Beneficial effects of tempol, a membrane-permeable radical scavenger, in a rodent model of collagen-induced arthritis.
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OBJECTIVE: To investigate the effects of tempol, a membrane-permeable radical scavenger, in rats with collagen-induced arthritis (CIA). METHODS: CIA was induced in Lewis rats by intradermal injection of 100 microl of an emulsion of 100 microg of bovine type II collagen (CII) in complete Freund's adjuvant (FCA) at the base of the tail. On day 21, a second injection of CII in FCA was administered. RESULTS: Lewis rats developed an erosive arthritis of the hind paws when immunized with CII in FCA. Macroscopic evidence of CIA first appeared as periarticular erythema and edema in the hind paws. The incidence of CIA was 100% by day 27 in the CII-challenged rats, and the severity of CIA progressed over a 35-day period. Radiographs revealed focal resorption of bone, with osteophyte formation in the tibiotarsal joint, and soft tissue swelling. The histopathologic features included erosion of the cartilage at the joint margins. Treatment of rats with tempol (10 mg/kg/day intraperitoneally) starting at the onset of arthritis (day 23) delayed the development of the clinical signs on days 24-35 and improved the histologic status of the knee and paw. Immunohistochemical analysis for nitrotyrosine and poly(ADP-ribose) synthetase (PARS) revealed positive staining in the inflamed joints of CII-treated rats. The degree of nitrotyrosine and PARS staining was markedly reduced in tissue sections obtained from CII-treated rats that had received tempol. Furthermore, radiographs revealed protection against bone resorption and osteophyte formation in the joints of tempol-treated rats. CONCLUSION: This study is the first to provide evidence that tempol, a small molecule that permeates biologic membranes and scavenges reactive oxygen species, attenuates the degree of chronic inflammation and tissue damage associated with CIA in the rat. (+info)
Bradyzide, a potent non-peptide B(2) bradykinin receptor antagonist with long-lasting oral activity in animal models of inflammatory hyperalgesia.
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Bradyzide is from a novel class of rodent-selective non-peptide B(2) bradykinin antagonists (1-(2-Nitrophenyl)thiosemicarbazides). Bradyzide has high affinity for the rodent B(2) receptor, displacing [(3)H]-bradykinin binding in NG108-15 cells and in Cos-7 cells expressing the rat receptor with K(I) values of 0.51+/-0.18 nM (n=3) and 0.89+/-0.27 nM (n=3), respectively. Bradyzide is a competitive antagonist, inhibiting B(2) receptor-induced (45)Ca efflux from NG108-15 cells with a pK(B) of 8.0+/-0.16 (n=5) and a Schild slope of 1.05. In the rat spinal cord and tail preparation, bradyzide inhibits bradykinin-induced ventral root depolarizations (IC(50) value; 1.6+/-0.05 nM (n=3)). Bradyzide is much less potent at the human than at the rodent B(2) receptor, displacing [(3)H]-bradykinin binding in human fibroblasts and in Cos-7 cells expressing the human B(2) receptor with K(I) values of 393+/-90 nM (n=3) and 772+/-144 nM (n=3), respectively. Bradyzide inhibits bradykinin-induced [(3)H]-inositol trisphosphate (IP(3)) formation with IC(50) values of 11.6+/-1.4 nM (n=3) at the rat and 2.4+/-0.3 microM (n=3) at the human receptor. Bradyzide does not interact with a range of other receptors, including human and rat B(1) bradykinin receptors. Bradyzide is orally available and blocks bradykinin-induced hypotension and plasma extravasation. Bradyzide shows long-lasting oral activity in rodent models of inflammatory hyperalgesia, reversing Freund's complete adjuvant (FCA)-induced mechanical hyperalgesia in the rat knee joint (ED(50), 0.84 micromol kg(-1); duration of action >4 h). It is equipotent with morphine and diclofenac, and 1000 times more potent than paracetamol, its maximal effect exceeding that of the non-steroidal anti-inflammatory drugs (NSAIDs). Bradyzide does not exhibit tolerance when administered over 6 days. In summary, bradyzide is a potent, orally active, antagonist of the B(2) bradykinin receptor, with selectivity for the rodent over the human receptor. British Journal of Pharmacology (2000) 129, 77 - 86 (+info)
Interrelationship of the kinin system, nitric oxide and eicosanoids in the antigen-induced arthritis in rabbits.
