S. Henzgen

Attenuation of murine antigen-induced arthritis by treatment with a decoy oligodeoxynucleotide inhibiting signal transducer and activator of transcription-1 (STAT-1)

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.

Influence of cyclosporin A on cytokine levels in synovial fluid and serum of rats with antigen-induced arthritis

The effects of the T-cell-directed immunosuppressant cyclosporin A (CsA) on the developmnnt of antigen-induced arthritis (AIA) in rats as well as on cytokine levels in synovial fluid and serum were determined. The treatment with CsA effectively inhibited the chronic phase of arthritis as demonstrated by decreased joint swelling and reduced histological arthritic score. In animals with AIA the level of IL-6 in the synovial fluid and serum is increased, showing good correlation with the severity of the disease. The CsA treatment reduced IL-6 levels to normal. IL-1, IL-2 and TNF-α do not seem to be directly involved in the pathogenic mechanisms of chronic arthritis.

Immunomodulation of rat antigen-induced arthritis by leflunomide alone and in combination with cyclosporin A

The effects of the new immunomodulating isoxazol derivative leflunomide, in comparison with cyclosporin A, on established antigen-induced arthritis in rats as well as serum antibody levels were determined. When treatment with leflunomide, at concentrations from 2.5 to 10 mg/kg/d, was started on day 3 of arthritis, the acute and chronic phases of arthritis were effectively inhibited. This was demonstrated by decreased joint swelling and reduced histopathological arthritis score at the end of experiment (day 26). Furthermore, the treatment resulted in a significantly reduced level of serum antibodies to the matrix components collagen type I, type II and proteoglycans. Neither leflunomide nor cyclosporin A, at doses of 1 mg/kg/d, had an effect on the severity of arthritis and antibody levels. However, when both drugs were used together, at these non-effective doses, the histopathological score of chronic arthritis was significantly reduced. The results of our experiments demonstrate that leflunomide has a strong suppressive effect on both acute and chronic phases of antigen-induced arthritis and formation of autoantibodies in rats. Furthermore, orally administered doses of leflunomide were as effective as doses of cyclosporin A given intraperitoneally. The combination of sub-effective doses of leflunomide and cyclosporin A resulted in significant inhibition of chronic arthritis.

Significance of cell-mediated and humoral immunity in the acute and chronic phase of antigen-induced arthritis in rabbits

In the course of antigen-induced arthritis of rabbit cell-mediated and humoral immune responses were repeatedly tested in order to prove their significance for the acute and chronic phase of inflammation. The arthritis was monitored during the progression of the inflammation by means of the joint swelling and at the end of experiments by histological evaluation of synovitis and cartilage degradation. Following the arthritis induction a strong increase of specific antibodies and of circulating immune complexes was evident. The correlations between antibody levels and joint swellings confirmed that the local formation of immune complexes is responsible for the initiation and perpetuation of arthritis. In the early phase after immunization the responsiveness of lymphocytes to antigenic and mitogenic stimulation was increased, in the late chronic phase of arthritis proliferative responses of lymphocytes to cartilage matrix components were revealed. No direct correlations could be demonstrated between any cell-mediated immune response and the severity of arthritis. The hyperreactivity of cell-mediated immunity is suggested to be responsible for the transition of the acute arthritis into the chronic stage. The deficiency of an effective suppression results in the activation of B-lymphocytes with increased production of antibodies, maintaining the inflammatory process for a long time. Under these conditions the release of cartilage matrix components during the acute joint reaction induces autoimmune responses against cartilage, which could contribute to the chronification of arthritis and to cartilage degradation.

Amelioration of murine antigen-induced arthritis by dehydroepiandrosterone (DHEA).

AbstractObjective and design:Sex hormones have immunomodulatory properties and may play an important role in the pathogenesis of autoimmune diseases like rheumatoid arthritis (RA). This study sought to examine the effects of the natural weak androgen dehydroepiandrosterone (DHEA) and its metabolite androstenediol (AED) on the development of murine antigen-induced arthritis (AIA). Methods:DHEA and AED were administered orally, approximately 10 mg/day, from the time of AIA induction (i.e., 3 weeks after start of immunization) in young male as well as young and old female C57BL/6 mice. The effects were assessed in terms of joint swelling, histological changes, and cell-mediated and humoral immunity. Results:Compared to untreated AIA animals, continuous administration of DHEA decreased knee joint swelling during acute and chronic AIA, as well as histological signs of inflammation and joint destruction during chronic AIA. These effects were age- and gender-independent. Delayed-type hypersensitivity (DTH) to the specific antigen methylated bovine serum albumin (mBSA) was significantly reduced, but there were no changes in the balance of the T helper (Th) cell subsets Th1/Th2, as tested by the ratio of IgG isotypes in the sera. Whereas serum levels of IgG antibodies to mBSA were not influenced, the formation of IgG autoantibodies to the matrix constituents collagen type I, collagen type II, and cartilage proteoglycans was significantly inhibited. In all experiments, the effects of AED were not significantly stronger than those of DHEA. Conclusions:Administration of exogenous DHEA ameliorates the severity of acute and chronic AIA, presumably by suppressing cell-mediated immunity against mBSA (the inducing antigen) and formation of autoantibodies. However, because of the fundamentally different DHEA physiology in rodents, the role of such a replacement therapy in human RA deserves further elucidation.

