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. PROCEDURE: 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.