiesis is induced to compensate for the loss of erythroid output until the inflammation is resolved and bone marrow erythropoiesis can resume. Steady state erythropoiesis constantly produces erythrocytes, while stress erythropoiesis generates a bolus of new erythrocytes through the rapid expansion of immature progenitor cells which is followed by the synchronous differentiation of progenitors. We hypothesized that the proliferation of early progenitor cells and their transition to differentiation is regulated by changes in metabolism.Metabolomics and isotope tracing analysis was performed to assess the intracellular metabolic profiles in proliferating progenitors isolated from in vitro stress erythropoiesis cultures. We observed an active engagement of glucose metabolism in glycolysis and anabolic biosynthesis, while the levels of TCA intermediates suggested that TCA cycle and mitochondrial respiration were blocked. Concomitantly, inducible nitric oxide synthase (iNOS) was induced in progenitor cells to increase the production of nitric oxide (NO), which was demonstrated to be crucial for proliferating progenitor metabolism. Inhibition or genetic mutation of iNOS decreased NO levels resulting in the suppression of progenitor proliferation in vitro and in vivo. As evaluated by RNA-seq, inhibition of iNOS suppressed cell proliferation-related pathways including cell cycle and nucleotide metabolism, while upregulating erythroid differentiation genes. These data suggest that iNOS-derived NO production establishes a metabolism that promotes the proliferation of progenitor cells while inhibiting their differentiation.Notably, proliferating progenitor cells displayed low levels of the metabolite itaconate and decreased expression of Immunoresponsive gene 1 (Irg1), the enzyme that catalyzes itaconate synthesis from cis-aconitate. Further analysis showed that the addition of 4-Octyl itaconate (OI), a cell-permeable itaconate derivative, inhibited iNOS-derived NO production by activating nuclear factor erythroid 2-related factor 2 (Nrf2), which in turn impaired progenitor expansion. These results indicate that itaconate production is inhibited to enable the accumulation of NO and the NO dependent metabolism required for progenitor cell proliferation during the initial expansion stage of stress erythropoiesis.In contrast, the transition to differentiation is marked by elevated itaconate synthesis, Nrf2 activation, and attenuated iNOS expression. We hypothesized that the inhibition of NO production alters metabolism and in concert with new cell signaling removes the NO-dependent inhibition of erythroid program, which allows the differentiation of progenitor cells. We tested this mechanism by examining the effects of iNOS inhibitors and mutants in iNOS, Irg1 and Nrf2 on progenitor cells isolated from differentiation cultures. iNOS deficiency led to the activation of erythroid transcriptional program, and increased numbers of mature progenitors as well as stress BFU-Es. In contrast, Irg1 and Nrf2 mutants showed impaired transition to erythroid differentiation, while they had elevated iNOS expression and NO production. Further analysis showed that treatment with either OI or iNOS inhibitor inhibited NO production in Irg1 and Nrf2 deficient progenitors, and consequently rescued the defects in erythroid differentiation.These data support a model in which inflammation inhibits steady state erythropoiesis, while at the same time promoting stress erythropoiesis to maintain homeostasis. Our work reveals a dynamic and tight coordination between pro-inflammatory signals and progenitor cell metabolism in regulating the proliferation and differentiation of stress erythroid progenitors, and highlights the therapeutic potential of targeting metabolic and inflammatory signaling pathways in inflammatory anemia. DisclosuresPaulson: Forma Therapeutics: Consultancy.

Embryonic exposure to low concentrations of aflatoxin B1 triggers global transcriptomic changes, defective yolk lipid mobilization, abnormal gastrointestinal tract development and inflammation in zebrafish

