Mechanisms and kinetics of glycosaminoglycan release following in vitro cartilage injury.
OBJECTIVE
Acute joint injury leads to increased risk for osteoarthritis (OA). Although the mechanisms underlying this progression are unclear, early structural, metabolic, and compositional indicators of OA have been reproduced using in vitro models of cartilage injury. This study was undertaken to determine whether glycosaminoglycan (GAG) loss following in vitro cartilage injury is mediated by cellular biosynthesis, activation of enzymatic activity, or mechanical disruption of the cartilage extracellular matrix.


METHODS
Immature bovine cartilage was cultured for up to 10 days. After 3 days, groups of samples were subjected to injurious mechanical compression (single uniaxial unconfined compression to 50% thickness, strain rate 100% per second). GAG release to the medium was measured, and levels were compared with those in location-matched, uninjured controls. The effects of medium supplementation with inhibitors of biosynthesis (cycloheximide), of matrix metalloproteinase (MMP) activity (CGS 27023A or GM 6001), and of aggrecanase activity (SB 703704) on GAG release after injury were assessed.


RESULTS
GAG release from injured cartilage was highest during the first 4 hours after injury, but remained higher than that in controls during the first 24 hours postinjury, and was not affected by inhibitors of biosynthesis or degradative enzymes. GAG release during the period 24-72 hours postinjury was similar to that in uninjured controls, but the MMP inhibitor CGS 27023A reduced cumulative GAG loss from injured samples between 1 day and 7 days postinjury. Other inhibitors of enzymatic degradation or biosynthesis had no significant effect on GAG release.


CONCLUSIONS
Injurious compression of articular cartilage induces an initially high rate of GAG release from the tissue, which could not be inhibited, consistent with mechanical damage. However, the finding that MMP inhibition reduced GAG loss in the days following injury suggests a potential therapeutic intervention.
DiMicco
Michael A
MA
Massachusetts Institute of Technology and Children's Hospital, Boston, Massachusetts 02139, USA. mdimicco@mit.edu
Patwari
Parth
P
Siparsky
Patrick N
PN
Kumar
Sanjay
S
Pratta
Michael A
MA
Lark
Michael W
MW
Kim
Young-Jo
YJ
Grodzinsky
Alan J
AJ
D000818:Animals; D002358:Cartilage, Articular / Q000293:injuries* / Q000473:pathology / Q000503:physiopathology; D002417:Cattle; D002470:Cell Survival; D004151:Dipeptides / Q000494:pharmacology; D010450:Endopeptidases / Q000187:drug effects; D006025:Glycosaminoglycans / Q000378:metabolism*; D006877:Hydroxamic Acids / Q000494:pharmacology; D066298:In Vitro Techniques; D007192:Indenes / Q000494:pharmacology; D007700:Kinetics; D011312:Pressure; D011480:Protease Inhibitors / Q000494:pharmacology; D011719:Pyrazines / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors; D014947:Wounds and Injuries / Q000209:etiology / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology
C097658:CGS 27023A; D004151:Dipeptides; D006025:Glycosaminoglycans; D006877:Hydroxamic Acids; D007192:Indenes; C078131:N-(2(R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl)-L-tryptophan methylamide; D011480:Protease Inhibitors; D011719:Pyrazines; C483299:SB 703704; D013449:Sulfonamides; D010450:Endopeptidases; C083641:aggrecanase