Induction of collagenase-3 (MMP-13) in rheumatoid arthritis synovial fibroblasts

• Published in: Biochimica et biophysica acta Vol. 1502; no. 2; pp. 307 - 318
• Main Authors: Moore, Bryan A., Aznavoorian, Sadie, Engler, Jeffrey A., Windsor, L.Jack
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 18.10.2000
• Subjects: Antibodies; Arthritis, Rheumatoid - enzymology; Arthritis, Rheumatoid - genetics; Base Sequence; Blotting, Western; Collagenase-3; Collagenases - biosynthesis; Collagenases - genetics; Collagenases - immunology; Cytokines - pharmacology; DNA Primers - genetics; Enzyme Induction; Fibroblasts - metabolism; Humans; Immunohistochemistry; In Vitro Techniques; Matrix metalloproteinase; Matrix Metalloproteinase 1 - biosynthesis; Matrix Metalloproteinase 1 - genetics; Matrix Metalloproteinase 13; Microscopy, Fluorescence; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; RNA, Messenger - metabolism; Synovial fibroblast; Synovial Membrane - enzymology; Tetradecanoylphorbol Acetate - pharmacology; Tissue Inhibitor of Metalloproteinases - genetics; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; RT-PCR, reverse transcriptase-polymerase chain reaction; CL-3, collagenase-3, MMP-13; mAb, monoclonal antibody; bp, base pair(s); IL-1, interleukin-1; Matrix metalloproteinase; Gel B, gelatinase B, M r 92 000 gelatinase, MMP-9; pAb, polyclonal antibody; Synovial fibroblast; MT1-MMP, membrane type-1 MMP, MMP-14; IL-6, interleukin-6; IL-6sR, interleukin-6 soluble receptor; TBS-TX, Tris buffered saline with 0.1% Triton X-100; TIMP, tissue inhibitor of metalloproteinases; CL-1, collagenase-1, fibroblast-type collagenase, MMP-1; TXRD, Texas-red dye; FBS, fetal bovine serum; PBS, phosphate buffered saline; FITC, fluorescein-5-isothiocyanate; SL-1, stromelysin-1, MMP-3; TNF-α, tumor necrosis factor-α; SL-2, stromelysin-2, MMP-10; TGF-β1, transforming growth factor-β1; DAPI, 4,6-diamidino-2-phenylindole; Collagenase-3; MMP, matrix metalloproteinase; Gel A, gelatinase A, M r 72 000 gelatinase, MMP-2; PMA, phorbol 12-myristate 13-acetate
• ISSN: 0925-4439; 0006-3002; 1879-260X
• DOI: 10.1016/S0925-4439(00)00056-9

There is a growing body of evidence that implicates matrix metalloproteinases (MMPs) as major players in numerous diseased conditions. The articular cartilage degradation that is characteristic of rheumatoid arthritis (RA) is believed to be mediated by the collagenase subfamily of matrix metalloproteinases. The preference of collagenase-3 (CL-3) for collagen type II makes it a likely candidate in the turnover of articular cartilage and a potential target for drug development. In this study, RA synovial membrane tissue was shown to express CL-3 mRNA by reverse transcriptase-polymerase chain reaction (RT-PCR) and protein by immunohistochemistry. Fibroblasts isolated and cultured from RA synovial membrane tissue were induced to express CL-3 mRNA. CL-3 mRNA was detected after PMA treatment in 16 of the 18 RA synovial membrane fibroblast cell lines established for this study. These fibroblasts also expressed mRNA for collagenase-1 (CL-1, MMP-1), membrane type-1 matrix metalloproteinase, gelatinase A, gelatinase B, stromelysin-1, stromelysin-2, TIMP-1, and TIMP-2. They were further shown to express CL-1 mRNA constitutively and CL-3 mRNA only after stimulation with PMA, IL-1, TGF-β1, TNF-α, or IL-6 with IL-6sR. These fibroblasts also expressed after induction both CL-1 and CL-3 at the protein level as determined by Western blot analyses and immunofluorescence.