Background Extracellular matrix degradation is one of the principal features of osteoarthritis (OA). Although several proteases are implicated in this process, MMP-13 presents several properties that lead it as a key point: it is the collagenase that degradates Col-II at a higher rate, and its expression is restricted to embriogenesis, cancer and OA conditions. Biomarkers development has been proposed for the OA diagnoses and progression. However, they have shown lack of specificity. We hypothesized that MMP-13 is able to digest cartilage components of the extracellular matrix others than Col-II. Furthermore, the fragments generated by the digestion are plausible OA-related biomarkers due to the restricted MMP-13 expression.
Objectives To extract total soluble protein from cartilage obtained from hip OA of patients undertaken to arthroplasty. To digest the protein extract with activated MMP-13 and to explore the fragments generated from different cartilage matrix components previously related with the OA pathofisiology.
Methods Cartilage samples presenting different degrees of OA development were selected from OA patients after hip replacement. Soluble proteins were extracted with guanidinium chloride method. MMP-13 was activated with APMA and afterward protein digestion was performed at a ratio of 1:100 (MMP:protein) for 16 hours. The digestion pattern of mimecan, asporin, keratocan, COMP, link protein, matrilin and Col-IXa was examined by western-blot.
Results 20 samples of each cartilage condition (slightly, moderate and severe OA) were included and soluble protein extraction performed. Analysis by Western blot of the digestions showed that asporin, keratocan and Link protein were resistant to MMP-13 degradation. However, mimecan, COMP, matrilin and Col-IXa present detectable MMP-13 digestion products. Fibronectin was degradated but no cleavage fragments were observed. No remarkable differences were observed in the digestion patterns between cartilage from different degrees of development. The sizes of the digestion products observed are summarized in the following table.
Conclusions COMP, Col-IXa, matrilin and mimecan present detectable fragments generated by MMP-13 and hence these products could represent new OA-related biomarkers. Since MMP-13 is not able to degrade all cartilage extracellular matrix components, synergies with other proteases are guaranteed to the completely cartilage digestion.
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Disclosure of Interest None declared
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