Dataset Information

A single blunt impact on cartilage promotes fibronectin fragmentation and upregulates cartilage degrading stromelysin-1/matrix metalloproteinase-3 in a bovine ex vivo model.

ABSTRACT: Post-traumatic osteoarthritis (PTOA) is characterized by progressive cartilage degeneration in injured joints. Since fibronectin-fragments (Fn-fs) degrade cartilage mainly through up-regulating matrix metalloproteinases (MMPs) and pro-inflammatory cytokines, we hypothesized that Fn-fs play a key role in PTOA by promoting chondrolysis in and around injured cartilage. To test this hypothesis, we profiled the catabolic events focusing on fibronectin fragmentation and proteinase expression in bovine osteochondral explants following a single blunt impact on cartilage with a drop tower device which created partial-thickness tissue damage. Injured and control explants were cultured for up to 14 days. The presence of Fn-fs, MMPs (-1, -3, -13), ADAMTS-5 in culture media and in cartilage was determined with immunoblotting. The daily proteoglycan (PG) depletion of cartilage matrix was assessed with DMMB assay. The effect of explant-conditioned media on chondrocytes was also examined with immunoblotting. Impacted cartilage released significantly higher amount of native Fn, three chondrolytic Fn-fs and PG than non-impacted controls did. Those increases coincided with up-regulation of MMP-3 both in culture media and in impacted cartilage. These findings support our hypothesis that PTOA may be propelled by Fn-fs which act as catabolic mediators through up-regulating cartilage-damaging proteinases.

SUBMITTER: Ding L

PROVIDER: S-EPMC4034576 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

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A single blunt impact on cartilage promotes fibronectin fragmentation and upregulates cartilage degrading stromelysin-1/matrix metalloproteinase-3 in a bovine ex vivo model.

Ding Lei L Guo Danping D Homandberg Gene A GA Buckwalter Joseph A JA Martin James A JA

Journal of orthopaedic research : official publication of the Orthopaedic Research Society 20140307 6

Post-traumatic osteoarthritis (PTOA) is characterized by progressive cartilage degeneration in injured joints. Since fibronectin-fragments (Fn-fs) degrade cartilage mainly through up-regulating matrix metalloproteinases (MMPs) and pro-inflammatory cytokines, we hypothesized that Fn-fs play a key role in PTOA by promoting chondrolysis in and around injured cartilage. To test this hypothesis, we profiled the catabolic events focusing on fibronectin fragmentation and proteinase expression in bovine ...[more]

PMID: 24610678

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Project description:The action of purified rabbit bone stromelysin was investigated on proteoglycan aggregates from pig laryngeal cartilage. The enzyme caused a rapid fall in viscosity of proteoglycan aggregate solution (6 mg/ml), and the products of a partial digest (60% loss of relative viscosity) and a complete digest (95% loss of relative viscosity) were characterized. Analysis by gel chromatography on Sepharose 2B under associative conditions showed that 95% of the glycosaminoglycans in the complete digest were in small-sized fragments, whereas most of the hyaluronan-binding G1 domain and link protein remained intact and bound to hyaluronan. In contrast, there was extensive digestion of the G2 domain which resulted in 76% loss in its detection by immunoassay. Analysis of the partial digest also showed considerable loss (40%) of detection of the G2 domain, but the glycosaminoglycan-rich fragments were much larger than in the complete digest. There was also much less cleavage to create small fragments containing the G1 domain. This was evident on SDS/PAGE analysis where a 58 kDa G1 domain fragment was abundant in the complete digest, but was only present in small amounts in the partial digest. There was also only very limited conversion of link protein from a 44 kDa form to a 40 kDa form. The digestion of proteoglycan aggregate (6 mg/ml) by stromelysin was unaffected by the addition of a high concentration of extra chondroitin sulphate chains (14 mg/ml), and the digestion of proteoglycan monomer showed that the G1 domain was resistant to stromelysin digestion even when not bound to hyaluronan and link protein. The results show that stromelysin degrades the proteoglycan protein core with major cleavages close to, but not within, the G1 domain, and extensive cleavage in other regions. Experiments with purified collagenase, a metalloproteinase structurally related to stromelysin, showed that it too cleaved proteoglycan at several sites within the glycosaminoglycan-rich region of the core protein. Metalloproteinase attack on proteoglycan thus not only occurs with stromelysin but also with collagenase.

Dataset Information

A single blunt impact on cartilage promotes fibronectin fragmentation and upregulates cartilage degrading stromelysin-1/matrix metalloproteinase-3 in a bovine ex vivo model.

Publications

A single blunt impact on cartilage promotes fibronectin fragmentation and upregulates cartilage degrading stromelysin-1/matrix metalloproteinase-3 in a bovine ex vivo model.

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