Modulation of matrix metalloprotease 13 (collagenase 3) gene expression in equine chondrocytes by interleukin 1 and corticosteroids.
OBJECTIVE
To determine whether matrix metalloprotease 13 (MMP-13; collagenase 3) is produced by equine chondrocytes and to investigate modulation of its expression by recombinant human interleukin 1 beta (rhIL-1 beta) and corticosteroids.


METHODS
Equine chondrocytes in monolayer culture were stimulated with rhIL-1 beta. Total RNA was extracted, purified, and reverse transcribed into DNA. Using appropriate primers, a putative MMP-13 fragment was amplified by polymerase chain reaction, and cloned into a bacterial vector. The resultant fragment was purified and sequenced, then was used to prepare a digoxigenin-labeled cRNA probe. Monolayer cultures of first-passage chondrocytes were treated with rhIL-1 beta in the presence or absence of dexamethasone (10(-6)M) or methylprednisolone acetate (10(-9)M to 10(-5)M), in addition to positive and negative controls. Cellular RNA was extracted and resolved on agarose gels and subjected to northern blot analysis, using the equine MMP-13 probe.


RESULTS
Reverse transcriptase-polymerase chain reaction enabled isolation of a 0.6-kb fragment of equine MMP-13 cDNA that had 93% homology with the human MMP-13 cDNA sequence. rhIL-1 significantly stimulated MMP-13 expression in the chondrocytes. Methylprednisolone acetate inhibited the stimulatory effects of rhIL-1 in dose-dependent manner that was statistically significant at 10(-5)M.


CONCLUSIONS
Novel information was gained on the existence of MMP-13 and its expression in equine chondrocytes, which suggests a possible role for this enzyme in matrix degradation in horses with arthritis.
Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing 48824, USA.
J P
JP
Caron
G
G
Tardif
J
J
Martel-Pelletier
J A
JA
DiBattista
C
C
Geng
J P
JP
Pelletier
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002358:Cartilage, Articular / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D017364:Collagenases / Q000096:biosynthesis / Q000235:genetics*; D003907:Dexamethasone / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005938:Glucocorticoids / Q000494:pharmacology*; D006736:Horses; D007375:Interleukin-1 / Q000494:pharmacology*; D053509:Matrix Metalloproteinase 13; D008775:Methylprednisolone / Q000031:analogs & derivatives* / Q000494:pharmacology; D000077555:Methylprednisolone Acetate; D008969:Molecular Sequence Data
D005938:Glucocorticoids; D007375:Interleukin-1; D000077555:Methylprednisolone Acetate; D003907:Dexamethasone; D017364:Collagenases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13; D008775:Methylprednisolone