scholarly journals Promotion of Anti-Tuberculosis Macrophage Activity by L-Arginine in the Absence of Nitric Oxide

2021 ◽  
Vol 12 ◽  
Author(s):  
Melanie C. McKell ◽  
Rebecca R. Crowther ◽  
Stephanie M. Schmidt ◽  
Michelle C. Robillard ◽  
Rachel Cantrell ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Host Defense ◽  
Defense Mechanisms ◽  
No Production ◽  
Macrophage Function ◽  
Macrophage Activity ◽  
Extracellular Stimuli ◽  
Essential Nutrient ◽  
Inhibition Of Glycolysis

Macrophages are indispensable immune cells tasked at eliminating intracellular pathogens. Mycobacterium tuberculosis (Mtb), one of the most virulent intracellular bacterial pathogens known to man, infects and resides within macrophages. While macrophages can be provoked by extracellular stimuli to inhibit and kill Mtb bacilli, these host defense mechanisms can be blocked by limiting nutritional metabolites, such as amino acids. The amino acid L-arginine has been well described to enhance immune function, especially in the context of driving macrophage nitric oxide (NO) production in mice. In this study, we aimed to establish the necessity of L-arginine on anti-Mtb macrophage function independent of NO. Utilizing an in vitro system, we identified that macrophages relied on NO for only half of their L-arginine-mediated host defenses and this L-arginine-mediated defense in the absence of NO was associated with enhanced macrophage numbers and viability. Additionally, we observed macrophage glycolysis to be driven by both L-arginine and mechanistic target of rapamycin (mTOR), and inhibition of glycolysis or mTOR reduced macrophage control of Mtb as well as macrophage number and viability in the presence of L-arginine. Our data underscore L-arginine as an essential nutrient for macrophage function, not only by fueling anti-mycobacterial NO production, but also as a central regulator of macrophage metabolism and additional host defense mechanisms.

scholarly journals Human Monocytic U937 Cells KillSalmonella In Vitro by NO-Independent Mechanisms

1999 ◽  
Vol 67 (7) ◽  
pp. 3670-3673 ◽  
Author(s):  
Päivi Ekman ◽  
Marja Saarinen ◽  
Qiushui He ◽  
Mika Virtala ◽  
Marko Salmi ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Host Defense ◽  
No Production ◽  
U937 Cells ◽  
Hla B27

ABSTRACT Nitric oxide (NO) has a central role in host defense against intracellular microbes. HLA-B27 has been shown to directly modulate host-microbe interaction in vitro, leading to the impaired elimination of Salmonella in human monocytic U937 cells. Here, we studied whether impaired elimination of Salmonella would result from differences in NO production between HLA-B27- and HLA-A2-transfected U937 cells. Both human monocytic transfectants produced NO equally well and killed Salmonella via NO-independent mechanisms.

scholarly journals Effects of chitosan on nitric oxide production and inducible nitric oxide synthase activity and mRNA expression in weaned piglets

2018 ◽  
Vol 60 (No. 8) ◽  
pp. 359-366
Author(s):  
J. Li ◽  
B. Shi ◽  
S. Yan ◽  
L. Jin ◽  
Y. Guo ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Mrna Expression ◽  
Inducible Nitric Oxide Synthase ◽  
No Production ◽  
Weaned Piglets ◽  
Inos Activity ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

The effects of chitosan on nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) activity and gene expression in vivo or vitro were investigated in weaned piglets. In vivo, 180 weaned piglets were assigned to five dietary treatments with six replicates. The piglets were fed on a basal diet supplemented with 0 (control), 100, 500, 1000, and 2000 mg chitosan/kg feed, respectively. In vitro, the peripheral blood mononuclear cells (PBMCs) from a weaned piglet were cultured respectively with 0 (control), 40, 80, 160, and 320 µg chitosan/ml medium. Results showed that serum NO concentrations on days 14 and 28 and iNOS activity on day 28 were quadratically improved with increasing chitosan dose (P < 0.05). The iNOS mRNA expressions were linearly or quadratically enhanced in the duodenum on day 28, and were improved quadratically in the jejunum on days 14 and 28 and in the ileum on day 28 (P < 0.01). In vitro, the NO concentrations, iNOS activity, and mRNA expression in unstimulated PBMCs were quadratically enhanced by chitosan, but the improvement of NO concentrations and iNOS activity by chitosan were markedly inhibited by N-(3-[aminomethyl] benzyl) acetamidine (1400w) (P < 0.05). Moreover, the increase of NO concentrations, iNOS activity, and mRNA expression in PBMCs induced by lipopolysaccharide (LPS) were suppressed significantly by chitosan (P < 0.05). The results indicated that the NO concentrations, iNOS activity, and mRNA expression in piglets were increased by feeding chitosan in a dose-dependent manner. In addition, chitosan improved the NO production in unstimulated PBMCs but inhibited its production in LPS-induced cells, which exerted bidirectional regulatory effects on the NO production via modulated iNOS activity and mRNA expression.

