Summary
Extracellular matrix vesicles from bovine fetal alveolar bone and from a dog osteosarcoma were isolated by differential centrifugation and then fractionated on a discontinuous sucrose density gradient. The fractions were examined by electron microscopy and were analyzed for protein, alkaline phosphatase, aminotripeptidase, and four different β-naphthylamidase activities. The low-density peak of enzyme activities was shown by electron microscopy to be much more homogeneous than the crude matrix vesicle fraction. Two major peaks of protein and enzyme activities were present, one in the high and one in the low density layers. There was good correlation between the activities of alkaline phosphatase and the various peptidases in the fractions from the sucrose density gradient. These results indicate a coexistence of peptidase and alkaline phosphatase in matrix vesicles. On the other hand, there was generally no correlation between the peptidase and alkaline phosphatase activities in vesicular specimens from bovine liver obtained in the same way. Most of the peptidase activity and about half of the alkaline phosphatase activity were solubilized from bone matrix vesicles by detergents. The extracted alkaline phosphatase and alanyl β-naphthylamidase activities were separated from each other on a DEAE-cellulose column.
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References
Anderson HC (1967) Electron microscopic studies of induced cartilage development and calcification. J Cell Biology 35:81–92
Bonuci E (1967) Fine structure of early cartilage calcification. J Ultrastruct Res 20:33–39
Sela J, Bab IA, Muhlrad A (1978) Ultrastructural and biochemical characterization of extracellular matrix vesicles in healing alveolar bone sockets: Preliminary indications for the presence of contractile proteins. Metab Bone Dis Rel Res 1:185–192
Muhlrad A, Stein H, Bab IA, Sela J (1978) Fine structure and enzymes of matrix vesicles in osteosarcoma: Possible occurrence of contractile proteins. Metab Bone Dis Rel Res 1:227–233
Hunziger EB, Hermann W, Schenk RK, Maiti T, Muller M, Moor H (1981) Structural integration of matrix vesicles in calcifying cartilage after cryofixation and freeze substitution. Proc Third Intl Conf on Matrix Vesicles. Monteluco, Spoleto June 7–11 Wichtig Editore, Milan, pp 25–31
Ornoy A, Zusman D, Atkin L (1981) Scanning and transmission electron microscopic studies of matrix vesicles in cartilage and bone of young rats and mice. Proc Third Intl Conf on Matrix Vesicles. Monteluco, Spoleto June 7–11 Wichtig Editore, Milan, pp 13–18
Ali SY, Sajdera SW, Anderson HC (1970) Isolation and characterization of calcifying matrix vesicles from epiphyseal cartilage. Proc Natl Acad Sci (USA) 67:1513–1520
Majeska RJ, Wuthier RE (1975) Studies on matrix vesicles isolated from chick epiphyseal cartilage. Association of pyrophosphatase and ATPase activities with alkaline phosphatase. Biochim Biophys Acta 391:51–60
Felix R, Fleisch H (1976) Pyrophosphatase and ATPase of isolated cartilage matrix vesicles. Calcif Tissue Res 22:1–7
Peress NS, Anderson HC, Sajdera SW (1974) The lipids of matrix vesicles from bovine fetal epiphyseal cartilage. Calcif Tissue Res 14:275–281
Wuthier RE, Gore ST (1977) Partition of inorganic ions and phospholipids in isolated cell, membrane, and matrix vesicle fractions: Evidence for Ca-Pi-acidic phospholipid complexes. Calcif Tissue Res 24:163–171
Siegel SA (1982) The mechanism of ATP-dependent matrix vesicle calcification. PhD Thesis, Department of Biochemistry, School of Graduate Studies, SUNY, Downstate Medical Center, Brooklyn, NY
Lipp W (1959) Aminopeptidase in bone cells. J Histochem Cytochem 7:205
Hirschman A, Hirschman M (1977) Naphthylamidase (arylaminopeptidase) activities in rat cartilage and bone. Enzyme 22:2–12
Hirschman A, Hirschman M (1971) Aminopeptidase profile and protease activity in rat cartilage at physiological pH. Isr J Med Sci 7:403–405
Deutsch D, Bab I, Muhlrad A, Sela J (1981) Purification and further characterization of isolated matrix vesicles from rat alveolar bone. Metab Bone Dis Rel Res 3:209–214
Bradford NM (1976) A rapid and sensitive method for the quantification of microgram quantities of protein, using the principle of protein dye binding. Anal Biochem 72:248–254
Matheson AT, Tattrie BL (1964) A modified Yemm and Cocking ninhydrin reagent for peptide assay. Can J Biochem 42:95–103
Arsenis C, Hackett MH, Huang SM (1976) Resolution, specificity and transphosphorylase activity of calcifying cartilage alkaline phosphatases. Calcif Tissue Res 20:159–171
Fortuna R, Anderson HC, Carty RP, Sajdera SW (1978) The purification and molecular characterization of alkaline phosphatases from chondrocytes and matrix vesicles of bovine fetal epiphyseal cartilage. Metab Bone Dis Rel Res 1:161–168
Sajdera SW, Franklin S, Fortuna R (1976) Matrix vesicles of bovine fetal cartilage: Metabolic potential and solubilization with detergents. Fed Proc 35:154–155
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Hirschman, A., Deutsch, D., Hirschman, M. et al. Neutral peptidase activities in matrix vesicles from bovine fetal alveolar bone and dog osteosarcoma. Calcif Tissue Int 35, 791–797 (1983). https://doi.org/10.1007/BF02405125
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DOI: https://doi.org/10.1007/BF02405125