Abstract
Increased quantities of the mast cell (MC)-restricted β tryptase, mouse MC protease (mMCP)-6 and its human ortholog hTryptase-β are present in the synovial tissue of mouse and human arthritic joints. mMCP-6-deficient mice lose less aggrecan from the articular cartilage during inflammatory arthritis than wild-type (WT) C57BL/6 mice. This thesis therefore aimed to elucidate the underlying mechanisms involved in MC β tryptase-mediated aggrecanolysis. Exposure of mouse femoral head explants ex vivo to rhTryptase-β, rmMCP-6, or lysates harvested from WT mouse peritoneal MCs (PMCs) significantly increased the levels of enzymatically active matrix metalloproteinases (MMPs), and significantly induced aggrecan loss. Treatment of cartilage explants with β tryptases generated aggrecan fragments containing both DIPEN and FFGVG neoepitopes, consistent with MMP-dependent aggrecanolysis. The abilities of mMCP-6 from WT PMCs to induce aggrecanolysis were prevented by inhibitors of MMP-3 and MMP-13, which supported the findings that rhTryptase-β was able to activate proMMP-3 and proMMP-13. To study the in vivo relevance of these findings, samples of arthritic knee joints were obtained after induction of the methylated BSA/IL-1β model of inflammatory arthritis in WT and mMCP-6-deficient C57BL/6 mice. In contrast to arthritic WT mice, the patella explants from mMCP-6-deficient mice exhibited significantly reduced MMP activity and subsequent aggrecan loss. Analysis of patella explants showed substantially greater levels of MMP-derived aggrecan fragments, and activated levels of MMP-3 and MMP-13 in WT mice compared to mMCP-6-deficient mice. Furthermore, the effects of hTryptase-β on the expression of ECM molecules in the chondrocytic ATDC5 cell line were analysed using PCR array. Culture of cells in the presence of rhTryptase-β for 48 h resulted in the down-regulation of Mmp13 and link protein mRNA and protein levels; two essential proteins required for the maintenance of normal cartilage homeostasis. The accumulated data presented in this thesis indicate that β tryptases regulate the activities of specific MMPs at the transcription, translation and post-translation levels, signifying an important pathophysiological pathway. The functions of MC β tryptases within the arthritic joint suggests that therapeutic inhibition may be beneficial in abrogating further joint destruction in patients with rheumatoid arthritis.