Objective
Cartilage of young but skeletally mature dogs is more susceptible to blood‐induced damage than that of old dogs. The aim of the present study was to investigate whether cartilage of ...skeletally immature individuals is even more adversely affected by exposure to blood than that of mature individuals, as suggested by clinical practice experience with humans.
Methods
Right knees of 3 groups of 6 beagle dogs (skeletally immature, young mature, and old animals) were injected with autologous blood on days 0 and 2. On day 4, cartilage matrix proteoglycan turnover (content, synthesis, and release), synovial inflammation, and cartilage‐destructive properties of the synovial tissue were determined and compared with those of the left uninjected control knees.
Results
Subsequent to intraarticular bleeding, cartilage proteoglycan content decreased in an age‐dependent manner, with the largest decrease occurring in cartilage of immature animals. Proteoglycan synthesis per cell also decreased in an age‐dependent manner, with the largest decrease occurring in the immature animals. Cartilage proteoglycan release increased in all 3 groups, but the decrease was not age dependent. Interestingly, immature animals showed a large increase in cartilage DNA content upon exposure to blood, whereas mature animals did not. Histologic analysis showed a mild synovitis in animals of all ages, but catabolic inflammatory activity was found only in immature animals.
Conclusion
Joints of skeletally immature dogs appeared to be more susceptible than joints of mature dogs to the adverse effects of a joint hemorrhage. These data suggest that for humans, specifically young children are at risk for joint damage after a joint hemorrhage.
Objective
To investigate the direct and indirect (via synovial inflammation) effects of intraarticular bleeding on cartilage in vivo.
Methods
Right knees of 14 beagle dogs were injected with ...autologous blood on days 0 and 2. Cartilage matrix proteoglycan turnover, collagen damage, and synovial inflammation of these knees, including the cartilage‐destructive properties of the synovial tissue, were determined and compared with those of the left control knees on day 4 (short‐term effects; n = 7) and day 16 (long‐term effects; n = 7).
Results
Injected knees had a diminished content of proteoglycans in the cartilage matrix, and release of proteoglycans was enhanced (days 4 and 16). The synthesis of proteoglycans was significantly inhibited on day 4 but was enhanced on day 16. On day 4 more collagen was denatured in the injected joint than in the control joint; this effect was no longer detectable on day 16. Synovial tissue showed signs of inflammation on day 4 and day 16 but had cartilage‐destructive properties only on day 16.
Conclusion
In vivo exposure of articular cartilage to blood for a relatively short time results in lasting changes in chondrocyte activity and in cartilage matrix integrity, changes that may predict lasting joint damage over time. Interestingly, the direct effect of blood on cartilage precedes the indirect effect via synovial inflammation.
Objective. Inflammation‐induced articular cartilage degradation is a major problem in rheumatoid arthritis (RA). Type 1 T cell activity (characterized by interferon‐γ/interleukin‐2 IL‐2 production), ...and consequently, the production of the proinflammatory cytokines IL‐1 and tumor necrosis factor α (TNFα), have been reported to play a major role in cartilage damage. IL‐10 and IL‐4, both produced by type 2 T cells, are cytokines with the capacity to down‐regulate proinflammatory responses. The present study was undertaken to investigate the way in which these cytokines affect activated mononuclear cells (MNC) of RA patients in relation to human articular cartilage degradation in vitro.
Methods. MNC from synovial fluid and peripheral blood of RA patients were stimulated with bacterial antigen and treated with IL‐10 and/or IL‐4. Bacterial antigen is known to activate type 1 T cells and to induce proinflammatory IL‐1/TNFα–dependent cartilage damage. Cytokine production and effects of conditioned media, as well as effects of IL‐10 and IL‐4 on proteoglycan (PG) turnover (as a measure for cartilage damage), were determined.
Results. IL‐10 and IL‐4 inhibited proinflammatory cytokine production of stimulated RA MNC and completely reversed inhibition of cartilage PG synthesis induced by these stimulated RA MNC. IL‐10 was more potent than IL‐4 in this respect, and the combination of IL‐10 and IL‐4 had an additive effect. In addition, IL‐10 directly stimulated cartilage PG synthesis.
