Selenium deficiency increases the expression of inducible nitric oxide synthase in RAW 264.7 macrophages: role of nuclear factor-κB in up-regulation

The inducible isoform of nitric oxide synthase (iNOS) is implicated in atherosclerosis, malignancy, rheumatoid arthritis, tissue and reperfusion injuries. A key determinant of the pro-oxidant versus protective effects of NO is the underlying redox status of the tissue. Selenoproteins, such as glutathione peroxidases (GPxs) and thioredoxin reductases, are key components of cellular defence and promote optimal antioxidant/oxidant balance. In this study, we have investigated the relationship between Se status, iNOS expression and NO production in Se-deficient and Se-supplemented RAW 264.7 macrophage cell lines. The cellular GPx activity, a measure of Se status, was 17-fold lower in Se-deficient RAW 264.7 cells and the total cellular oxidative tone, as assessed by flow cytometry with 2′,7′-dichlorodihydrofluorescein diacetate, was higher in the Se-deficient cells than the Se-supplemented cells. Upon lipopolysaccharide (LPS) stimulation of these cells in culture, we found significantly higher iNOS transcript and protein expression levels with an increase in NO production in Se-deficient RAW 264.7 cells than the Se-supplemented cells. Electrophoretic mobility-shift assays, nuclear factor-κB (NF-κB)—luciferase reporter assays and Western blot analyses indicate that the increased expression of iNOS in Se deficiency could be due to an increased activation and consequent nuclear localization of the redox-sensitive transcription factor NF-κB. These results suggest an inverse relationship between cellular Se status and iNOS expression in LPS-stimulated RAW 264.7 cells and provide evidence for the beneficial effects of dietary Se supplementation in the prevention and/or treatment of oxidative-stress-mediated inflammatory diseases.

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Inhibition of inducible nitric oxide synthase and cyclooxygenase II by Platycodon grandiflorum saponins via suppression of nuclear factor-κB activation in RAW 264.7 cells

Life Sciences ◽

10.1016/j.lfs.2004.10.042 ◽

2005 ◽

Vol 76 (20) ◽

pp. 2315-2328 ◽

Cited By ~ 208

Author(s):

Kwang Seok Ahn ◽

Eun Jung Noh ◽

Hai Lin Zhao ◽

Sang Hoon Jung ◽

Sam Sik Kang ◽

...

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Inducible Nitric Oxide Synthase ◽

Nuclear Factor Κb ◽

Raw 264.7 Cells ◽

Raw 264.7 ◽

Nuclear Factor ◽

Platycodon Grandiflorum ◽

Oxide Synthase ◽

Inducible Nitric Oxide

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Ikarisoside A inhibits inducible nitric oxide synthase in lipopolysaccharide-stimulated RAW 264.7 cells via p38 kinase and nuclear factor-κB signaling pathways

European Journal of Pharmacology ◽

10.1016/j.ejphar.2008.09.032 ◽

2008 ◽

Vol 601 (1-3) ◽

pp. 171-178 ◽

Cited By ~ 43

Author(s):

Hwa Jung Choi ◽

Jae-Soon Eun ◽

Young-Ran Park ◽

Dae Keun Kim ◽

Rihua Li ◽

...

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Inducible Nitric Oxide Synthase ◽

Nuclear Factor Κb ◽

Raw 264.7 Cells ◽

Raw 264.7 ◽

Nuclear Factor ◽

P38 Kinase ◽

Oxide Synthase ◽

Inducible Nitric Oxide

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Activation of Nuclear Factor κB and Induction of Inducible Nitric Oxide Synthase by Ureaplasma urealyticumin Macrophages

Infection and Immunity ◽

10.1128/iai.68.12.7087-7093.2000 ◽

2000 ◽

Vol 68 (12) ◽

pp. 7087-7093 ◽

Cited By ~ 28

Author(s):

Y.-H. Li ◽

Z.-Q. Yan ◽

J. Skov Jensen ◽

K. Tullus ◽

A. Brauner

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Inducible Nitric Oxide Synthase ◽

