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Alkaline phosphatase inhibition by parathyroid hormone and isoproterenol in a clonal rat osteosarcoma cell line. Possible mediation by cyclic AMP

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The effect of parathyroid hormone (PTH 1–34 bovine) on alkaline phosphatase activity was investigated in an osteoblast-like clonal cell line derived from rat osteosarcoma (ROS 17/2). ROS 17/2 alkaline phosphatase resembled the bone enzyme in levamisole sensitivity and electrophoretic mobility but differed in heat stability. The specific activity of ROS 17/2 alkaline phosphatase increased with time in culture. This increase was inhibited by PTH (1–34) and (-)-isoproterenol in a dose-dependent manner starting at near-physiological hormone concentrations. The ID50 values were 0.02 nM for PTH (1–34) and 1.7 nM for isoproterenol. The two hormones stimulated ROS 17/2 adenylate cyclase, albeit at higher concentrations: Km values were 13 nM for PTH (1–34) and 16 nM for isoproterenol. The rise in alkaline phosphatase was also inhibited by dibutyryl cyclic AMP and 8-bromocyclic AMP (0.1 mM). These findings further document the osteoblastic properties of the ROS 17/2 osteosarcoma cell line, suggest that PTH inhibition of alkaline phosphatase represents a physiological response to the hormone in these cells, and implicate cyclic AMP as a mediator of this PTH effect.

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References

  1. Luben, R. A., Wong, G. L., Cohn, D. V.: Biochemical characterization with parathormone and calcitonin of isolated bone cells: provisional identification of osteoclasts and osteoblasts, Endocrinology99:526–534, 1976

    Article  CAS  PubMed  Google Scholar 

  2. Peck, W. A., Burks, J. K., Wilkins, J., Rodan, S. B., Rodan, G. A.: Evidence for preferential effects of parathyroid hormone, calcitonin and adenosine on bone and periosteum, Endocrinology100:1357–1366, 1977

    CAS  PubMed  Google Scholar 

  3. Thomas, M. L., Ramp, W. K.: Effects of parathyroid hormone on alkaline phosphatase activity and mineralization of cultured chick embryo tibiae, Calcif. Tissue Int.27:137–142, 1979

    CAS  PubMed  Google Scholar 

  4. Felix, R., Fleisch, H.: Increase in alkaline phosphatase activity in calvaria cells cultured with diphosphonates, Biochem. J.183:73–81, 1979

    CAS  PubMed  Google Scholar 

  5. Dietrich, J. W., Canalis, E. M., Maina, D. M., Raisz, L. G.: Hormonal control of bone collagen synthesis in vitro: effects of parathyroid hormone and calcitonin, Endocrinology98:943–949, 1976

    CAS  PubMed  Google Scholar 

  6. Raisz, L. G., Lorenzo, J., Gworek, S., Kream, B., Rosenblatt, M.: Comparison of the effects of a potent synthetic analog of bovine parathyroid hormone with native bPTH (1–84) and synthetic bPTH-(1–34) on bone resorption and collagen synthesis, Calcif. Tissue Int.29:215–218, 1979

    Article  CAS  PubMed  Google Scholar 

  7. Peck, W. A.: Cyclic AMP as a second messenger in the skeletal actions of parathyroid hormone: a decade-old hypothesis, Calcif. Tissue Int.29:1–4, 1979

    Article  CAS  PubMed  Google Scholar 

  8. Majeska, R. J., Rodan, S. B., Rodan, G. A.: Parathyroid hormone responsive clonal cell lines from rat osteosarcoma, Endocrinology107:1494–1503, 1980

    CAS  PubMed  Google Scholar 

  9. Manolagas, S. C., Haussler, M. R., Deftos, L. J.: 1,25-Dihydroxyvitamin D3 receptor-like macromolecule in rat osteogenic sarcoma cell lines, J. Biol. Chem.255:4414–4417, 1980

    CAS  PubMed  Google Scholar 

  10. Nishimoto, S. K., Price, P. A.: Secretion of the vitamin K-dependent protein of bone by rat osteosarcoma cells. Evidence for an intracellular precursor, J. Biol. Chem.255:6577–6583, 1980

    Google Scholar 

  11. Weidman, E. R., Gill, G. N.: Differential effects of ACTH and 8BrcAMP on growth and replication in a functional adrenal tumor cell line, J. Cell. Physiol.90:91–104, 1977