scholarly journals New Cyclic Diarylheptanoids from Platycarya strobilacea

Molecules ◽  
2020 ◽  
Vol 25 (24) ◽  
pp. 6034
Author(s):  
Wen-bing Ding ◽  
Rui-yuan Zhao ◽  
Guan-hua Li ◽  
Bing-lei Liu ◽  
Hua-liang He ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Pc12 Cells ◽  
Stem Bark ◽  
No Production ◽  
Electronic Circular Dichroism ◽  
Pheochromocytoma Pc12 ◽  
Pheochromocytoma Pc12 Cells ◽  
Induced Apoptosis ◽  
Circular Dichroïsm

Five new cyclic diarylheptanoids (platycary A–E, compounds 1–5) and three previously identified analogues (i.e., phttyearynol (compound 6), myricatomentogenin (compound 7), and juglanin D (compound 8)) were isolated from the stem bark of Platycarya strobilacea. The structures of these compounds were determined using NMR, HRESIMS, and electronic circular dichroism (ECD) data. The cytotoxicity of compounds 1–5 and their ability to inhibit nitric oxide (NO) production, as well as protect against the corticosterone-induced apoptosis of Pheochromocytoma (PC12) cells, were evaluated in vitro using the appropriate bioassays. Compounds 1 and 2 significantly inhibited the corticosterone-induced apoptosis of PC12 cells at a concentration of 20 μΜ.

Acid and neutral sphingomyelinases: roles and mechanisms of regulation

2004 ◽  
Vol 82 (1) ◽  
pp. 27-44 ◽  
Author(s):  
Norma Marchesini ◽  
Yusuf A Hannun
Keyword(s):  
Nitric Oxide ◽  
Molecular Mechanisms ◽  
Caveolin 1 ◽  
Niemann Pick Disease ◽  
Extracellular Stimuli ◽  
Niemann Pick ◽  
Hydrolysis Of ◽  
Pick Disease

Ceramide, an emerging bioactive lipid and second messenger, is mainly generated by hydrolysis of sphingomyelin through the action of sphingomyelinases. At least two sphingomyelinases, neutral and acid sphingo myelinases, are activated in response to many extracellular stimuli. Despite extensive studies, the precise cellular function of each of these sphingomyelinases in sphingomyelin turnover and in the regulation of ceramide-mediated responses is not well understood. Therefore, it is essential to elucidate the factors and mechanisms that control the activation of acid and neutral sphingomyelinases to understand their the roles in cell regulation. This review will focus on the molecular mechanisms that regulate these enzymes in vivo and in vitro, especially the roles of oxidants (glu ta thi one, peroxide, nitric oxide), proteins (saposin, caveolin 1, caspases), and lipids (diacylglycerol, arachidonic acid, and ceramide).Key words: sphingomyelinase, ceramide, apoptosis, Niemann-Pick disease, FAN (factor associated with N-SMase activation).

scholarly journals A novel long intergenic non-coding RNA, Nostrill, regulates iNOS gene transcription and neurotoxicity in microglia

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Nicholas W. Mathy ◽  
Olivia Burleigh ◽  
Andrew Kochvar ◽  
Erin R. Whiteford ◽  
Matthew Behrens ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
No Production ◽  
Cortical Tissue ◽  
Non Coding Rna ◽  
Transcriptional Regulatory ◽  
Lncrna Locus ◽  
Rna Immunoprecipitation ◽  
Long Non Coding Rna

