l-663536 and Arthritis--Rheumatoid

To investigate quantification of expression of LTB4 inducing IL-1beta and TNF-alpha at mRNA level in synovial membrane cells of rheumatoid arthritis.. Primary cultured synovial cells from RA patients were treated with exogenous LTB4, MK-886 (inhibitor of 5-lipoxygenase activating protein) and Bestatin(inhibitor of leukotriene A4 hydrolase) in the presence of LIT respectively, expressions of TNF-alpha and IL-1beta were detected at mRNA level by Real-time Quantitative PCR.. Expressions of basic TNF-alpha (TNF-alpha/GAPDH) and IL-beta (IL-beta/GAPDH) at mRNA level in primary cultured synovial cells were 0.02 +/- 0.00 and 0.16 +/- 0.01 respectively. LTB4 (10(-9) mol/L-10(-8) mol/L) was shown to induce dose-dependent increase of mRNA expression of TNF-alpha. (7-15 times) and IL-1beta (1 time) , endogenous product of LTB4 by LIT significantly increased mRNA expressions of TNF-alpha (145 times) and IL-1beta (12 times) respectively. LIT-treated synoviocytes with addition of MK-886 (5-LOX exciting protein FLAP inhibitor) (1-10 micromol/L) were inhibited to secrete LTB4 dose-dependently, following the markedly down-regulated expressions of TNF-alpha (15%-66%) and IL-1beta (41%-71%) at mRNA level . Bestatin(100 mg/L) could also remarkably diminish LTB4-induced mRNA expressions of TNF-alpha(86%) and IL-1beta (79%).. LTB4 of synovial membrance cells in rheumatoid arthritis could induce expressions of TNF-alpha and IL-1beta at mRNA level, and their expression at mRNA level had been quantified successfully. It is a beneficial help to quantify all kinds of cytokines in methodology.

Topics: Arthritis, Rheumatoid; Cells, Cultured; Gene Expression; Humans; Indoles; Interleukin-1beta; Leukotriene B4; Polymerase Chain Reaction; RNA, Messenger; Synovial Membrane; Tumor Necrosis Factor-alpha

IL-18 expression and functional activity have been associated with a range of autoimmune diseases. However, the precise mechanism by which IL-18 induces such pathology remains unclear. In this study we provide direct evidence that IL-18 activates neutrophils via TNF-alpha induction, which drives the production of leukotriene B(4) (LTB(4)), which in turn leads to neutrophil accumulation and subsequent local inflammation. rIL-18 administered i.p. resulted in the local synthesis of LTB(4) and a rapid influx of neutrophils into the peritoneal cavity, which could be effectively blocked by the LTB(4) synthesis inhibitor MK-886 (MK) or its receptor antagonist CP-105,696. IL-18-induced neutrophils recruitment and LTB(4) production could also be blocked by a neutralizing anti-TNF-alpha Ab. In addition, IL-18 failed to induce neutrophil accumulation in vivo in TNFRp55(-/-) mice. In an IL-18-dependent murine collagen-induced arthritis model, administration of MK significantly inhibited disease severity and reduced articular inflammation and joint destruction. Furthermore, MK-886-treated mice also displayed suppressed proinflammatory cytokine production in response to type II collagen in vitro. Finally, we showed that IL-18-activated human peripheral blood neutrophils produced significant amounts of LTB(4) that were effectively blocked by the MK. Together, these findings provide a novel mechanism whereby IL-18 can promote inflammatory diseases.

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Arthritis, Experimental; Arthritis, Rheumatoid; Cells, Cultured; Chemotaxis, Leukocyte; Collagen Type II; Humans; Indoles; Inflammation Mediators; Injections, Intradermal; Injections, Intraperitoneal; Interleukin-18; Leukotriene B4; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Knockout; Neutrophil Infiltration; Neutrophils; Severity of Illness Index; Tumor Necrosis Factor-alpha