Alveolar Macrophages ◽

Nuclear Factor Κb ◽

Inos Expression ◽

Dependent Manner ◽

Nuclear Factor ◽

Oxide Synthase ◽

Inducible Nitric Oxide

ABSTRACT Chronic lung disease (CLD) of prematurity is an inflammatory disease with a multifactorial etiology. The importance ofUreaplasma urealyticum in the development of CLD is debated, and steroids produce some improvement in neonates with this disease. In the present study, the capability of U. urealyticum to stimulate rat alveolar macrophages to produce nitric oxide (NO), express inducible nitric oxide synthase (iNOS), and activate nuclear factor κB (NF-κB) in vitro was characterized. The effect of NO on the growth of U. urealyticum was also investigated. In addition, the impact of dexamethasone and budesonide on these processes was examined. We found that U. urealyticum antigen (≥4 × 107 color-changing units/ml) stimulated alveolar macrophages to produce NO in a dose- and time-dependent manner (P < 0.05). This effect was further enhanced by gamma interferon (100 IU/ml; P < 0.05) but was attenuated by budesonide and dexamethasone (10−4 to 10−6 M) (P < 0.05). The mRNA and protein levels of iNOS were also induced in response to U. urealyticum and inhibited by steroids.U. urealyticum antigen triggered NF-κB activation, a possible mechanism for the induced iNOS expression, which also was inhibited by steroids. NO induced by U. urealyticum caused a sixfold reduction of its own growth after infection for 10 h. Our findings imply that U. urealyticum may be an important factor in the development of CLD. The host defense response againstU. urealyticum infection may also be influenced by NO. The down-regulatory effect of steroids on NF-κB activation, iNOS expression, and NO production might partly explain the beneficial effect of steroids in neonates with CLD.

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Inhibitory Effect of 1,5-Dimethyl Citrate from Sea Buckthorn (Hippophae rhamnoides) on Lipopolysaccharide-Induced Inflammatory Response in RAW 264.7 Mouse Macrophages

Foods ◽

10.3390/foods9030269 ◽

2020 ◽

Vol 9 (3) ◽

pp. 269 ◽

Cited By ~ 1

Author(s):

Su Cheol Baek ◽

Dahae Lee ◽

Mun Seok Jo ◽

Kwang Ho Lee ◽

Yong Hoon Lee ◽

...

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Sea Buckthorn ◽

Raw 264.7 Cells ◽

No Production ◽

Raw 264.7 ◽

Tnf Α ◽

Mouse Macrophages ◽

Cox 2 ◽

Oxide Synthase

Hippophae rhamnoides L. (Elaeagnaceae; commonly known as “sea buckthorn” and “vitamin tree”), is a spiny deciduous shrub whose fruit is used in foods and traditional medicines. The H. rhamnoides fruit (berry) is rich in vitamin C, with a level exceeding that found in lemons and oranges. H. rhamnoides berries are usually washed and pressed to create pomace and juice. Today, the powder of the aqueous extract of H. rhamnoides berries are sold as a functional food in many countries. As part of our ongoing effort to identify bioactive constituents from natural resources, we aimed to isolate and identify those from the fruits of H. rhamnoides. Phytochemical analysis of the extract of H. rhamnoides fruits led to the isolation and identification of six compounds, namely, a citric acid derivative (1), a phenolic (2), flavonoids (3 and 4), and megastigmane compounds (5 and 6). Treatment with compounds 1–6 did not have any impact on the cell viability of RAW 264.7 mouse macrophages. However, pretreatment with these compounds suppressed lipopolysaccharide (LPS)-induced NO production in RAW 264.7 mouse macrophages in a concentration-dependent manner. Among the isolated compounds, compound 1 was identified as the most active, with an IC50 of 39.76 ± 0.16 μM. This value was comparable to that of the NG-methyl-L-arginine acetate salt, a nitric oxide synthase inhibitor with an IC50 of 28.48 ± 0.05 μM. Western blot analysis demonstrated that compound 1 inhibited the LPS-induced expression of IKKα/β (IκB kinase alpha/beta), I-κBα (inhibitor of kappa B alpha), nuclear factor kappa-B (NF-κB) p65, iNOS (inducible nitric oxide synthase), and COX-2 (cyclooxygenase-2) in RAW 264.7 cells. Furthermore, LPS-stimulated cytokine production was detected using a sandwich enzyme-linked immunosorbent assay. Compound 1 decreased interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α) production in LPS-stimulated RAW 264.7 cells. In summary, the mechanism of action of 1 included the suppression of LPS-induced NO production in RAW 264.7 cells by inhibiting IKKα/β, I-κBα, NF-κB p65, iNOS, and COX-2, and the activities of IL-6 and TNF-α.

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