S C M van Dalen1, N N L Kruisbergen2, B Walgreen3, M M A Helsen4, A W Slöetjes5, N A J Cremers6, M I Koenders7, F A J van de Loo8, J Roth9, T Vogl10, A B Blom11, P M van der Kraan12, P L E M van Lent13, M H J van den Bosch14. 1. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Stephanie.vanDalen@radboudumc.nl. 2. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Nik.Kruisbergen@radboudumc.nl. 3. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Birgitte.Walgreen@radboudumc.nl. 4. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Monique.Helsen@radboudumc.nl. 5. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Annet.Sloetjes@radboudumc.nl. 6. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Niels.Cremers@radboudumc.nl. 7. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Marije.Koenders@radboudumc.nl. 8. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Fons.vandeLoo@radboudumc.nl. 9. Institute of Immunology, University of Münster, Germany. Electronic address: rothj@uni-muenster.de. 10. Institute of Immunology, University of Münster, Germany. Electronic address: vogl@uni-muenster.de. 11. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Arjen.Blom@radboudumc.nl. 12. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Peter.vanderKraan@radboudumc.nl. 13. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Peter.vanLent@radboudumc.nl. 14. Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address: Martijn.vandenBosch@radboudumc.nl.
Abstract
OBJECTIVE: Synovitis in collagenase-induced osteoarthritis (CiOA) is driven by locally released S100A8/A9 proteins and enhances joint destruction. S100A8/A9 can induce reactive oxygen species (ROS) release by phagocytes in OA synovium via neutrophil cytosolic factor-1 (Ncf1)-regulated NOX2 activation. In the present study we investigated whether NOX2-derived ROS affect joint pathology during CiOA. METHODS: CiOA was induced in knee joints of wild type (WT) and Ncf1-deficient (Ncf1**) mice. Synovial gene expression of NOX2-subunits was measured with quantitative real-time polymerase chain reaction (qRT-PCR). Joint pathology was assessed using histology and immunohistochemistry for aggrecan neo-epitope VDIPEN. Levels of inflammatory proteins were measured with Luminex or ELISA. Phagocytes in synovium, blood, bone marrow (BM) and spleen were analyzed with flow cytometry. ROS release by phagocytes was measured with a ROS detection kit. RESULTS: CiOA induction in knee joints of WT mice caused significantly increased synovial gene expression of NOX2 subunits. On day 7 of CiOA, cartilage damage and MMP activity, as measured by VDIPEN, were comparable between WT and Ncf1** mice. Synovial thickening, synovial S100A8/A9 levels and percentages of synovial macrophages, polymorphonuclear cells (PMNs), and monocytes were not different, as were levels of inflammatory mediators in serum and phagocyte percentages in blood, BM and spleen. On day 42 of CiOA, synovitis, cartilage damage, and osteophyte formation in Ncf1** mice were unaltered when compared to WT mice. ROS detection confirmed that Ncf1** PMNs lack functional NOX2, but in vitro macrophages showed ROS production, suggesting activation of compensatory mechanisms. CONCLUSIONS: Absence of Ncf1-mediated ROS production does not alter joint pathology in CiOA.
OBJECTIVE:Synovitis in collagenase-induced osteoarthritis (CiOA) is driven by locally released S100A8/A9 proteins and enhances joint destruction. S100A8/A9 can induce reactive oxygen species (ROS) release by phagocytes in OA synovium via neutrophil cytosolic factor-1 (Ncf1)-regulated NOX2 activation. In the present study we investigated whether NOX2-derived ROS affect joint pathology during CiOA. METHODS:CiOA was induced in knee joints of wild type (WT) and Ncf1-deficient (Ncf1**) mice. Synovial gene expression of NOX2-subunits was measured with quantitative real-time polymerase chain reaction (qRT-PCR). Joint pathology was assessed using histology and immunohistochemistry for aggrecan neo-epitope VDIPEN. Levels of inflammatory proteins were measured with Luminex or ELISA. Phagocytes in synovium, blood, bone marrow (BM) and spleen were analyzed with flow cytometry. ROS release by phagocytes was measured with a ROS detection kit. RESULTS:CiOA induction in knee joints of WT mice caused significantly increased synovial gene expression of NOX2 subunits. On day 7 of CiOA, cartilage damage and MMP activity, as measured by VDIPEN, were comparable between WT and Ncf1** mice. Synovial thickening, synovial S100A8/A9 levels and percentages of synovial macrophages, polymorphonuclear cells (PMNs), and monocytes were not different, as were levels of inflammatory mediators in serum and phagocyte percentages in blood, BM and spleen. On day 42 of CiOA, synovitis, cartilage damage, and osteophyte formation in Ncf1** mice were unaltered when compared to WT mice. ROS detection confirmed that Ncf1** PMNs lack functional NOX2, but in vitro macrophages showed ROS production, suggesting activation of compensatory mechanisms. CONCLUSIONS: Absence of Ncf1-mediated ROS production does not alter joint pathology in CiOA.