Abstract Background Microglia are resident immunocompetent and phagocytic cells in the CNS. Pro-inflammatory microglia, stimulated by microbial signals such as bacterial lipopolysaccharide (LPS), viral RNAs, or inflammatory cytokines, are neurotoxic and associated with pathogenesis of several neurodegenerative diseases. Long non-coding RNAs (lncRNA) are emerging as important tissue-specific regulatory molecules directing cell differentiation and functional states and may help direct proinflammatory responses of microglia. Characterization of lncRNAs upregulated in proinflammatory microglia, such as NR_126553 or 2500002B13Rik, now termed Nostrill (iNOS Transcriptional Regulatory Intergenic LncRNA Locus) increases our understanding of molecular mechanisms in CNS innate immunity. Methods Microglial gene expression array analyses and qRT-PCR were used to identify a novel long intergenic non-coding RNA, Nostrill, upregulated in LPS-stimulated microglial cell lines, LPS-stimulated primary microglia, and LPS-injected mouse cortical tissue. Silencing and overexpression studies, RNA immunoprecipitation, chromatin immunoprecipitation, chromatin isolation by RNA purification assays, and qRT-PCR were used to study the function of this long non-coding RNA in microglia. In vitro assays were used to examine the effects of silencing the novel long non-coding RNA in LPS-stimulated microglia on neurotoxicity. Results We report here characterization of intergenic lncRNA, NR_126553, or 2500002B13Rik now termed Nostrill (iNOS Transcriptional Regulatory Intergenic LncRNA Locus). Nostrill is induced by LPS stimulation in BV2 cells, primary murine microglia, and in cortical tissue of LPS-injected mice. Induction of Nostrill is NF-κB dependent and silencing of Nostrill decreased inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production in BV2 and primary microglial cells. Overexpression of Nostrill increased iNOS expression and NO production. RNA immunoprecipitation assays demonstrated that Nostrill is physically associated with NF-κB subunit p65 following LPS stimulation. Silencing of Nostrill significantly reduced NF-κB p65 and RNA polymerase II recruitment to the iNOS promoter and decreased H3K4me3 activating histone modifications at iNOS gene loci. In vitro studies demonstrated that silencing of Nostrill in microglia reduced LPS-stimulated microglial neurotoxicity. Conclusions Our data indicate a new regulatory role of the NF-κB-induced Nostrill and suggest that Nostrill acts as a co-activator of transcription of iNOS resulting in the production of nitric oxide by microglia through modulation of epigenetic chromatin remodeling. Nostrill may be a target for reducing the neurotoxicity associated with iNOS-mediated inflammatory processes in microglia during neurodegeneration.

scholarly journals Effect ofTityus serrulatusvenom on cytokine production and the activity of murine macrophages

2002 ◽  
Vol 11 (1) ◽  
pp. 23-31 ◽  
Author(s):  
Vera L. Petricevich
Keyword(s):  
Cytokine Production ◽  
Peritoneal Macrophages ◽  
Interferon Γ ◽  
Enzyme Linked Immunosorbent Assay ◽  
No Production ◽  
Dependent Manner ◽  
Immune Functions ◽  
Macrophage Activity ◽  
Ifn Γ

The purpose of this study was to investigate the effects ofTityus serrulatusvenom (TSV) on murine peritoneal macrophages evaluated in terms of activation. The effects of crude TSV were analysed by detection of cytokines, oxygen intermediate metabolites (H2O2) and nitric oxide (NO) in supernatants of peritoneal macrophages. Several functional bioassays were employed including anin vitromodel for envenomating: cytotoxicity of TSV was assessed using the lyses percentage. Tumor necrosis factor (TNF) activity was assayed by measuring its cytotoxic activity on L-929 cells, and interleukin-6 (IL-6) and interferon-γ (IFN-γ) were assayed by enzyme-linked immunosorbent assay, whereas NO levels were detected by Griess colorimetric reactions in culture supernatant of macrophages incubated with TSV and subsequently exposed to either lipopolysaccharide or IFN-γ. Incubation of macrophages with TSV increased production of IL-6 and IFN-γ in a dose-dependent manner. TNF production was not detected in supernatants treated with TSV at any concentration. The increase in IL-6 secretion was not associated with concentration-dependent cytoxicity of TSV on these cells. These data suggest that the cytotoxicity does not appear to be the main cause of an increased cytokine production by these cells. Although NO is an important effector molecule in macrophage microbicidal activity, the inducing potential of the test compounds for its release was found to be very moderate, ranging from 125 to 800 mM. Interestingly, NO levels of peritoneal macrophages were increased after IFN-γ. Moreover, NO production had an apparent effect on macrophage activity. The results obtained here also shown that the TSV induces an important elevation in H2O2release. These results combined with NO production suggest that TSV possesses significant immunomodulatory activities capable of stimulating immune functionsin vitro.