    Article  CAS  PubMed  Google Scholar 

  12. Rae, P. A., Gutmann, N. S., Tsao, J., Schimmer, B. P.: Mutations in cyclic AMP-dependent protein kinase and corticotropin (ACTH)-sensitive adenylate cyclase affect adrenal steroidogenesis, Proc. Natl. Acad. Sci. U.S.A.76:1896–1900, 1979

    CAS  PubMed  Google Scholar 

  13. deVellis, J., Brooker, G.: Induction of enzymes by glucocorticoids and catecholamines in a rat glial cell line. In G. Sato (ed.): Tissue Culture of the Nervous System, pp. 231–245. Academic Press, New York, 1974

    Google Scholar 

  14. Schwartz, J. P., Costa, E.: Protein kinase translocation followingβ-adrenergic receptor activation in C6 glioma cells, J. Biol. Chem.255:2943–2948, 1980

    CAS  PubMed  Google Scholar 

  15. McKeehan, W. L., Hamilton, W. G., Ham, R. G.: Selenium is an essential trace nutrient for growth of WI-38 diploid human fibroblasts, Proc. Natl. Acad. Sci. U.S.A.73:2023–2027, 1976

    CAS  PubMed  Google Scholar 

  16. Lowry, O. H.: Micromethods for the assay of enzyme. II. Specific procedures. Alkaline phosphatase, Meth. Enzymol.4:371–372, 1955

    Google Scholar 

  17. Spector, T.: Refinement of the Coomassie Blue method of protein quantitation, Anal. Biochem.86:142–146, 1978

    Article  CAS  PubMed  Google Scholar 

  18. Humes, J. L., Bounbehler, M., Kuehl, F. A., Jr.: A new assay for measuring adenyl cyclase activity in intact cells, Anal. Biochem.32:210–217, 1969

    Article  CAS  PubMed  Google Scholar 

  19. Salomon, Y., London, C. D., Rodbell, M.: An improved method for measuring adenyl cyclase, Anal. Biochem.58:541–549, 1974

    Article  CAS  PubMed  Google Scholar 

  20. Laemmli, U. K.: Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature227:680–685, 1970

    Article  CAS  PubMed  Google Scholar 

  21. Farley, J. R., Ivey, J. L., Baylink, D. J.: Human skeletal alkaline phosphatase. Kinetic studies including pH dependence and inhibition by theophylline, J. Biol. Chem.255:4680–4686, 1980

    CAS  PubMed  Google Scholar 

  22. Goldstein, D. J., Rogers, C. E., Harris, H.: Expression of alkaline phosphatase loci in mammalian tissues, Proc. Natl. Acad. Sci. U.S.A.77:2857–2860, 1980

    CAS  PubMed  Google Scholar 

  23. van Belle, H.: Kinetics and inhibition of alkaline phosphatases from canine tissues, Biochim. Biophys. Acta289:158–168, 1972

    PubMed  Google Scholar 

  24. Singh, I., Tsang, K. Y.: An in vitro production of bone specific alkaline phosphatase, Exp. Cell Res.95:347–358, 1975

    Article  CAS  PubMed  Google Scholar 

  25. Hamada, H., Amitani, K., Ono, K., Sakamoto, Y., Koshimoto, R., Nakata, Y.: Osseous alkaline phosphatase from osteosarcoma in mouse and in culture, Cell. Mol. Biol.25:77–84, 1979

    CAS  Google Scholar 

  26. 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

    CAS  PubMed  Google Scholar 

  27. Goldstein, D. J., Harris, H.: Human placental alkaline phosphatase differs from that of other species, Nature280:602–605, 1979

    Article  CAS  PubMed  Google Scholar 

  28. McKenna, M. J., Hamilton, T. A., Sussman, H. H.: Comparison of human alkaline phosphatase isoenzymes. Structural evidence for three protein classes, Biochem. J.181:67–73, 1979

    CAS  PubMed  Google Scholar 

  29. Wada, H. G., VandenBerg, S. R., Sussman, H. H., Grove, W. E., Herman, M. M.: Characterization of two different alkaline phosphatases of mouse teratoma: partial purification, electrophoretic, and histochemical studies, Cell9:37–44, 1976

    Article  CAS  PubMed  Google Scholar 

  30. Waters, M. D., Summer, G. K., Switzer, B. R., Moore, R. D., Heitkamp, D. H.: Alkaline phosphatase activation and collagen synthesis in human skin fibroblasts in culture, Exp. Cell Res.80:170–174, 1973