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The aim of the present study was to investigate the interrelationship of the kinin system, nitric oxide and eicosanoids in the acute phase of antigen-induced arthritis (AIA) in rabbits. The arthritis was induced in immunized rabbits and the following parameters were evaluated 24 hours later: leukocyte influx (total and differential white cell count), vascular permeability (Evans's blue method), and synovial PMN cell infiltrate. PGE2 and LTB4 (radioimmunoassay) levels were quantified in the synovial fluid. The animals were pre-treated with 20mg/kg/day during 14 days with L-NAME or D-NAME and/or Enalapril (0.12 mg/kg/day-14 days), and/or the B2 antagonist of Bradykinin HOE 140 (0.9 mg/kg). Our results showed that L-NAME was effective in the prevention of AIA with reduction of all Inflammatory parameters analyzed. Enalapril partially reverted the L-NAME anti-inflammatory effects. The simultaneous treatment with HOE 140 abolished this reversion and returned the inflammatory parameters to the levels observed in L-NAME treated animals. Our results suggest that pressoric alterations induced by L-NAME could not account for all its anti-inflammatory action in this model of experimental arthritis. Additionally the contribution of the kinin system in AIA was characterized as well as its interaction with eicosanoids and nitric oxide. (+info)
Alpha 4 integrin-dependent leukocyte recruitment does not require VCAM-1 in a chronic model of inflammation.
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Rats immunized with Mycobacterium butyricum in Freund's adjuvant develop a chronic vasculitis, with large increases in leukocyte rolling and adhesion in mesenteric postcapillary venules that are significantly inhibited with an alpha 4 integrin Ab. Using intravital microscopy to visualize chronically inflamed microvessels, we demonstrated that alpha 4 integrin-dependent leukocyte rolling and adhesion was inhibited with a beta 1 integrin, but not a beta 7 integrin Ab. To date, VCAM-1 has been presumed to be the primary ligand for alpha 4 beta 1 integrin in the vasculature. However, alpha 4 beta 1 integrin-dependent interactions were not reduced by monoclonal or polyclonal VCAM-1 Abs or a VCAM-1 antisense oligonucleotide despite increased VCAM-1 expression in the mesenteric vasculature. To ensure that the VCAM-1 Abs were functional and used at saturating concentrations, blood from Ab-treated rats was perfused over monolayers of CHO cells transfected with rat VCAM-1. Sufficient alpha 4 integrin or VCAM-1 Ab was present to inhibit leukocyte interactions with rat VCAM-1 by 95-100%. Under in vitro flow conditions, only mononuclear leukocytes were recruited from blood of control rats onto purified VCAM-1. However, neutrophils were also recruited onto VCAM-1 from whole blood of adjuvant-immunized animals via alpha 4 integrin. Another ligand for alpha 4 beta 1 integrin is the connecting segment-1 (CS-1) region of fibronectin. An Ab to the CS-1 portion of fibronectin, which did not reduce rolling and adhesion in adjuvant arthritis animals, completely inhibited leukocyte adhesion to CS-1 under static conditions. These findings provide the first evidence that alpha 4 beta 1 integrin-dependent leukocyte rolling and adhesion can occur in vivo via a mechanism other than VCAM-1. (+info)
The route of administration of an immunodominant peptide derived from heat-shock protein 65 dramatically affects disease outcome in pristane-induced arthritis.
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Previous studies have shown that immunization of mice with an immunodominant epitope from heat-shock protein 65 (hsp 65) (amino acids 261-271) can protect from the development of pristane-induced arthritis (PIA) and this protection is mediated by an antigen-specific T helper type 2 (Th2) cytokine response. Here we confirm these findings and show that frequent intranasal administration of this peptide exacerbates disease. In naive mice given peptide intranasally an antigen-specific T-cell population is systemically activated similar to that induced by peptide immunization in incomplete Freund's adjuvant. Thus, a paradox exists whereby apparently similar peptide-specific populations are either associated with protection from, or exacerbation of, PIA. However, comparison of cytokine profiles reveals differences between these two cell populations. Peptide inhalation induces the production of Th1-type cytokines (interferon-gamma) whereas intraperitoneal immunization leads to the production of Th2-type cytokines (interleukin-4, interleukin-5 and interleukin-10) by splenic T cells upon stimulation with peptide. Thus, for the application of nasal 'tolerance' in clinical medicine, it is important to identify antigens and dosing regimes that counteract but do not activate adverse immune responses. (+info)
Conditioning regimens for the treatment of experimental arthritis with autologous bone marrow transplantation.