scholarly journals In Vitro Study of Nitric Oxide Metabolites Effects on Human Hydatid ofEchinococcus granulosus

2009 ◽  
Vol 2009 ◽  
pp. 1-7 ◽  
Author(s):  
Razika Zeghir-Bouteldja ◽  
Manel Amri ◽  
Saliha Aitaissa ◽  
Samia Bouaziz ◽  
Dalila Mezioug ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Hydatid Cyst ◽  
Echinococcus Granulosus ◽  
In Vitro Study ◽  
No Production ◽  
In Vitro Effects ◽  
Nitric Oxide Metabolites

Hydatidosis is characterized by the long-term coexistence of larvaEchinococcus granulosusand its host without effective rejection. Previous studies demonstrated nitric oxide (NO) production (in vivo and in vitro) during hydatidosis. In this study, we investigated the direct in vitro effects of NO species: nitrite (NO2−), nitrate (NO3−) and peroxynitrite (ONOO−) on protoscolices (PSCs) viability and hydatid cyst layers integrity for 24 hours and 48 hours. Our results showed protoscolicidal activity ofNO2−andONOO−24 hours and 3 hours after treatment with 320 μM and 80 μM respectively. Degenerative effects were observed on germinal and laminated layers. The comparison of the in vitro effects of NO species on the PSCs viability indicated thatONOO−is more cytotoxic thanNO2−. In contrast,NO3−has no effect. These results suggest possible involvement ofNO2−andONOO−in antihydatic action and point the efficacy of these metabolites as scolicidal agents.

Losartan Increases No Production from the Bovine Aortic Wall That is Stimulated by Angiotensin II

2003 ◽  
Vol 1 (3) ◽  
pp. 113-117 ◽  
Author(s):  
M. Myronidou ◽  
B. Kokkas ◽  
A. Kouyoumtzis ◽  
N. Gregoriadis ◽  
A. Lourbopoulos ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Angiotensin Ii ◽  
Aortic Wall ◽  
Vascular Wall ◽  
Protective Role ◽  
Normal Function ◽  
No Production ◽  
Chemical Inactivation

In these studies we investigated if losartan, an AT1- receptor blocker has any beneficial effect on NO production from the bovine aortic preparations in vitro while under stimulation from angiotensin II. Experiments were performed on intact specimens of bovine thoracic aorta, incubated in Dulbeco's MOD medium in a metabolic shaker for 24 hours under 95 % O2 and 5 % CO2 at a temperature of 37°C. We found that angiotensin II 1nM−10 μM does not exert any statistically significant action on NO production. On the contrary, angiotensin II 10nM increases the production of NO by 58.14 % (from 12.16 + 2.9 μm/l to 19.23 + 4.2 μm/l in the presence of losartan 1nM (P<0.05). Nitric oxide levels depend on both rate production and rate catabolism or chemical inactivation. Such an equilibrium is vital for the normal function of many systems including the cardiovascular one. The above results demonstrate that the blockade of AT1-receptors favors the biosynthesis of NO and indicate the protective role of losartan on the vascular wall.

scholarly journals Estimation of nitric oxide generation by endothelial cells EA.hy926 under flow-induced mechanical stress in a microfluidic system

2020 ◽  
pp. 71-77
Author(s):  
А.А. Московцев ◽  
А.Н. Мыльникова ◽  
Д.В. Колесов ◽  
А.А. Микрюкова ◽  
Д.М. Зайченко ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Mechanical Stress ◽  
Microfluidic System ◽  
Hydrodynamic Characteristics ◽  
No Production ◽  
Hydrodynamic Conditions ◽  
Cellular Mechanisms ◽  
Vascular Walls

