Summary
Chondrocyte, matrix vesicle, and membrane fractions, as well as interstitial fluid samples from the proliferating and hypertrophic zones of chicken epiphyseal cartilage were analyzed for electrolyte content. Intracellular Ca levels were 1.4–2.1 mM, over 90% of which was nondiffusible. Isolated hypertrophic chondrocytes had higher intracellular Na and lower K than proliferating cells. Matrix vesicles contained 25 to 50 times higher concentrations of Ca than the adjacent cells. Vesicles from the zone of hypertrophy contained twice as much Ca as did those from the proliferating area. Ca/P1 molar ratios of matrix vesicles were much higher than those of cells or of later mineral deposits. These findings indicate that Ca is concentrated in matrix vesicles during formation, but acuumulation of Ca and P1 must continue in the matrix. X-ray diffraction of freeze-dried vesicle and membrane fractions failed to detect crystalline apatite, suggesting that crystals seen in electron micrographs of matrix vesicles may be artifacts. Interstitial fluid expressed from epiphyseal cartilage was higher in K, Pi, Mg and nucleotides, and lower in Na and Cl, than blood plasma. Fluid from the hypertrophic zone was higher in K and nucleotides, but not Pi or Mg, than that from the proliferating layer. These data suggest that selective leakage or extrusion of these constituents, which are normally intracellular, must occur, especially in the hypertrophic zone. More of the Ca and Mg, and less of the Pi, was protein-bound in cartilage fluid than in blood plasma. There was more binding of the divalent cations in fluid from proliferating than from hypertrophic cartilage. The presence of greater amounts of ultrafilterable peptides in fluid from hypertrophic than from proliferating cartilage or blood plasma, suggests that proteolytic activity may release bound divalent cations during mineralization.
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Ali, S.Y.: Analysis of matrix vesicles and their role in the calcification of epiphyseal cartilage. Fed. Proceed.35, 135–142 (1976)
Anderson, H.C.: Vesicles associated with calcification in the matrix of epiphyseal cartilage. J. Cell Biol.41, 59–72 (1969)
Anderson, H.C.: Calcium-accumulating vesicles in the intercellular matrix of bone. In: Hard tissue growth repair and remineralization, p. 213–246. Amsterdam: CIBA Foundation Symposium II, 1973
Anghileri, L.J., Dermietzel, R.: Calcium-phosphate-phospholipid complexes in experimental tumors: Their possible relationship with tumor calcification. Z. Krebsforsch.79, 148–156 (1973)
Asper, S.P., Schales, O., Schales, S.S.: Importance of controlling pH in the Schales and Schales method of chloride determination. J. Biol. Chem.168, 779–780 (1947)
Bonucci, E.: Fine structure and histochemistry of “calcifying globules” in epiphyseal cartilage. Z. Zellforsch.103, 192–217 (1970)
Borle, A.: Effects of purified parathyroid hormone on the calcium metabolism of monkey kidney cells. Endocrinol83, 1316–1322 (1968)
Boskey, A., Posner, A.S.: Extraction of calcium-phospholipid-phosphate complex from bone. Calcif. Tiss. Res.19, 273–284 (1976)
Brighton, C.T., Heppenstall, R.B.: Oxygen tension in zones of the epiphyseal plate, the metaphysis and diaphysis. J. Bone Jt. Surg.53-A, 719–728 (1971)
Brighton, C.T., Hunt, R.M.: Mitochondrial calcium and its role in calcification. Clin. Orthop. Rel. Res.100, 406–416 (1974)
Cotmore, J.M., Nichols, G., Jr., Wuthier, R.E.: Phospholipid-calcium-phosphate complex. Enhanced calcium migration in the presence of phosphate. Science172, 1339–1341 (1971)
Cuervo, L.A., Pita, J.C., Howell, D.S.: Ultramicroanalysis of pH, Pco 2 and carbonic anhydrase activity at calcifying sites in cartilage. Calcif. Tiss. Res.7, 220–231 (1971)
Dziak, R., Brand, J.S.: Calcium transport in isolated bone cells. I. Bone cell isolation procedures. J. Cell. Physiol.84, 75–84 (1974a)