Conclusion. IL‐10 reverses the cartilage degradation induced by antigen‐stimulated MNC, and IL‐4 has an additive effect on this process. Furthermore, IL‐10 has a direct stimulatory effect on PG synthesis, and IL‐4, as a growth factor for type 2 T cells, can reduce the ratio of type 1 to type 2 T cell activity. These results provide evidence in favor of the use of a combination of the two cytokines in the treatment of RA.
This study was designed to investigate whether methotrexate (MTX), used in the treatment of rheumatoid arthritis (RA), affects proliferation and differentiation of human osteoblasts in culture. The ...effects of MTX were assessed by analyzing markers of proliferation and differentiation of human trabecular bone-derived osteoblast-like cells cultured in the presence or absence of 1,25-dihydroxyvitamin D3 (1,25OH2D3). Treatment of the osteoblastic cells with MTX resulted in a strong dose-dependent inhibition of cell proliferation with half maximal response at a dose of 30 nM. MTX did not interfere with cellular alkaline phosphatase (AP) activity, the number of cells expressing cytochemical AP, or basal osteocalcin production. Addition of 1,25(OH)2D3 to the cultures caused an enhanced AP expression and osteocalcin production coinciding with a decreased osteoblast proliferation. Coincubation of 1,25(OH)2D3 with MTX in doses > or = 100 nM further inhibited osteoblast growth and induced a significant stimulation of AP expression and activity, and production of osteocalcin above the values reached in the 1,25(OH)2D3 cultures. In conclusion, MTX proved to be a potent inhibitor of osteoblast proliferation but did not affect basal osteoblastic phenotypic expression. In the presence of the osteoblast differentiation-promoter, 1,25(OH)2D3, MTX further inhibited cell growth which was associated with enhanced AP activity and osteocalcin production. Thus, MTX may have profound effects on bone metabolism and remodeling by interfering with bone cell turnover.
Objective
To determine whether monocyte/macrophages from rheumatoid arthritis (RA) patients can be selectively eliminated by a toxin‐conjugated antibody CD64–ricin A (CD64‐RiA) directed toward the ...high‐affinity receptor for IgG (FcγRI), exploiting the capacity of FcγRI to efficiently endocytose antibody which it has bound.
Methods
Mononuclear cells from peripheral blood (PB) and synovial fluid (SF) obtained from RA patients were cultured in the presence of CD64‐RiA. Cell death of monocyte/macrophages was measured by phenotypic changes (light‐scatter patterns and CD14 and FcγRI expression) and apoptosis (nuclear DNA fragmentation). We then tested whether CD64‐RiA–induced cell death of macrophages affected their capacity to stimulate antigen‐induced lymphocyte proliferation and to secrete cytokines. Additionally, the capacity of CD64‐RiA to inhibit proinflammatory activity and cartilage degradation by RA synovial tissue explants was evaluated.
Results
Inflammatory macrophages from RA SF expressed elevated levels of FcγRI and were selectively eliminated by CD64‐RiA via apoptotic cell death. Monocyte/macrophages from RA PB, which had lower levels of FcγRI expression, were much less affected. Induction of SF macrophage apoptosis was associated with efficient inhibition of antigen‐induced lymphocyte proliferation and a reduction in tumor necrosis factor α (TNFα) release. Consistent with these effects on SF macrophages, CD64‐RiA also inhibited TNFα production, interleukin‐1β production, and cartilage‐degrading activity of RA synovial tissue explants.
Conclusion
Together, these data underscore the crucial role of synovial macrophages in RA joint inflammation and indicate that selective elimination of these cells through FcγRI‐directed immunotoxins could be a novel approach to the treatment of RA.
Removal of the corpuscles of Stannius (STX) in the freshwater European eel causes a marked increase in the concentrations of blood ionic calcium and protein-bound calcium. The hypercalcaemia peaks 20 ...days after STX and lasts at least another 20 days. In stanniectomized eels hypocalcin decreased both blood ionic and total calcium concentrations. The reduction of plasma total calcium concentration by hypocalcin is attributed to a reduction in blood ionic calcium concentration. We conclude that hypocalcin regulates blood ionic calcium levels in fish.