Эндотелиальные клетки, выстилающие стенки сосудов, преобразовывают деформацию собственных структур, вызванную током крови, в химические сигналы, одним из которых является важный регулятор просвета сосуда - оксид азота (NO). К настоящему моменту накоплен большой объём данных о клеточных механизмах активации продукции NO, однако сведений о динамике генерации оксида азота эндотелиальными клетками в зависимости от гидродинамических условий недостаточно. В этой связи разработка микрофлюидных систем in vitro, имитирующих кровеносное русло, и изучение в них эндотелия в сложных гидродинамических условиях является актуальной задачей. В данной работе для создания контролируемых гидродинамических условий для монослоя эндотелиоцитоподобных клеток EA.hy926 была спроектирована и разработана микрофлюидная система, имитирующая линейные участки микрососудистого русла. Методом непрямого определения содержания оксида азота (II) NO с использованием флуоресцентного зонда 4,5-диаминофлуоресцеина DAF-2 впервые получены данные об увеличении продукции NO клетками EA.hy926 при механическом стрессе, создаваемом потоком ростовой среды. Представлены расчетные гидродинамические характеристики микрофлюидной системы, а также методика измерения продукции NO. Возможность исследования функциональной активности эндотелия позволяет использовать разработанную микрофлюидную модельную систему как для изучения клеточно-автономных регуляторных свойств эндотелия при действии ряда вазоактивных фармакологических препаратов и других методов воздействия на эндотелий, так и при моделируемой дисфункции эндотелия. Endothelial cells lining vascular walls transform the flow-induced deformation of their own structures into chemical signals, one of which, nitric oxide (NO), is an important regulator of the vascular lumen diameter. By present, a large amount of data on cellular mechanisms for activation of NO production has been accumulated. However, there is insufficient information on changes in endothelial NO generation under different hydrodynamic conditions. Therefore, development of microfluidic systems that model blood vessels in vitro and using them to study the endothelium under complex hydrodynamic conditions are relevant tasks. In this study, a microfluidic system was developed to create controlled hydrodynamic conditions for a monolayer of endotheliocyte-like cells EAhy.926. This system simulates linear sections of the microvasculature. By indirect measurement of NO (II) content with a fluorescent 4,5-diaminofluorescein (DAF-2) probe, we showed an increase in the NO production by EAhy.926 cells under mechanical stress generated by the medium flow. The article presents the method for measuring NO production and the calculated hydrodynamic characteristics of the microfluidic system. The results showed that the developed microfluidic model system is promising for studying cell-autonomous regulatory properties of the endothelium both under the action of vasoactive agents and in simulated endothelial dysfunction.

Determination of the enhancing action of HSP90 on neuronal nitric oxide synthase by EPR spectroscopy

2001 ◽  
Vol 281 (6) ◽  
pp. C1819-C1824 ◽  
Author(s):  
Yao Song ◽  
Jay L. Zweier ◽  
Yong Xia
Keyword(s):  
Nitric Oxide ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Heat Shock Protein 90 ◽  
Tryptophan Fluorescence ◽  
Hsp90 Inhibitor ◽  
Resonance Spectroscopy ◽  
No Production ◽  
Dependent Manner ◽  
Intact Cells

Recent studies showed that heat shock protein 90 (HSP90) enhances nitric oxide (NO) synthesis from endothelial and neuronal NO synthase (eNOS and nNOS, respectively). However, these findings were based on indirect NO measurements. Moreover, although our previous studies showed that the action of HSP90 involves increased Ca2+/calmodulin (Ca2+/CaM) binding, quantitative measurements of the effect of HSP90 on CaM binding to nNOS have been lacking. With electron paramagnetic resonance spectroscopy, we directly measured NO signals from purified nNOS. HSP90 augmented NO formation from nNOS in a dose-dependent manner. Tryptophan fluorescence-quenching measurements revealed that HSP90 markedly reduced the K d of CaM to nNOS (0.5 ± 0.1 nM vs. 9.4 ± 1.8 nM in the presence and absence of HSP90, P < 0.01). Ca2+ ionophore triggered strong NO production from nNOS-transfected cells, and this was significantly reduced by the HSP90 inhibitor geldanamycin. Thus these studies provide direct evidence demonstrating that HSP90 enhances nNOS catalytic function in vitro and in intact cells. The effect of HSP90 is mediated by the enhancement of CaM binding to nNOS.

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