THU0093 Nadph oxidases associated production of reactive oxygen species in rheumatoid arthritis

Background Reactive oxygen species (ROS) is produced during metabolism of Oxygen. ROS is important in cell signalling and homeostasis. Production of ROS can be elevated in stressful condition. Oxidative stress has been known to be related with the disease like infection and malignancy. NADPH oxidases (Nox) are membrane proteins which produce ROS. Objectives In this study we aimed to investigate the role of Nox in rheumatoid arthritis (RA) associated with production of ROS. Methods Nox and Granulocyte macrophage colony-stimulating factor (GMCSF) messenger ribonucleic acid (mRNA) were analysed in fibroblast like synoviocyte (FLS) of patients with RA and osteoarthritis (OA) by reverse transcription polymerase chain reaction. Amount of ROS which is produced in FLS of patients with RA and OA is determined using the cell permeant fluoroprobe 5-(and-6)-chloromethyl-2’,7’-dichlorodihydrofluorescein diacetate acetyl ester (CM-H2DCFDA) by flowcytometry. Same experiments were performed after treatment with cytokine, interleukin (IL)−17 and tumour necrosis factor-α (TNF-α). Results Among Nox subunits (Nox1, Nox2, Nox4, Nox5, DUOX1, DUOX2, NOXA1, NOXO1), NOXA1 and NOXO1 mRNA were expressed higher in RA FLS than in OA FLS. After treatment with IL-17 and TNF-α for 24 hours GMCSF, Nox1 and NOXO1 mRNA in RA FLS were elevated. Amount of ROS production was also elevated after treatment with IL-17 and TNF-α. When RA FLS were treated with bromopyruvic acid (BrPa), glucolysis inhibitor by inhibition of hexokinase II, GMCSF mRNA and ROS were decreased and Nox1 and Nox4 mRNA showed no diffrence. Conclusions Several factors may be involved between ROS and Nox in RA FLS. Both ROS and Nox were elevated in inflammatory condition in RA FLS. From this result we expect that Nox-targeted therapy may be effective for treatment with RA. References [1] Drummond GR, Selemidis S, Griendling KK, Sobey CG. Combating oxidative stress in vascular disease: NADPH oxidases as therapeutic targets. Nat Rev Drug Discov2011;10:453–71. [2] Neumann E, Lefevre S, Zimmermann B, Gay S, Muller-Ladner U. Rheumatoid arthritis progression mediated by activated synovial fibroblasts. Trends Mol Med2010;16:458–68. [3] Rodino-Janeiro BK, Paradela-Dobarro B, Castineiras-Landeira MI, Raposeiras-Roubin S, Gonzalez-Juanatey JR, Alvarez E. Current status of NADPH oxidase research in cardiovascular pharmacology. Vasc Health Risk Manag2013;9:401–28. [4] Xiao C, Li J, Dong X, He X, Niu X, Liu C, Zhong G, Bauer R, Yang D, Lu A. Anti-oxidativeand TNF-α suppressive activities of puerarin derivative (4AC) in RAW264.7 cells and collagen-induced arthritic rats. Eur J Pharmacol2011;666:242–50. [5] Yun JM, Chien A, Jialal I, Devaraj S. Resveratrol up-regulates SIRT1 and inhibits cellular oxidative stress in the diabetic milieu: mechanistic insights. J Nutr Biochem2012;23:699–705. Disclosure of Interest None declared