Matrix vesicles (MVs) in the growth plate bind to cartilage collagens and initiate mineralization of the extracellular matrix. Native MVs have been shown to contain a nucleational core responsible for mineral formation that is comprised of Mg 2؉ -containing amorphous calcium phosphate and lipid-calciumphosphate complexes (CPLXs) and the lipid-dependent Ca 2؉ -binding proteins, especially annexin-5 (Anx-5), which greatly enhances mineral formation. Incorporation of non-Ca 2؉ -binding MV lipids impedes mineral formation by phosphatidylserine (PS)-CPLX. In this study, nucleators based on amorphous calcium phosphate (with or without Anx-5) were prepared with PS alone, PS ؉ phosphatidylethanolamine (PE), or PS ؉ PE and other MV lipids. These were incubated in synthetic cartilage lymph containing no collagen or containing type II or type X collagen. Dilution of PS with PE and other MV lipids progressively retarded nucleation. Incorporation of Anx-5 restored nucleational activity to the PS:PE CPLX; thus PS and Anx-5 proved to be critical for nucleation of mineral. Without Anx-5, induction of mineral formation was slow unless high levels of Ca 2؉ were used. The presence of type II collagen in synthetic cartilage lymph improved both the rate and amount of mineral formation but did not enhance nucleation. This stimulatory effect required the presence of the nonhelical telopeptides. Although type X collagen slowed induction, it also increased the rate and amount of mineral formation. Both type II and X collagens markedly increased mineral formation by the MV-like CPLX, requiring Anx-5 to do so. Thus, Anx-5 enhances nucleation by the CPLXs and couples this to propagation of mineral formation by the cartilage collagens.
Matrix vesicles (MVs),2 extracellular lipid bilayer-enclosed microstructures released by calcifying cells, initiate mineral formation in newly forming bone (1-4); there is little evidence that they play a role during the remainder of mineral deposition under the aegis of osteoblasts. However, MVs also appear to initiate ectopic calcification in calcific tendonitis, apatite-deposition osteoarthritis, cardiac valve calcification, and atherosclerotic lesions (5-8). MVs interact with both the matrix collagens (9 -11) and proteoglycans (12, 13) in the extracellular matrix. The interaction between MVs and the matrix collagens has been shown to be mediated by annexin 5 (14 -16), the major protein in MV (17-22). MV are enriched in phosphatidylserine (PS) (23, 24), a lipid that has high affinity for Ca 2ϩ (25,26) and is able to form complexes with both Ca 2ϩ and P i (27). Such complexes have the ability to nucleate hydroxyapatite formation (28 -32). PS is initially localized to the internal membrane of MV (33) where it is found in PS:Ca 2ϩ :P i complexes (CPLXs) (34). However, during programmed cell death (apoptosis) PS becomes externalized in a variety of cells (35,36); in fact externalization of PS is widely used to identify apoptotic cells (37,38). Although there are similarities between apoptotic bodies and MV, th...