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Abstract

The transcription factor STAT-1 (signal transducer and activator of transcription-1) plays a pivotal role in the expression of inflammatory gene products involved in the pathogenesis of arthritis such as various cytokines and the CD40/CD40 ligand (CD40/CD40L) receptor-ligand dyad. The therapeutic efficacy of a synthetic decoy oligodeoxynucleotide (ODN) binding and neutralizing STAT-1 was tested in murine antigen-induced arthritis (AIA) as a model for human rheumatoid arthritis (RA). The STAT-1 decoy ODN was injected intra-articularly in methylated bovine serum albumin (mBSA)-immunized mice 4 h before arthritis induction. Arthritis was evaluated by joint swelling measurement and histological evaluation and compared to treatment with mutant control ODN. Serum levels of pro-inflammatory cytokines, mBSA-specific antibodies and auto-antibodies against matrix constituents were assessed by enzyme-linked immunosorbent assay (ELISA). The transcription factor neutralizing efficacy of the STAT-1 decoy ODN was verified in vitro in cultured synoviocytes and macrophages. Single administration of STAT-1 decoy ODN dose-dependently suppressed joint swelling and histological signs of acute and chronic arthritis. Delayed-type hypersensitivity (DTH) reaction, serum levels of interleukin-6 (IL-6) and anti-proteoglycan IgG titres were significantly reduced in STAT-1 decoy ODN-treated mice, whereas mBSA, collagen type I and type II specific immunoglobulins were not significantly affected. Intra-articular administration of an anti-CD40L (anti-CD154) antibody was similarly effective. Electrophoretic mobility shift analysis (EMSA) of nuclear extracts from synoviocytes incubated with the STAT-1 decoy ODN in vitro revealed an inhibitory effect on STAT-1. Furthermore, the STAT-1 decoy ODN inhibited the expression of CD40 mRNA in stimulated macrophages. The beneficial effects of the STAT-1 decoy ODN in experimental arthritis presumably mediated in part by affecting CD40 signalling in macrophages may provide the basis for a novel treatment of human RA. IndroductionHuman rheumatoid arthritis (RA) is a chronic systemic disorder of unknown aetiology, characterized by intimal lining layer hyperplasia, infiltration of the sublining area by macrophages, T and B lymphocytes, plasma cells and other inflammatory cells as well as progressive destruction of joint structures [1,2]. Despite the uncertainty about its aetiology, RA is thought to be an immune-mediated disease promoting inflammation and tissue destruction. Besides the pro-inflammatory cytokines tumour necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6, which mainly derive from macrophages, a dominant T-helper (Th)1-response is associated with the disease, which is characterized by an imbalance of interferon (IFN)-γ over 4]. AIA = antigen-induced arthritis; AP = alkaline phosphatase; bp = base pairs; CD40L = CD40 ligand (CD154); DMEM = Dulbecco's modified Eagle's medium; DTH = delayed-type hypersensitivity; ELISA = enzyme-linked immunosorbent assay; EMSA = electrophoretic mobility s...

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“…Four-micrometer-thick sections were stained with H&E, safranin O, or toluidine blue. The inflammation score and joint destruction score were measured using the procedure of Huckel et al by three individual researchers40. The inflammation score of each group was measured by the severity of infiltration and pannus formation.…”

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