    Article  CAS  PubMed  Google Scholar 

  31. Cox, R. P., Elson, N. A., Tu, S.-H., Griffin, M. J.: Hormonal induction of alkaline phosphatase activity by an increase in catalytic efficiency of the enzyme, J. Mol. Biol.58:197–215, 1971

    Article  CAS  PubMed  Google Scholar 

  32. Singh, I., Tsang, K. Y., Blakemore, W. S.: Placenta-like alkaline phosphatase from human osteosarcoma cells, Cancer Res.38:193–198, 1978

    CAS  PubMed  Google Scholar 

  33. Haussler, M. R., Nagode, L. A., Rasmussen, H.: Induction of intestinal brush border alkaline phosphatase by vitamin D and identity with Ca-ATPase, Nature228:1199–1201, 1970

    Article  CAS  PubMed  Google Scholar 

  34. Wong, G. L., Luben, R. A., Cohn, D. V.: 1,25-Dihydroxycholecalciferol and parathormone: effects on isolated osteoclast-like and osteoblast-like cells, Science197:663–665, 1977

    CAS  PubMed  Google Scholar 

  35. Chen, T. L., Feldman, D.: Glucocorticoid receptors and actions in subpopulations of cultured rat bone cells. Mechanism of dexamethasone potentiation of parathyroid hormone-stimulated cyclic AMP production, J. Clin. Invest.63:750–758, 1979

    Article  CAS  PubMed  Google Scholar 

  36. Chen, T. L., Feldman, D.: Glucorcorticoid potentiation of the adenosine 3',5'-monophosphate response to parathyroid hormone in cultured rat bone cells, Endocrinology102:589–596, 1978

    CAS  PubMed  Google Scholar 

  37. Parsons, J. A., Rafferty, B., Gray, D., Reit, B., Zanelli, J. M., Keutmann, H. T., Tregear, G. W., Callahan, E. N., Potts, J. T., Jr.: Pharmacology of parathyroid hormone and some of its fragments and analogues. In R. V. Talmage, M. Owen, J. A. Parsons, (eds.): Calcium-Regulating Hormones, pp. 33–39. Elsevier, Amsterdam, 1975

    Google Scholar 

  38. Dufau, M. L., Tsuruhara, T., Horner, K. A., Podesta, E., Catt, K. H.: Intermediate role of adenosine 3′,5′-cyclic monophosphate and protein kinase during gonadotropin-induced steroidogenesis in testicular intestitial cells, Proc. Natl. Acad. Sci. U.S.A.74:3419–3423, 1977

    CAS  PubMed  Google Scholar 

  39. Saez, J. M., Evain, D., Gallet, D.: Role of cyclic-AMP and protein kinase on the steroidogenic action of ACTH, prostaglandin E1 and dibutyryl cyclic AMP in normal adrenal cells and adrenal tumor cells from humans, J. Cyclic Nucleotide Res.4:311–321, 1978

    CAS  PubMed  Google Scholar 

  40. Schimmer, B. P.: Adenylate cyclase activity and steroidogenesis in phenotypic revertants of an ACTH-insensitive adrenal tumour cell line, Nature259:482–483, 1976

    Article  CAS  PubMed  Google Scholar 

  41. Wang, T., Sheppard, J. R., Foker, J. E.: Rise and fall of cyclic AMP required for onset of lymphocyte DNA synthesis, Science201:155–157, 1978

    CAS  PubMed  Google Scholar 

  42. Green, H., Meuth, M.: An established pre-adipose cell line and its differentiation in culture, Cell3:127–133, 1974

    Article  CAS  PubMed  Google Scholar 

  43. Kream, B. E., Rowe, D. W., Gworek, S. C., Raisz, L. G.: Parathyroid hormone alters collagen synthesis and procollagen mRNA levels in fetal rat calvaria, Proc. Natl. Acad. Sci. U.S.A.77:5654–5658, 1980

    CAS  PubMed  Google Scholar 

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  1. Department of Oral Biology, University of Connecticut, School of Dental Medicine, 06032, Farmington, Connecticut, USA

    Robert J. Majeska & Gideon A. Rodan

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  1. Robert J. Majeska
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Majeska, R.J., Rodan, G.A. Alkaline phosphatase inhibition by parathyroid hormone and isoproterenol in a clonal rat osteosarcoma cell line. Possible mediation by cyclic AMP. Calcif Tissue Int 34, 59–66 (1982). https://doi.org/10.1007/BF02411210

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