Aflatoxin B1-contaminated feeds and foods induce various health problems in domesticated animals and humans, including tumor development and hepatotoxicity. Aflatoxin B1 also has embryotoxic effects in different livestock species and humans. However, it is difficult to distinguish between the indirect, maternally-mediated toxic effects and the direct embryotoxicity of aflatoxin B1 in mammals. In the present study, we investigated the aflatoxin B1-induced direct embryotoxic effects in a zebrafish embryo model system combining toxicological, transcriptomic, immunological, and biochemical approaches. Embryonic exposure to aflatoxin B1 induced significant changes at the transcriptome level resulting in elevated expression of inflammatory gene network and repression of lipid metabolism and gastrointestinal tract development-related gene sets. According to the gene expression changes, massive neutrophil granulocyte influx, elevated nitric oxide production, and yolk lipid accumulation were observed in the abdominal region of aflatoxin B1-exposed larvae. In parallel, aflatoxin B1-induced defective gastrointestinal tract development and reduced L-arginine level were found in our model system. Our results revealed the complex direct embryotoxic effects of aflatoxin B1, including inhibited lipid utilization, defective intestinal development, and inflammation.

Antioxidant and Anti-Inflammatory Effects of White Mulberry (Morus alba L.) Fruits on Lipopolysaccharide-Stimulated RAW 264.7 Macrophages.

In this study, the protective effects of white mulberry (Morus alba) fruits on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were investigated. The ethanol (EtOH) extract of white mulberry fruits and its derived fractions contained adequate total phenolic and flavonoid contents, with good in vitro antioxidant radical scavenging activity. The extract and fractions also markedly inhibited ROS generation and antioxidant activity. After treatment with the EtOH extract and its fractions, LPS stimulation-induced elevated nitric oxide (NO) production was restored, which was primarily mediated by downregulation of inducible NO synthase expression. A total of 20 chemical constituents including flavonoids, steroids, and phenolics were identified in the fractions using ultra-high-performance liquid chromatography (UHPLC)-quadrupole time-of-flight (QTOF) high-resolution mass spectrometry (HRMS). These findings provide experimental evidence of the protective effects of white mulberry fruit extract against oxidative stress and inflammatory responses, suggesting their nutraceutical and pharmaceutical potential as natural antioxidant and anti-inflammatory agents.

ГИПОТАЛАМИЧЕСКИЕ МЕХАНИЗМЫ РЕГУЛЯЦИИ СТАРЕНИЯ

The hypothalamus is the most important integrator of functions of the autonomous, endocrine, somatic nervous systems; it is involved in the regulation of rhythmic processes and systemic aging. Putative hypothalamic nuclei responsible for the implementation of the developmental program of aging include the suprachiasmatic nucleus and the median group of nuclei, including the arcuate, ventromedial, and dorsomedial nuclei. Hypothalamic aging is accompanied by an excitation-inhibition imbalance due to dysregulation of the GABA-ergic system, decreased frequency of neuronal firing, impaired calcium metabolism, elevated nitric oxide production, decreased autophagy, and disturbed intracellular signaling, in particular Sirt1, mTOR, NF-κB, AMPK, P53, and FoxO. Also, an important role in hypothalamic aging belongs to specialized ependymal cells, tanycytes, as manifested in the disruption of tanycyte-neuron relationships and impaired barrier function of these cells. Hypothalamic neural stem cells, located mainly in the region of the middle hypothalamic nuclei, secrete exosomal microRNA also responsible for anti-aging effects of stem cells. The pool of hypothalamic neural stem cells, as well as microRNA production therein, decline with age.

Crosstalk between hydrogen peroxide and nitric oxide mediates priming‐induced drought tolerance in wheat

AbstractDrought is a limiting factor for wheat production. Nitric oxide (NO) and hydrogen peroxide (H2O2) are key signalling molecules produced in response to drought stress. Drought priming is a promising strategy for enhancing drought tolerance. Here, the roles of NO and H2O2 in drought priming were investigated. The results showed that the scavengers of NO or H2O2 reduced the effects of priming‐induced tolerance to drought stress during seedling and grain filling stages. Drought priming induced the production of H2O2 and NO, suggesting that H2O2 and NO were involved in priming‐induced drought tolerance and that the synthesis of H2O2 could be related to respiratory burst oxidase homologues (RBOH). Further, nitrate reductase (NR) activity was not the unique source for NO production. The elevated production of NO in the primed plants was blocked by scavengers of both H2O2 and NO, whereas the H2O2 concentration was not affected by a scavenger of NO. Osmolyte accumulation induced by drought priming was partly inhibited by the removal of NO, but was higher than in non‐primed plants. NO worked downstream of H2O2‐mediated the accumulation of proline and glycine‐betaine in primed plants under drought stress during grain filling. The results increase our understanding of the roles of H2O2 and NO in priming‐induced drought tolerance in wheat.

