Assessment of cartilage degradation effects of matrix metalloproteinase-13 in equine cartilage cocultured with synoviocytes.
OBJECTIVE
To determine the effects of matrix metalloproteinase (MMP)-13, compared with interleukin (IL)-1alpha, on cartilage matrix molecule gene expression in a coculture system of equine cartilage explants and synoviocytes.


METHODS
Articular cartilage and synovium specimens harvested from femoropatellar joints of 4 horses, aged 3 to 5 years.


METHODS
Synoviocytes were isolated and cocultured with cartilage explants. Cultures were treated with human recombinant MMP-13 (1, 25, or 100 ng/mL) or IL-1alpha (0.01, 0.1, 1.0, or 10 ng/mL) for 96 hours, with medium exchange at 48 hours. Cartilage extracts and media were analyzed for glycosaminoglycan (GAG) content, and results were adjusted to cartilage DNA content. Quantitative PCR was performed on mRNA from cartilage (MMP-3, MMP-13, aggrecan, and collagen type IIB [COL2A1]) and synoviocytes (MMP-3 and MMP-13), and results were adjusted to 18S ribosomal subunit mRNA expression. Treatments were performed in triplicate, and the experiment was repeated 4 times.


RESULTS
Cultures treated with MMP-13 or IL-1alpha had increased media GAG concentration at 48 and 96 hours. Aggrecan and COL2A1 mRNA expression were increased by application of MMP-13 or IL-1alpha. Gene expression of the catabolic mediator, MMP-3, in cartilage and synoviocytes was increased in cultures treated with MMP-13 or IL-1alpha. Expression of MMP-13 mRNA in cartilage was increased by IL-1alpha, but decreased in synoviocytes by MMP-13 treatment.


CONCLUSIONS
Results support the use of recombinant MMP-13 in a coculture system of synoviocytes and cartilage explants for the study of osteoarthritis.
Fortier
Lisa A
LA
Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, USA.
Schnabel
Lauren V
LV
Mohammed
Hussni O
HO
Mayr
Karen G
KG
D000704:Analysis of Variance; D000818:Animals; D002356:Cartilage / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects*; D006736:Horses / Q000378:metabolism*; D053509:Matrix Metalloproteinase 13 / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology*; D013997:Time Factors
D016326:Extracellular Matrix Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D053509:Matrix Metalloproteinase 13