|Budget Amount *help
¥12,700,000 (Direct Cost : ¥12,700,000)
Fiscal Year 1999 : ¥4,100,000 (Direct Cost : ¥4,100,000)
Fiscal Year 1998 : ¥8,600,000 (Direct Cost : ¥8,600,000)
Degradation of the extracellular matrix (ECM) is an essential step to the destruction of articular cartilage in rheumatoid athritis (RA), and members of the matrix metalloproteinase (MMP) gene family are involved in the degradation. In the present studies, we characterized the biochemical properties of some MMP species and examined their functions in the destruction of the cartilage in RA joints. In addition, synthetic MMP inhibitors and tissue inhibitor of metalloproteinases 3 (TIMP-3)-inducible drug were developed by collaboration with pharmaceutical companies. The major results are as follows:
(1). Activation of proMMP-2 mediated by membrane-type1 MMP (MT1-MMP) was accelerated in the presence of TIMP-2, and thus TIMP-2 was considered to be essential to the efficient activation of proMMP-2 on the cell membranes. We also demonstrated that MT1-MMP cleaves aggrecan at three sites in the interglobular domain and MT3-MMP degrades the ECM components such as type III collagen and aggrecan. O
n the other hand, the activities of MT1-MMP and MT3-MMP were not inhibited by TIMP-1. MT-MMP was shown to be shed from the cell membranes of tumor cells treated with concanvalin A.
(2). ProMMP-2 was efficiently activated in RA synovial tissues and its activation ratio directly correlated with the expression levels of MT1-MMP among MT1, 2, 3-MMPs. MT1-MMP was expressed in the lining cells of RA synovial membrane and gelatinolytic activity was detected by in situ zymography in the lining cell layer.
(3). When steady state levels of MMP-1, 2, 3, 7, 8, 9, 13 and TIMP-1, 2 in synovial fluids were assayed by sandwich enzyme immunoassays, the levels of MMP 1, 2, 3, 8, 9 and TIMP-1 were significantly higher in RA than in OA. The molar ratio was significantly higher in RA than in OA, and metalloproteinase activity was detected with a direct correlation to MMP/TIMP molar ratio, suggesting an imbalance in favor of proteinase.
(4). By collaboration with a pharmaceutical company, we developed synthetic inhibitors more selective to MT1-MMP, We also demonstrated that pentosan polysulfate stimulates RA synovial fibroblasts to produce TIMP-3 by the posttranscriptional level. Less