Characterization and identification of proteinases and proteinase inhibitors synthesized by temporomandibular joint disc cells.
The adult mammalian temporomandibular joint (TM) disc is a fibrocartilaginous tissue that undergoes normal developmental remodeling, requiring removal of the existing extracellular matrix and its replacement by new matrix macromolecules. This remodeling is probably mediated by matrix-degrading enzymes, but to date none has been demonstrated in association with the TMJ disc. We characterized, identified, and determined the regulation of proteinases and proteinase inhibitor (PIs) synthesized by TMJ disc cells in organ and cell cultures. TMJ discs were retrieved from 14-week-old male NZW rabbits and both tissue- and disc-derived cells were cultured in serum-free medium. The conditioned media were retrieved at 12-hour intervals and assayed for proteinases and PIs in gelatin- and casein-impregnated polyacrylamide gels. Three proteinases with gelatinolytic activities at 92 kDa, 72 kDa, and 42/57 kDa and one caseinolytic activity at 51/54 kDa were detected. All were inhibited by 1,10-1 phenanthroline, thus characterizing these enzymes as matrix metalloproteinases (MMPs), most likely 92-kDa gelatinase (proMMP-9), 72-kDa gelatinase (proMMP-2), procollagenase (proMMP-1), and prostromelysin (proMMP-3). The identity of the latter two MMPs was confirmed by Western blots. Two PIs and 30 kDa and 20 kDa, probably tissue inhibitors of metalloproteinase (TIMP) and TIMP-2, were observed on reverse zymograms. TPA, a protein kinase-C agonist, increased the expression of 92-kDa gelatinase and 30-kDa PI by both explanted discs and isolated disc cells. The profile of MMPs constitutively expressed by disc cells is similar to that of synovial fibroblasts but different from that of chondrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
Kapila
S
S
Department of Growth and Development, School of Dentistry, University of California, San Francisco 94143-0640, USA.
Lee
C
C
Richards
D W
DW
D000818:Animals; D002358:Cartilage, Articular / Q000201:enzymology*; D002478:Cells, Cultured; D017364:Collagenases / Q000096:biosynthesis / Q000737:chemistry; D046508:Culture Techniques; D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000737:chemistry; D005109:Extracellular Matrix / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D018093:Gelatinases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000737:chemistry; D006651:Histocytochemistry; D015151:Immunoblotting; D008297:Male; D061965:Matrix Metalloproteinase Inhibitors; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000737:chemistry; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000737:chemistry / Q000378:metabolism*; D011817:Rabbits; D013704:Temporomandibular Joint / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004792:Enzyme Precursors; D061965:Matrix Metalloproteinase Inhibitors; D011480:Protease Inhibitors; D017364:Collagenases; D018093:Gelatinases; D008666:Metalloendopeptidases; C022440:procollagenase; C048047:prostromelysin; D013755:Tetradecanoylphorbol Acetate