The Effect of Sex Differences on Endothelial Function and Circulating Endothelial Progenitor Cells in Hypertriglyceridemia.

Background Men have a higher risk and earlier onset of cardiovascular diseases compared with premenopausal women. Hypertriglyceridemia is an independent risk factor for the occurrence of ischemic heart disease. Endothelial dysfunction is related to the development of ischemic heart disease. Whether sex differences will affect the circulating endothelial progenitor cells (EPCs) and endothelial function in hypertriglyceridemia patients or not is not clear. Methods Forty premenopausal women and forty age- and body mass index (BMI)-matched men without cardiovascular and metabolic disease were recruited and then divided into four groups: normotriglyceridemic women (women with serum triglycerides level <150 mg/dl), hypertriglyceridemic women (women with serum triglycerides level ≥150 mg/dl), normotriglyceridemic men (men with serum triglycerides level <150 mg/dl), and hypertriglyceridemic men (men with serum triglycerides level ≥150 mg/dl). Peripheral blood was obtained and evaluated. Flow-mediated dilatation (FMD), the number and activity of circulating EPCs, and the levels of nitric oxide (NO), vascular endothelial growth factor (VEGF), and granulocyte-macrophage colony-stimulating factor (GM-CSF) in plasma and culture medium were measured. Results The number and activity of circulating EPCs, as well as the level of NO in plasma or culture medium, were remarkably increased in premenopausal females compared with those in males both in the hypertriglyceridemic group and the normotriglyceridemic group. The EPC counts and activity, as well as the production of NO, were restored in hypertriglyceridemic premenopausal women compared with those in normal women. However, in hypertriglyceridemic men, the EPC counts and activity, as well as levels of NO, were significantly reduced. The values of VEGF and GM-CSF were without statistical change. Conclusions The present study firstly demonstrated that there were sex differences in the number and activity of circulating EPCs in hyperglyceridemia patients. Hypertriglyceridemic premenopausal women displayed restored endothelial functions, with elevated NO production, probably mediated by estradiol. We provided a new insight to explore the clinical biomarkers and therapeutic strategies for hypertriglyceridemia-related vascular damage.

A role for the Sts phosphatases in negatively regulating IFNγ‐mediated production of nitric oxide in monocytes

IntroductionThe atypical Sts phosphatases negatively regulate signaling pathways in diverse immune cell types, with two of their molecular targets being the related kinases Syk and Zap‐70. Mice lacking Sts expression (Sts −/−) are resistant to infection by the live vaccine strain (LVS) of Francisella tularensis. Although the mechanisms underlying the enhanced resistance of Sts −/− mice have not been definitively established, Sts −/− bone marrow‐derived monocytes (BMMs) demonstrate greater clearance of intracellular LVS following ex vivo infection, relative to wild type cells. To determine how the Sts proteins regulate monocyte bactericidal properties, we analyzed responses of infected cells.MethodsMonocyte bacterial clearance was assayed using ex vivo coculture infections followed by colony‐forming unit analysis of intracellular bacteria. Levels of gene expression were quantified by quantitative reverse‐transcription polymerase chain reaction, levels of Nos2 protein levels were quantified by Western blot analysis, and levels of nitric oxide (NO) were quantified directly using the Griess reagent. We characterized monocyte cytokine production via enzyme‐linked immunosorbent assay.ResultsWe demonstrate that Sts −/− monocyte cultures produce elevated levels of interferon‐γ (IFNγ) after infection, relative to wild type cultures. Sts −/− monocytes also demonstrate heightened responsiveness to IFNγ. Specifically, Sts −/− monocytes produce elevated levels of antimicrobial NO following IFNγ stimulation, and this NO plays an important role in LVS restriction. Additional IFNγ‐stimulated genes, including Ip10 and members of the Gbp gene family, also display heightened upregulation in Sts −/− cells. Both Sts‐1 and Sts‐2 contribute to the regulation of NO production, as evidenced by the responses of monocytes lacking each phosphatase individually. Finally, we demonstrate that the elevated production of IFNγ‐induced NO in Sts −/− monocytes is abrogated following chemical inhibition of Syk kinase.ConclusionOur results indicate a novel role for the Sts enzymes in regulating monocyte antibacterial responses downstream of IFNγ.

