Upregulated ank expression in osteoarthritis can promote both chondrocyte MMP-13 expression and calcification via chondrocyte extracellular PP i excess

Abstract

Objective: In idiopathic chondrocalcinosis and in osteoarthritis (OA), increased extracellular PP i (ecPP i) promotes calcification. In chromosome 5p-associated familial chondrocalcinotic degenerative arthropathy, certain mutations in the membrane protein ANK may chronically raise ecPP i via enhanced PP i channeling. Therefore, we assessed if dysregulated wild-type ANK expression could contribute to pathogenesis of idiopathic degenerative arthropathy through elevated ecPP i. Design: Using cells with genetic alterations in expression of ANK and the PP i-generating nucleotide pyrophosphatase phosphodiestrase (NPP) PC-1, we examined how increased ANK expression elevates ecPPI, testing for codependent effects with PC-1. We also evaluated the effects of ANK expression on chondrocyte growth, matrix synthesis, and MMP-13 expression and we immunohistochemically examined ANK expression in situ in human knee OA cartilages. Results: Using cells expressing defective ANK, as well as PC-1 knockout cells, we demonstrated that ANK required PC-1 (and vice versa) to raise ecPP i and that the major ecPP i regulator TGFβ required both ANK and PC-1 to elevate ecPP i. Upregulation of wild-type ANK by transfection in normal chondrocytes not only raised ecPP i 5-fold to ∼100 nM but also directly stimulated matrix calcification and inhibited collagen and sulfated proteoglycans synthesis. In addition, upregulated ANK induced chondrocyte MMP-13, an effect that also was stimulated within 2 h by treatment of chondrocytes with 100 nM PP i alone. Finally, ANK expression was upregulated in situ in human knee OA cartilages. Conclusion: Elevation of ecPP i by ANK critically requires the fraction of cellular PP i generated by PC-1. The upregulation of ANK expression in OA cartilage and the capacity of increased ANK expression to induce MMP-13 and to promote matrix loss suggest that increased ANK expression and ecPP i exert noxious effects in degenerative arthropathies beyond stimulation of calcification.