Dziak, R., Brand, J.S.: Calcium transport in isolated bone cells. II. Calcium transport studies. J. Cell Physiol.84, 85–96 (1974b)
Eisenmann, D.R., Glick, P.L.: Ultrastructure of initial crystal formation in dentin. J. Ultrastruct. Tes.41, 18–28 (1972)
Gamble, J.L.: Chemical anatomy, physiology and extracellular fluid, 6th ed. Cambridge: Harvard Univ. Press 1954
Granda, J.L., Posner, A.S.: Distribution of four hydrolases in the epiphyseal plate. Clin. Orthop. Rel. Res.74, 269–272 (1971)
Guest, G.M., Organic phosphates of the blood and mineral metabolism in diabetic acidosis. Am. J. Diseases Children64, 401–412 (1942)
Höhling, H.J., Steffens, H., Stamm, G., Mays, U.: Transmission microscopy of freeze dried, unstained epiphyseal cartilage of the guinea pig. Cell Tiss. Res.167, 243–263 (1976)
Howell, D.S., Delchamps, E., Riemer, W., Kiem, I.: A profile of electrolytes in the cartilaginous plate of growing ribs. J. Clin. Invest.39, 919–929 (1960)
Howell, D.S., Pita, J.C., Marquez, J.F., Madruga, J.E.: Partition of calcium, phosphate and protein in the fluid phase aspirated at calcifying sites in epiphyseal cartilage, J. Clin. Invest.47, 1121–1132 (1968)
Kobayashi, Y., Maudsley, D.V.: Practical aspects of double isotope counting. In: The current status of liquid scintillation counting, (E.D. Bransome, Jr., ed.), pp. 76–85. New York: Grune and Stratton 1970
Linn, F.C., Sokoloff, L.: Movement and composition of interstitial fluid of cartilage. Arth. Rheum.8, 481–494 (1965)
Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J.: Protein measurement with the Folin phenol reagent, J. Biol. Chem.193, 265–276 (1951)
Majeska, R.J., Wuthier, R.E.: Studies on matrix vesicles isolated from chick epiphyseal cartilage: Association of pyrophosphatase and ATPase activities with alkaline phosphatase. Biochim. Biophys. Acta391, 51–60 (1975)
Nichols, G., Jr., Rogers, P.: Bone cell calcium stores: Their size, location and kinetics of exchange. Calcif. Tiss. Res.9, 80–94 (1972)
Peress, N., Sajdera, S.W., Anderson, H.C.: The lipids on matrix vesicles from bovine fetal epiphyseal cartilage. Calcif. Tiss. Res.14, 274–282 (1974)
Rouser, G., Siakotos, A.N., Fleischer, S.: Quantitative analysis of phospholipids by thin-layer chromatography and phosphorus analysis of spots. Lipids1, 85–86 (1966)
Scarpace, P.J., Neuman, W.F.: The blood: bone disequilibrium. 1. The active accumulation of K+ into the bone extracellular fluid. Calcif. Tiss. Res.20, 137–150 (1976)
Sisca, R.F., Provenza, D.V.: Initial dentin formatjion in human deciduous teeth: An electron microscope study. Calcif. Tiss. Res.9, 1–16 (1972)
Vogel, J.J., Boyan-Salyers, B.D.: Acidic lipids associated with the local mechanism of calcification. Clin. Orthop. Rel. Res.118, 230–241 (1976)
Wallach, S., Reigenstein, D., Bellavia, J.: Cellular transport of calcium in rat liver. J. Gen. Physiol.49, 743–761 (1966)
Wuthier, R.E.: Zonal analysis of inorganic and organic constituents of the epiphysis during endochondral calcification. Calcif. Tiss. Res.4, 20–38 (1969)
Wuthier, R.E.: Zonal analysis of elctrolytes in epiphyseal cartilage and bone of normal and rachitic chickens and pigs. Calcif. Tiss. Res.8, 24–35 (1971)
Wuthier, R.E.: Lipid composition of isolated epiphyseal cartilage cells, membranes and mstrix vesicles. Biochim. Biophys. Acta409, 128–143 (1975)
Wuthier, R.E., Eanes, E.D.: Effect of phospholipids on the transformation of amorphous calcium phosphate to hydroxyapatite in vitro. Calcif. Tiss. Res.19, 197–210 (1975)
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Wuthier, R.E. Electrolytes of isolated epiphyseal chondrocytes, matrix vesicles, and extracellular fluid. Calc. Tis Res. 23, 125–133 (1977). https://doi.org/10.1007/BF02012777
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DOI: https://doi.org/10.1007/BF02012777