Activation and inhibition of endogenous matrix metalloproteinases in articular cartilage: effects on composition and biophysical properties.
Bovine cartilage explants were cultured with 1 mM 4-aminophenylmercuric acetate (APMA) to activate endogenous matrix metalloproteinases (MMPs) and changes in biochemical, biomechanical, and physicochemical properties were assessed. Additionally, graded levels of either rhTIMP-1 (recombinant human tissue inhibitor of metalloproteinases-1) or L-696-418 (a synthetic metalloproteinase inhibitor) were used to inhibit degradation induced by APMA. Treatment with APMA resulted in as much as 80% loss in tissue GAG content, a greater than threefold increase in denatured type II collagen as determined by the presence of CB11B epitope, and complete loss of biosynthetic activity after 3 days in culture. Physicochemical studies revealed that APMA treatment resulted in a significant increase in tissue swelling response, consistent with damage to the collagen network. Activation of MMPs by APMA also resulted in > 80% decrease in equilibrium modulus, dynamic stiffness, and streaming potential and > 50% decrease in electrokinetic coupling coefficient. The addition of 4 microM, 400 nM, and 40 nM TIMP inhibited PG loss by 95, 50, and 20%, respectively, and all doses effectively inhibited swelling response. The addition of 4 microM and 400 nM L-696-418 inhibited PG loss by 95% while 40 nM L-696-418 inhibited PG loss by 60%, and all doses effectively inhibited swelling response. The inhibition of APMA-induced GAG loss by 4 microM TIMP was accompanied by maintenance of streaming potential, electrokinetic coupling coefficient, dynamic stiffness, and equilibrium modulus.
Bonassar
L J
LJ
Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge 02139, USA.
Stinn
J L
JL
Paguio
C G
CG
Frank
E H
EH
Moore
V L
VL
Lark
M W
MW
Sandy
J D
JD
Hollander
A P
AP
Poole
A R
AR
Grodzinsky
A J
AJ
D000818:Animals; D000831:Animals, Newborn; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D002417:Cattle; D003094:Collagen / Q000378:metabolism; D004151:Dipeptides / Q000494:pharmacology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors; D005109:Extracellular Matrix / Q000201:enzymology*; D006023:Glycoproteins / Q000494:pharmacology*; D006025:Glycosaminoglycans / Q000096:biosynthesis; D006801:Humans; D007700:Kinetics; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009924:Organ Culture Techniques; D010662:Phenylmercuric Acetate / Q000031:analogs & derivatives* / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D019714:Tissue Inhibitor of Metalloproteinases
D004151:Dipeptides; D004791:Enzyme Inhibitors; D006023:Glycoproteins; D006025:Glycosaminoglycans; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D013439:Sulfhydryl Reagents; D019714:Tissue Inhibitor of Metalloproteinases; C085083:N-(1-carboxyethyl)-alpha-(2-phenylethyl)glycyl-leucine, N-phenylamide; C010024:4-aminophenylmercuriacetate; D003094:Collagen; D008666:Metalloendopeptidases; D010662:Phenylmercuric Acetate