Upregulated ank expression in osteoarthritis can promote both chondrocyte MMP-13 expression and calcification via chondrocyte extracellular PPi excess.
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).


METHODS
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 TGFbeta 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 approximately 100nM 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 2h by treatment of chondrocytes with 100nM PP(i) alone. Finally, ANK expression was upregulated in situ in human knee OA cartilages.


CONCLUSIONS
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.
Johnson
K
K
Veterans Affairs Medical Center, UCSD, La Jolla, CA 92161, USA.
Terkeltaub
R
R
D000818:Animals; D002114:Calcinosis / Q000235:genetics / Q000503:physiopathology*; D002358:Cartilage, Articular / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D003094:Collagen / Q000096:biosynthesis; D017364:Collagenases / Q000032:analysis*; D011756:Diphosphates / Q000032:analysis*; D045604:Extracellular Fluid / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D007719:Knee Joint / Q000378:metabolism; D053509:Matrix Metalloproteinase 13; D051379:Mice; D008815:Mice, Inbred Strains; D020370:Osteoarthritis, Knee / Q000235:genetics / Q000503:physiopathology*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011509:Proteoglycans / Q000096:biosynthesis; D011755:Pyrophosphatases / Q000378:metabolism; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000378:metabolism; D015854:Up-Regulation
D011756:Diphosphates; D011509:Proteoglycans; D016212:Transforming Growth Factor beta; D003094:Collagen; D010727:Phosphoric Diester Hydrolases; D017364:Collagenases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13; C507081:Mmp13 protein, mouse; D011755:Pyrophosphatases; C405782:nucleotide pyrophosphatase - phosphodiesterase I