OBJECTIVES The influence of dexamethasone on interleukin 10 (IL10) production and the type 1 (T1)/type 2 (T2) T cell balance found in rheumatoid arthritis (RA) was studied. METHODS Peripheral blood ...mononuclear cells (PB MNC) were isolated from 14 RA patients both before and 7 and 42 days after high dose dexamethasone pulse therapy. The ex vivo production of IL10, interferon GAMMA (IFNGAMMA) (T1 cell), and IL4 (T2 cell) by PB MNCs was assessed, along with parameters of disease activity (erythrocyte sedimentation rate, C reactive protein, Visual Analogue Scale, Thompson joint score). In addition, the in vitro effect of dexamethasone (0.02, 0.2, and 2 μM) on PB MNC IL10, IFNGAMMA, and IL4 production was studied. RESULTS Dexamethasone pulse therapy resulted in a rapid and sustained decrease in RA disease activity. IL10 production increased after dexamethasone treatment and this was sustained for at least six weeks. A transient strong decrease in IFNGAMMA was seen shortly after corticosteroid treatment, while IL4 only decreased slightly. This led to an increased IL-4/IFNGAMMA ratio. In vitro, IL10 production was not detectable, IFNGAMMA and IL4 decreased, but the effect was more pronounced for IFNGAMMA than for IL4, which again resulted in an increased IL4/IFNGAMMA ratio. CONCLUSION Dexamethasone therapy in RA patients leads to a rapid, clinically beneficial effect. The upregulation of IL10 production may be involved in the prolonged clinical benefit. The strong immunosuppressive effect is most evident in the decrease in IFNGAMMA, and is therefore accompanied by a relative shift towards T2 cell activity. In vitro evaluation showed that this shift in T cell balance was a direct effect of dexamethasone and thus independent of the hypothalamic-pituitary-adrenal axis.
Products of the Stannius corpuscles (SC) of rainbow trout were tested in an established PTH bioassay involving bone resorption in embryonic mouse calvaria. Aqueous extracts from Stannius corpuscles ...(SC-homogenate) showed a bone-resorbing activity comparable to PTH in 24-h cultures of calvaria, indicated by a dose-dependent stimulation of lactate production and of calcium, phosphate as well as beta-glucuronidase release. Moreover, SC-homogenates induced an increase in osteoclastic activity. The PTH-like SC-principle is released during in vitro incubations of the glands. These results and the lack of an additive effect of SC-products and PTH on bone resorption suggest that both products activate the same receptor. We hypothesize that the hypocalcemic hormone of the SC of fish shares structural resemblance with PTH, the major hypercalcemic hormone of terrestrial vertebrates.
Objective
Human Hsp60 is expressed in the joints of patients with rheumatoid arthritis (RA) and can elicit a regulatory T cell response in the peripheral blood and synovial fluid. However, Hsp60 can ...also trigger strong proinflammatory pathways. Thus, to understand the nature of these Hsp60‐directed responses in RA, it is necessary to study such responses at the molecular, epitope‐specific level. This study was undertaken to characterize the disease specificity and function of pan–DR‐binding Hsp60–derived epitopes as possible modulators of autoimmune inflammation in RA.
Methods
Lymphocyte proliferation assays (using 3H‐thymidine incorporation and carboxyfluorescein diacetate succinimidyl ester CFSE staining) and measurement of cytokine production (using multiplex immunoassay and intracellular staining) were performed after in vitro activation of peripheral blood mononuclear cells from patients with RA, compared with healthy controls.
Results
A disease (RA)–specific immune recognition, characterized by T cell proliferation as well as increased production of tumor necrosis factor α (TNFα), interleukin‐1β (IL‐1β), and IL‐10, was found for 3 of the 8 selected peptides in patients with RA as compared with healthy controls (P < 0.05). Intracellular cytokine staining and CFSE labeling showed that CD4+ T cells were the subset primarily responsible for both the T cell proliferation and the cytokine production in RA. Interestingly, the human peptides had a remarkably different phenotype, with a 5–10‐fold higher IL‐10:TNFα ratio, compared with that of the microbial peptides.
Conclusion
These results suggest a disease‐specific immune‐modulatory role of epitope‐specific T cells in the inflammatory processes of RA. Therefore, these pan–DR‐binding epitopes could be used as a tool to study the autoreactive T cell response in RA and might be suitable candidates for use in immunotherapy.
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