The Effect And The Pharmacologic Mechanism Of Leflunomide On IL-18 Production In Synoviocytes

Objectives: Leflunomide, a potent diseae-modifying antirheumatic drug used in the treatment of rheumatoid arthritis (RA), exhibits anti-inflammatory, antiproliferative and immunosuppressive effects. Although most of the beneficial effects of leflunomide have been attributed to its antimetabolite activity. Interleukin-18(IL-18) plays an important role in the pathogenesis of RA. We carried out this study to investigate the effects of the active metabolite of leflunomide (A77 1726;LEF-M) on interleukin-18(IL-18) production in cultured fibroblast-like synoviocytes (FLS) obtained from patients with rheumatoid arthritis(RA) and the restrainable effects of uridine to leflunomide active metaboliten A77 1726 .Methods: Synovial tissue was obtained during surgery from patients undergoing total knee or elbow metathesis owing to RA or traumatism (NC). Synovial cells were isolated by collagenase digestion, and passages 3-8 cells were used for the experiments. To choice the dose of LEF, we evaluated the synoviocytes viability by methyl thiazalyl tetrazolium salt test(MTT), which incubated with different concentration of LEF ranged from 10 μ g/ml to 160 μg/ml. Intracytoplasmic IL-18 protein production was detected by immunocytochemistry, and extracellular concentration of IL-18 protein was detected by enzyme-linked immunosorbent assay (ELISA) after 48 hours incubated with differentdoses of A77 1726 and stimulated by lip polysaccharide (LPS).Result: l.The level of intracellular and extracellular IL-18 protein expression of RA is higher than that of NC. 2. A77 1726 inhibits IL-18 production in the synoviocytes in a dose dependent manner comparison with untreated cells.3. As the concentration of LEF was no more than 90u g/ml, the inhibitive effect of it can be restrained by uridine. 4. Synoviocytes viability evaluated by MTT was unaffected by the A77 1726 ,which concentration was no more than 120u g/ml.Conclusion: The high expression of IL-18 in RA synoviocytes shows that IL-18 is related with the occurrence and progress of RA;A77 1726 inhibited the production of IL-18 in human synovial fibroblasts in presence of LPS, this might explain some of the protective effects exerted by leflunomide in RA and other autoimmune disease;Exogenous addition of uridine could restrained the effect of LEF(<90 u g/ml), might revealed that LEF might act as the inhibitor of the L-dihydro-orotate dehydrogenase (DHOH) enzyme and pyrimidine denovo synthesis in this dose.