β-Cyclodextrin Inhibits Monocytic Adhesion to Endothelial Cells through Nitric Oxide-Mediated Depletion of Cell Adhesion Molecules.

Cyclodextrins (CDs) are used as drug delivery agents. In this study, we examined whether CDs have an inflammatory effect on endothelial cells. First, we found that β-CD promoted cell proliferation in bovine aortic endothelial cells and elevated nitric oxide (NO) production through dephosphorylation of threonine-495 (T-495) in endothelial nitric oxide synthetase (eNOS). Dephosphorylation of T-495 is known to activate eNOS. Phosphorylation of T-495 was found to be catalyzed by protein kinase Cε (PKCε). We then found that β-CD inhibits binding of PKCε to diacylglycerol (DAG) via formation of a β-CD-DAG complex, indicating that β-CD inactivates PKCε. Furthermore, β-CD controls activation of PKCε by reducing the recruitment of PKCε into the plasma membrane. Finally, β-CD inhibits expression of intercellular and vascular cell adhesion molecule-1 by increasing NO via control of PKCε/eNOS and suppression of THP-1 cell adhesion to endothelial cells. These findings imply that β-CD plays an important role in anti-inflammatory processes.

Biological effects of chronic and acute exposure of human endothelial cell line EA.hy926 to bisphenol A: New tricks from an old dog

Although epidemiological and animal studies suggest a possible correlation between bisphenol A (BPA) exposure and atherosclerosis, very few in vitro mechanistic and functional studies regarding the effect of BPA on vascular cells have been conducted. Here, we applied a “real-life” exposure scenario by continuously exposing human endothelial cell (EC) line EA.hy926 to environmentally relevant concentrations of BPA (10−9, 10−8, and 10−7 M) during 14 weeks. We also exposed EA.hy926 cells to higher concentrations of BPA (10−7, 10−6, and 10−5 M) for up to 48 h to gain mechanistic insight into the BPA’s action in ECs. Chronic exposure to BPA produced some unexpected effects in EA.hy926 cells including a transient decrease in the adhesion of monocytes to the EC monolayer and decrease in the expression of cellular adhesion molecules, improvement in endothelial barrier function and elevated expression of tight junction proteins occludin and zonula occludens-1 (ZO-1), increased adhesion of ECs, and increased nitric oxide (NO) production. Some of these effects, such as diminished adhesion of monocytes to the EC monolayer and elevated NO production have also been replicated during acute exposure experiments. Using Western blotting and specific pharmacological inhibitors in the acute study, we have shown that direct BPA’s action in EA.hy926 cells involves activation of estrogen receptor (ER), phosphorylation of protein kinase B (PKB/Akt) and endothelial nitric oxide synthase (eNOS)-mediated production of NO. Collectively, these data indicate that BPA induces functional and molecular changes in EA.hy926 cells associated with the promotion of endothelial integrity through activation of the ER/Akt/eNOS pathway.

