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Adenoviral Gene Transfer of the Endogenous Inhibitor IkBa into Human Osteoarthritis Synovial Fibroblasts Demonstrates That Several Matrix Metalloproteinases and Aggrecanases Are Nuclear Factor-kB-Dependent
JAN BONDESON, SARAH LAUDER, SHANE WAINWRIGHT, NICK AMOS, AMY EVANS, CLARE HUGHES, MARC FELDMANN, and BRUCE CATERSON ABSTRACT. Objective. To investigate the role of the transcription factor nuclear factor-kB (NF-kB) in promoting inflammatory and destructive responses in human osteoarthritis (OA) synovial fibroblasts, by assessing the effect of NF-kB blockade on the production of cytokines and destructive enzymes. Methods. Infection with adenoviruses transferring the ß-galactosidase gene was used to ascertain that the OA fibroblasts could be infected (> 95%). Using an adenovirus transferring the inhibitory subunit IkBa, effective inhibition of NF-kB was achieved. The expression and production of several pro- and antiinflammatory cytokines and mediators, the major matrix metalloproteinases (MMP 1, 3, and 13), their main inhibitor tissue inhibitor of metalloproteinase-1 (TIMP-1), and the aggrecanases (ADAMTS4 and ADAMTS5) were measured by ELISA and/or reverse transcription-polymerase chain reaction, and their dependence on NF-kB evaluated. Results. The production of interleukin 6 (IL-6), monocyte chemoattractant protein-1, and RANTES was potently inhibited by IkBa overexpression, irrespective of stimulus, but IL-8 was unaffected. The p55 soluble tumor necrosis factor (TNF) receptor was unaffected, but the p75 soluble TNF receptor was potently inhibited by IkBa overexpression. MMP-1, MMP-3, and MMP-13 were inhibited by IkBa overexpression, at both the mRNA and protein levels, whereas TIMP-1 mRNA was unaffected. The mRNA gene expression of ADAMTS4 was also inhibited by IkBa overexpression, particularly in IL-1-stimulated cells, but ADAMTS5 was unaffected. Conclusion. In OA synovial fibroblasts, inhibition of NF-kB has a beneficial effect on the balance between the expression of proinflammatory cytokines and antiinflammatory mediators. Inhibition of this transcription factor also results in the decreased expression of several destructive metalloproteinases and also the ADAMTS4 aggrecanase. (First Release Jan 15 2007; J Rheumatol 2007;34:523-33) Key Indexing Terms: ADENOVIRUS From the Department of Rheumatology; Connective Tissue Biology Laboratories, Cardiff School of Biosciences, Cardiff University, Cardiff; and Kennedy Institute of Rheumatology Division, Imperial College School of Medicine, London, UK. Supported by the Arthritis Research Campaign (UK), grants W0596, 13172, and 14570. J. Bondeson, MD, PhD, Senior Lecturer; S. Lauder, BSc, PhD Student; N. Amos, MSc, Clinical Scientist; A. Evans, BSc, Research Assistant, Department of Rheumatology; S. Wainwright, PhD, Postdoctoral Fellow; C. Hughes, PhD, Lecturer; B. Caterson, PhD, Professor, Connective Tissue Biology Laboratories, Cardiff School of Biosciences, Cardiff University; M. Feldmann, PhD, Professor, Kennedy Institute of Rheumatology Division, Imperial College School of Medicine. Address reprint requests to Dr. J. Bondeson, Department of Rheumatology, Cardiff University, Heath Park, Cardiff CF14 4XN, UK. E-mail BondesonJ@cf.ac.uk Accepted for publication October 10, 2006.
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