scholarly journals Regulation of the mouse inducible-type nitric oxide synthase gene promoter by interferon-γ, bacterial lipopolysaccharide and NG-monomethyl-l-arginine

1996 ◽  
Vol 316 (1) ◽  
pp. 209-215 ◽  
Author(s):  
Alessandro WEISZ ◽  
Luigi CICATIELLO ◽  
Hiroyasu ESUMI
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Gene Promoter ◽  
Interferon Γ ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Promoter Activation ◽  
Inos Gene ◽  
Ifn Γ ◽  
Oxide Synthase

Cytokines and bacterial lipopolysaccharides (LPSs) stimulate nitric oxide production in macrophages by inducing transcription of the gene coding for the inducible isoform of nitric oxide synthase (iNOS). We have cloned the mouse iNOS gene promoter and analysed its structural features and its response to interferon-γ (IFN-γ) and Escherichia coli LPS in RAW 264.7 mouse macrophage-like cells. Transcription of a recombinant reporter gene including the promoter and 4 kb of its 5′-flanking DNA, linked to the bacterial chloramphenicol acetyltransferase (CAT) reporter gene, is stimulated by IFN-γ and, more efficiently, by LPS upon transient transfection in RAW 264.7 cells. Two upstream DNA regions are required for maximal promoter activation by LPS: the first maps between positions -1541 and -775 and the other between -420 and -47, with respect to the major transcriptional start site of the iNOS gene. The upstream-most region also mediates promoter trans-activation by IFN-γ. As reported earlier for transcription of the endogenous iNOS gene, combined stimulation of RAW 264.7 cells with IFN-γ and LPS results in lower activation of the transfected promoter, when compared with LPS alone. NG-Monomethyl-L-arginine, a competitive inhibitor of nitric oxide synthase activity, enhances iNOS gene mRNA induction and promoter activation by IFN-γ and LPS, indicating that nitric oxide can influence negatively the responsiveness of this gene to inducers. These results suggest the possibility of a negative regulatory feedback exerted by iNOS on the transcriptional activation of its own gene.

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

Foods ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 269 ◽  
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.

scholarly journals Suppression of Nitric Oxide Synthase by Thienodolin in Lipopolysaccharide-stimulated RAW 264.7 Murine Macrophage Cells

2012 ◽  
Vol 7 (6) ◽  
pp. 1934578X1200700 ◽  
Author(s):  
Eun-Jung Park ◽  
John M. Pezzuto ◽  
Kyoung Hwa Jang ◽  
Sang-Jip Nam ◽  
Sergio A. Bucarey ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Nuclear Translocation ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Dependent Manner ◽  
Signal Transducers ◽  
Protein Levels ◽  
Mitogen Activated Protein ◽  
Oxide Synthase

The measurement of nitric oxide in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells is used as a model for evaluating the anti-inflammatory or chemopreventive potential of substances. Thienodolin, isolated from a Streptomyces sp. derived from Chilean marine sediment, inhibited nitric oxide production in LPS-stimulated RAW 264.7 cells (IC50 = 17.2 ± 1.2 μM). At both the mRNA and protein levels, inducible nitric oxide synthase (iNOS) was suppressed in a dose-dependent manner. Mitogen-activated protein kinases (MAPKs), one major upstream signaling pathway involved in the transcription of iNOS, were not affected by treatment of thienodolin. However, the compound blocked the degradation of IκBα resulting in inhibition of NF-κB p65 nuclear translocation, and inhibited the phosphorylation of signal transducers and activators of transcription 1 (STAT1) at Tyr701. This study supports further exploration of thienodolin as a potential therapeutic agent with a unique mechanistic activity.

Tannins and related compounds induce nitric oxide synthase and cytokines gene expressions in Leishmania major-infected macrophage-like RAW 264.7 cells

2005 ◽  
Vol 13 (23) ◽  
pp. 6470-6476 ◽  
Author(s):  
Herbert Kolodziej ◽  
Anne Burmeister ◽  
Weronika Trun ◽  
Oliver A. Radtke ◽  
Albrecht F. Kiderlen ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Leishmania Major ◽  
Infected Macrophage ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Gene Expressions ◽  
Oxide Synthase ◽  
Related Compounds

Induction of nitric oxide synthase by lipopolysaccharide and its inhibition by auranofin in RAW 264.7 cells

1997 ◽  
Vol 338 (2) ◽  
pp. 151-158 ◽  
Author(s):  
Masamichi Yamashita ◽  
Hisae Niki ◽  
Masateru Yamada ◽  
Suetsugu Mue ◽  
Kazuo Ohuchi
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Oxide Synthase
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