Characterization, pro-inflammatory response and cytotoxic profile of bioaerosols from urban and rural residential settings in Pune, India

Microbiota associated with airborne particulate matter (PM) is an important indicator of indoor pollution as they can be pathogenic and cause serious health threats to the exposed occupants. Present study aimed to investigate the level of culturable microbes associated with PM and their toxicological characterization in urban and rural houses of Pune city. Highest concentration of bacterial aerosols observed to be associated with PM10 size fraction in urban site (2136 ± 285 CFU/m3) whereas maximum fungal concentration has been measured in rural houses (1521 ± 302 CFU/m3). Predominantly found bacterial species were Bacillus sp., S. aureus, and Pseudomonas aeruginosa and fungal species were Aspergillus sp., Cladosporium sp., and Penicillium sp. in both urban and rural residential premises. Concentration of endotoxin measured using the kinetic Limulus Amebocyte Lysate assay exhibited that the level of endotoxin in both urban and rural sites are associated with household characteristics and the activities performed in indoor as well as outdoor. Cell free DTT assay confirmed the ability of these airborne microbes to induce the production of reactive oxygen species (ROS) varying along with the types of microorganisms. On exposure of A549 cells to airborne microbes, a significant decrease in cell viability was observed in terms of both necrosis and apoptosis pathway. Elevated production of nitric oxide (NO) and proinflammatory cytokines in epithelial cells and macrophages clearly suggest the inflammatory nature of these airborne microbes. Results derived from the present study demonstrated that the indoor air of urban and rural houses of Pune is contaminated in terms of microbial load. Therefore, attention should be paid to control the factors favoring the microbial growth in order to safeguard the health of exposed inhabitants.

Silver nanoparticles restrict microbial growth by promoting oxidative stress and DNA damage.

Bacterial infections remain a serious health issue; hence there is a need for continuous search for improved antimicrobials. In addition, it is important to understand the antibacterial mechanism of prospective antimicrobials to fully harness their benefits. In this study, the antimicrobial action of silver nanoparticles was investigated. The antimicrobial potential of silver nanoparticles against different strains of bacteria was evaluated after which Escherichia coli and Staphylococcus aureus were selected as model for gram-negative and gram-positive bacteria respectively. Additionally, to determine mechanism of action, some biochemical assays including determination of kynurenine level, DNA fragmentation, lipid peroxidation and antioxidant status were carried out. Results showed that silver nanoparticles caused DNA damage and induced oxidative stress as reflected in elevated nitric oxide production and lipid peroxidation level. In contrast silver nanoparticles increased the antioxidant capacity viz-a-viz, elevated levels of total thiol, superoxide dismutase (SOD), and total antioxidant capacity (TAC) compared to untreated cells. They also initiated inconsistent alteration to the kynurenine pathway. Taken together, the findings indicate that silver nanoparticles exhibited antimicrobial action through the promotion of oxidative stress.

Sasa quelpaertensis Nakai extract induces p53-independent apoptosis via the elevation of nitric oxide production in human HCT116 colon cancer cells.

Induction of apoptosis in human cancer cells by Sasa quelpaertensis Nakai has been considered to be a potential therapeutic target for cancer treatment; however, the underlying mechanisms of action are not well understood. The present study investigated the role of nitric oxide (NO•) and inhibitors of apoptosis (IAPs) during apoptosis induced by Sasa quelpaertensis Nakai extracts (SQE) in p53-wild type (WT) and p53-null HCT116 colon carcinoma cells. Trypan blue exclusion and Annexin V/propidium iodide assays were used to test for antiproliferation, and apoptosis and cell cycle. Griess and reverse transcription-polymerase chain reaction and western blotting assays were carried out to assay NO• production, and to detect the mRNA and protein levels of Bcl-2, PARP and IAPs. A colorimetric assay was utilized to measure the time-dependent increase in caspase-3 activity. SQE inhibited cell growth and promoted apoptosis by the elevation of NO• in a dose- and time-dependent manner. In addition, both cell types underwent a reduction in mRNA and protein levels of IAPs (survivin, CIAP-1 and -2, and X-linked inhibitor of apoptosis) as well as anti-apoptotic Bcl-2, whereas an increase in protein expression of poly (ADP-ribose) polymerase 1 and caspase 3 activity was observed; however, an equivalent cytotoxic and apoptotic effect by SQE was observed in p53-WT and p53-null cells. Collectively, the results indicated that SQE-induced apoptosis was independent of p53 status and associated with modulation of endogenous NO• and IAP family gene expression.