| The objective of this work was to investigate the role of cytokines in rheumatoid arthritis (RA) and to compare cytokine{09}profiles, observed in RA, with those in allergic diseases, such as allergic asthma, atopic dermatitis and hymenoptera allergy.; In order to investigate cytokines, we applied a single cell flow cytometric technique that determines intracellular blocked cytokine production. Appropriate inhibitoon of secretion of the produced cytokines is required for studying intracellular cytokine expression by flow cytometry. In the first part of the study (Chapter II), the efficiency of the most widely used inhibitors of cytokine secretion, the antibiotics monensin and brefeldin A, was evaluated.; The intracellular flow cytometric method can identify the cytokine producing cell type but gives no absolute quantitative measurement of the produced cytokine. In Chapter III, we investigated whether the technique that determines intracellular cytokine production is well correlated with extracellular cytokine determination.; Chapter IV presents a transversal study, in which we applied the intracellular flow cytometric method to examine cytokine production in patients with different types of immune mediated disorders.; In Chapter V, the influence of immunotherapy on the cytokine profile in T-lymphocytes was evaluated in a type 2 allergic disease, in casu hymenoptera venom allergy.; In Chapter VI, the influence of therapy with methotrexate in combination with low dose corticosteroids on T-cell cytokine profiles was evaluated in RA patients.; In Chapter VII, the influence of anti-TNF-α therapy on cytokine profiles in T-lymphocytes and monocytes of RA patients was examined, to further investigate the possible mechanísm, responsible for the clinical efficacy of anti-TNF-α therapy.; In the next Chapter (Chapter VIII), the direct influence of pro-inflammatory (IL-1α, IL-6, TNF-α, IFN-γ) and anti-inflammatory (IL-4) cytokines on chondrocyte survival (cell proliferation/necrosis/apoptosis), proliferation and nitric oxide production was investigated in an in vitro model.; Finally, Chapter IX focused on the interaction between chondrocytes, pro-inflammatory cytokines and immune complexes (IC). We observed a significant decrease in chondrocyte proliferation and viability, when the cells were incubated Krith sera and synovial fluid IC derived from RA patients. Moreover, IC derived from RA sera and synovial fluid, were able to induce apoptosis of chondrocytes, determined with three independent techniques (TUNEL/annexine-V/microscopy). Thus, immune complex mediated apoptosís of cytokine-stimulated chondrocytes, might be at least in part responsible for the cartilage breakdown in rheumatoid arthritis. (Abstract shortened by UMI.)... |