Different immunological mechanisms contribute to cartilage destruction in antigen-induced arthritis

Antigen-induced arthritis in guinea pigs was used as a model to investigate the pathogenic mechanisms responsible for cartilage destruction in chronic joint inflammation. The activation of macrophages, their effects on cartilage metabolism, and the development of autoimmunity to cartilage constituents were studied during the progression of arthritis. The results show that in arthritic animals the macrophages are systemically activated, with a peak in the early phase of inflammation. Interleukin 1, produced by the activated cells, suppresses the proteoglycan synthesis in cartilage explants and cultured chondrocytes and increases the proliferation of the cells in vitro. During the progression of arthritis humoral and cell-mediated immune responses to collagen type II and cartilage proteoglycans occur correlating with the severity of arthritis. It is concluded that different immunological mechanisms may be involved in cartilage destruction during antigen-induced arthritis. Mediator-induced metabolic reactions dominate in the early phase, whereas autoimmunity to cartilage might play an essential role in later phases of arthritis.

High-dose clodronate therapy prevents joint destruction in chronic antigen-induced arthritis of the rat but inhibits bone formation at the axial skeleton

Abstract:Objective: To investigate the effects of clodronate on clinical disease activity, inflammatory alterations and cartilage destruction, periarticular and axial bone volume and bone turnover in chronic antigen-induced arthritis (AIA; day 28).¶Methods: Rats with AIA were treated with clodronate (5 mg/kg/day continuously; 20 mg/kg/day intermittently or high-dose with 300 mg/kg 3 hours after arthritis induction +20 mg/kg/day continuously, respectively). Joint pathology was examined by histology. Bone volume and cellular turnover parameters of the right tibia head and the third lumbar vertebra were evaluated by histomorphometry. The findings were compared with those of healthy controls, sham-treated AIA and AIA treated continuously with 250 μg/kg of dexamethasone.¶Results: All three therapy regimens with clodronate resulted in a significant reduction of joint swelling, histopathological inflammatory changes and cartilage destruction in comparison with sham-treated AIA. The antiinflammatory effect of high-dose clodronate was comparable with dexamethasone. The intermittent administration of 20 mg/kg/day of clodronate completely prevented periarticular bone loss by reduction of bone resorption without affecting bone formation at the periarticular and axial bone. Both continuous treatment with 5 mg/kg/day of clodronate and high-dose clodronate therapy partially prevented periarticular bone loss and reduced parameters of bone formation at the axial bone to values below those of healthy controls.¶Conclusion: High-dose clodronate therapy exerts an excellent preventive effect on clinical disease activity and on joint destruction in AIA. However, continuous treatment with high doses of clodronate may result in a low turnover state of bone remodelling.¶

CHANGES OF IMMUNOREGULATORY PROPERTIES AND INDUCTION OF AUTOIMMUNE REACTIVITY TO CARTILAGE IN RABBITS WITH ANTIGEN-INDUCED ARTHRITIS

The chronicity of the antigen-induced arthritis is characterized as dependent on the development of cell-mediated immunity to the antigen, but the exact mechanisms underlying are unclear. We have evidenced decreased suppressor and increased helper cell potential in the early phase of arthritis as result of the immunization procedure. In the late phase of arthritis proliferative responses of spleen lymphocytes to cartilage proteoglycans were revealed which were neither present in immunized animals without arthritis induction nor in the early phase of arthritis. The changes of the regulatory properties on the T-cell level are probably responsible for the transition of acute arthritis into the chronic stage. The deficiency of an effective suppression and/or the increased helper cell potential results in the activation of B- and T-lymphocytes with increased cell-mediated and humoral immune responsiveness to the antigen maintaining the inflammatory process for a long time. In this situation the release of cartilage proteoglycans during the acute joint reaction induces autoimmune responses against cartilage which could contribute to the chronification of inflammation and to cartilage degradation.

Degradation of articular cartilage during the progression of antigen-induced arthritis in mice: A scanning and transmission electron microscopic study

Ultrastructural changes of knee articular cartilage in C57B1/6 mice were studied in the course of antigen-induced arthritis by means of scanning and transmission electron microscopy. First damages of the cartilage surface were seen one hour after arthritis induction. The earliest signs were the loss of the superficial electron-dense layer as well as a progressive loss of proteoglycans in the cartilaginous matrix on the surface. Breaks of collagen fibres were already detected at the first day of arthritis. The chondrocytes of the superficial cartilage layer showed an increase of the intracellular membraneous system in the early phase of arthritis. Thereafter chondrocytes on the surface became more and more necrotic. On the 7th day of arthritis acute alterations of cartilage had developed completely. Many lacunae of chondrocytes were opened and the cartilage surface showed deep structural defects with adhering cells, probably lymphocytes and macrophages. In the following time these destructive processes were demonstrated along with cellular proliferation as a sign of repair attempts in hyaline cartilage.

Effects of the immunomodulator diacetyl-splenopentin on antigen-induced arthritis in rabbits

Long-term treatment with the immunomodulator diacetyl-splenopentin reduces the severity of chronic joint inflammation and cartilage destruction in rabbits with antigen-induced arthritis. The level of specific antibodies as well as specific and non-specific cell-mediated immune reactivities including the proliferative response of spleen lymphocytes to cartilage proteoglycans in treated animals are lower than in untreated arthritic rabbits. Moreover, suppressor cell activity, which normally decreases during the early phase of inflammation, is enhanced and hyperreactive helper cell potential is reduced. These findings suggest that treatment with diacetyl-splenopentin normalizes the immune regulation, which is disturbed in the early phase of inflammation. This might result in a depression of the hyperreactive immune system including the autoimmunity developed against cartilage. Lowered immune reactivity in the joint in turn reduces the severity of chronic joint inflammation.