Selenium deficiency increases the expression of inducible nitric oxide synthase in RAW 264.7 macrophages: role of nuclear factor-kappaB 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-kappaB (NF-kappaB)-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-kappaB. 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.
Prabhu
K Sandeep
KS
Department of Veterinary Science and The Center for Molecular Toxicology and Carcinogenesis, 115 Henning, The Pennsylvania State University, University Park, PA 16802, U.S.A.
Zamamiri-Davis
Faith
F
Stewart
Jennifer B
JB
Thompson
Jerry T
JT
Sordillo
Lorraine M
LM
Reddy
C Channa
CC
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D005609:Free Radicals / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008264:Macrophages / Q000201:enzymology*; D051379:Mice; D016328:NF-kappa B / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis* / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D018384:Oxidative Stress; D010100:Oxygen / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012643:Selenium / Q000172:deficiency* / Q000378:metabolism; D013997:Time Factors; D015854:Up-Regulation
D005609:Free Radicals; D016328:NF-kappa B; D005979:Glutathione Peroxidase; D008156:Luciferases; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D012643:Selenium; D010100:Oxygen