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Adjuvant induced arthritis (AA) in Buffalo rats is a chronic progressive disease that responds very well to treatment with myeloablation and rescue with autologous BM. These previous results were obtained by conditioning with a lethal single dose of TBI. In the present study various other conditioning regimens were compared with single dose TBI. Fractionated TBI using adjusted total dose was equally effective. CY and BU when used as single agents at the highest tolerated dose were less effective. Combinations of CY (2 x 60 mg/kg) with lower dose (4 Gy) TBI and of BU with CY were as beneficial as high-dose TBI. These results indicate that a very intense reduction of T lymphocytes, in the order of 3-4 log, is required for obtaining the highest rate of long-lasting complete remissions. A similar conclusion was reached from our studies of various conditioning regimens in rats suffering from experimental allergic encephalitis (EAE). If extrapolated to the clinic, such a degree of T lymphocyte eradication poses upper limits to the number of T lymphocytes that can be safely reintroduced with the autograft. The exact limits cannot be derived from these experiments because the addition of autologous T lymphocytes to the graft yielded different results in the two models of autoimmune disease (AID). (+info)
Reversal of the antiinflammatory effects of methotrexate by the nonselective adenosine receptor antagonists theophylline and caffeine: evidence that the antiinflammatory effects of methotrexate are mediated via multiple adenosine receptors in rat adjuvant arthritis.
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OBJECTIVE: Weekly low-dose methotrexate (MTX) remains the mainstay of second-line therapy for rheumatoid arthritis (RA). We have previously reported that adenosine, acting at specific receptors on inflammatory cells, mediates the antiinflammatory effects of MTX in both in vitro and in vivo models of acute inflammation, but the mechanism by which MTX suppresses the chronic inflammation of arthritis remains controversial. The present study was undertaken to further investigate the means by which adenosine mediates the antiinflammatory effects of MTX. METHODS: The effects of 2 nonselective adenosine receptor antagonists, theophylline and caffeine, were examined, using the rat adjuvant arthritis model of RA. These agents were given alone and in conjunction with MTX, and arthritis severity was assessed clinically, radiologically, and histologically. Since rodent adenosine A3 receptors are not blocked by theophylline, selective A1, A2A, and A2B receptor antagonists were tested as well. RESULTS: Control animals developed severe arthritis, which was markedly attenuated by weekly treatment with MTX (0.75 mg/kg/week). Neither theophylline alone nor caffeine alone (each at 10 mg/kg/day) significantly affected the severity of the arthritis, but both agents markedly reversed the effect of MTX as measured by a severity index, hindpaw swelling, and hindpaw ankylosis. Radiographic and histologic analyses confirmed these observations. Neither A1, A2A, nor A2B receptor antagonists affected the capacity of MTX to ameliorate inflammation in adjuvant arthritis. CONCLUSION: These results provide strong evidence that adenosine mediates the antiinflammatory effects of MTX in this model of RA. Moreover, the findings suggest that abstinence from caffeine, a ubiquitous food additive and medication, may enhance the therapeutic effects of MTX in RA. (+info)
A role for complement in antibody-mediated inflammation: C5-deficient DBA/1 mice are resistant to collagen-induced arthritis.
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Collagen-induced arthritis (CIA) represents an animal model of autoimmune polyarthritis with significant similarities to human rheumatoid arthritis that can be induced upon immunization with native type II collagen. As in rheumatoid arthritis, both cellular and humoral immune mechanisms contribute to disease pathogenesis. Genotypic studies have identified at least six genetic loci contributing to arthritis susceptibility, including the class II MHC. We have examined the mechanism of Ab-mediated inflammation in CIA joints, specifically the role of complement activation, by deriving a line of mice from the highly CIA-susceptible DBA/1LacJ strain that are congenic for deficiency of the C5 complement component. We show that such C5-deficient DBA/1LacJ animals mount normal cellular and humoral immune responses to native type II collagen, with the activation of collagen-specific TNF-alpha-producing T cells in the periphery and substantial intra-articular deposition of complement-fixing IgG Abs. Nevertheless, these C5-deficient mice are highly resistant to the induction of CIA. These data provide evidence for an important role of complement in Ab-triggered inflammation and in the pathogenesis of autoimmune arthritis. (+info)