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A proteomic approach to identify proteins involved in Redox regulation of inflammation and immunity
Author(s) -
salzano sonia,
Coppo Lucia,
Bowler Lucas,
Cervellini Ilaria,
Mengozzi Manuela,
Ghezzi Pietro
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.lb671
Subject(s) - western blot , peroxiredoxin , lipopolysaccharide , oxidative stress , microbiology and biotechnology , chemistry , inflammation , blot , secretion , glutathione , antioxidant , biochemistry , biology , immunology , enzyme , gene , peroxidase
To better understand the role of redox regulation in immunity, we studied the glutathionylation of proteins secreted in response to inflammatory stimuli such as lipopolysaccharide (LPS). These proteins were detected by cell labeling with biotinylated glutathione ethyl ester (BioGEE) and identified by one‐dimensional gel electrophoresis, subjected to tryptic digestion followed by mass spectrometry analysis. Several proteins were identified in supernatants from Raw 264.7 cells (murine macrophages) stimulated with 100 ng/ml LPS for 24 hours. We focused our study on Peroxiredoxin 2 (Prx2), an antioxidant enzyme involved in cells protection against oxidative stress which breaks down H2O2. Following its identification, we studied secretion of Prx2 by Raw 264.7 cells using Western blot. In vivo studies were also performed to measure Prx2 levels in serum from mice injected intraperitoneally with saline (vehicle) or with 400 μg/mouse LPS (LPS‐treated mice). On the basis of our Western blot data, we confirmed higher levels of secreted Prx2 in Raw 264.7 cells after LPS stimulation. Increase in Prx2 levels were also observed in serum from LPS‐treated mice. Supported by BSMS and interreg TC2N