Sphingomyelin Storage in Lymphoid Cell Lines from Patients with Niemann-Pick Disease Types A, B and C: Influence of Culture Conditions

  • Thierry Levade
  • Robert Salvayre
  • Arlette Maret
  • Louis Douste-Blazy
Conference paper
Part of the NATO ASI Series book series (NSSA, volume 150)

Abstract

Epstein-Barr-virus-transformed lymphoid cell lines from normal individuals and from patients with Niemann-Pick disease types A, B or C were subjected to various culture conditions in order to study the source of the characteristic lysosomal storage of sphingomyelin (SPM) observed in the tissues of Niemann-Pick patients. RPMI 1640 containing L-glutamine and antibiotics was used as culture medium and was supplemented with 2% Ultroser HY (a serum substitute devoid of lipoproteins and lipids) or with one of the following lipid sources: fetal calf serum (10%), human LDL (100 to 400 µg protein/ml) or human HDL (400 µg protein/ml). By determining the phosphorus content in the SPM fraction of cell lipid extracts, storage of SPM was demonstrated under all tested culture conditions in cells deficient in acid sphingomyelinase, i.e., lymphoid cell lines from Niemann-Pick disease types A and B. In contrast, the lymphoid cell line from a Niemann-Pick type C patient (not deficient in sphingomyelinase), like normal cells, exhibited no storage of SPM. Even after growing the cells for more than 30 days in a medium devoid of SPM, the Niemann-Pick types A and B lymphoid cell lines showed considerable accumulation of SPM (about 2.0–2.3 times more SPM than in the control lymphoid cell lines). SPM concentration was higher when cells were grown in a medium supplemented with lipids, particularly human LDL or HDL. These results are consistent with the hypothesis that there are both exogenous and endogenous factors in the lysosomal storage of SPM observed in lymphoid cell lines from patients with Niemann-Pick disease types A and B.

Keywords

Cholesterol Hydrolysis Phosphorus Albumin Foam 

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References

  1. 1.
    R.O. Brady, Sphingomyelin lipidoses: Niemann-Pick disease, in: “The Metabolic Basis of Inherited Disease”, J.B. Stanbury, J.B. Wyngaarden, D.S. Fredrickson, J.L. Goldstein and M.S. Brown, eds., McGraw Hill, New York, pp. 831–841 (1983).Google Scholar
  2. 2.
    M. Elleder and A. Jirasek, Niemann-Pick disease, Acta Universitatis Carolinae Medica, 29: 259 (1983).PubMedGoogle Scholar
  3. 3.
    T. Levade, R. Salvayre and L. Douste-Blazy, Sphingomyelinases and Niemann-Pick disease, J. Clin. Chem. Clin. Biochem.. 24: 205 (1986).PubMedGoogle Scholar
  4. 4.
    D.A. Wenger, Niemann-Pick disease, in: “Practical Enzymology of the Sphingolipidoses”, R. H. Glew and S. P. Peters, eds., Alan R. Liss, New York, pp. 39–70 (1977).Google Scholar
  5. 5.
    T. Levade, R. Salvayre, G. Lenoir and L. Douste-Blazy, Sphingomyelinase and non-specific phosphodiesterase activities in Epstein-Barr virustransformed lymphoid cell lines from Niemann-Pick disease A, B and C, Biochim. Biophys. Acta, 793: 321 (1984).PubMedGoogle Scholar
  6. 6.
    T. Levade, R. Salvayre and L. Douste-Blazy, Molecular forms of sphingomyelinase and non-specific phosphodiesterases in Epstein-Barr virus-transformed lymphoid cell lines from Niemann-Pick disease A and B, Eur. J. Biochem., 149: 405 (1985).PubMedCrossRefGoogle Scholar
  7. 7.
    T. Levade, R. Salvayre, J.C. Bes, A. Maret and L. Douste-Blazy, Biochemical and ultrastructural findings in a lymphoid cell line from Niemann-Pick disease type A, Biol. Cell, 55: 143 (1985).PubMedGoogle Scholar
  8. 8.
    T. Levade, R. Salvayre, J.C. Bes, M. Nezri and L. Douste-Blazy, New tools for the study of Niemann-Pick disease: analogues of natural substrate and Epstein-Barr virus-transformed lymphoid cell lines, Pediatr. Res., 19: 153 (1985).PubMedCrossRefGoogle Scholar
  9. 9.
    T. Levade, A. Maret, R. Salvayre, N. Livni, P. Rogalle and L. Douste-Blazy, Biochemical and ultrastructural studies on an Epstein-Barr virustransformed lymphoid cell line from a Niemann-Pick disease type C patient, Biochim. Biophvs. Acta, 877: 415 (1986).Google Scholar
  10. 10.
    V.N. Schumaker and D.L. Puppione, Sequential flotation ultracentrifugation, Meth. Enzvmol., 128: 155 (1986).CrossRefGoogle Scholar
  11. 11.
    J.L. Goldstein and M.S. Brown, Binding and degradation of low density lipoproteins by cultured human fibroblasts, J. Biol. Chem.. 249: 5153 (1974).PubMedGoogle Scholar
  12. 12.
    E.F. Hartree, Determination of protein: a modification of the Lowry method that gives a linear photometric response, Anal. Biochem., 48: 422 (1972).PubMedCrossRefGoogle Scholar
  13. 13.
    J. Folch, M. Lees and G.H. Sloane-Stanley, A simple method for the isolation and purification of total lipids from animal tissues, J. Biol. Chem.. 226: 497 (1957).PubMedGoogle Scholar
  14. 14.
    B.N. Ames, Assay of inorganic phosphate, total phosphate and phosphatases, Meth. Enzvmol., 8: 115 (1966).CrossRefGoogle Scholar
  15. 15.
    T.M. Forte, J.J. Bell-Quint and F. Cheng, Lipoproteins of fetal and newborn calves and adult steer: a study of developmental changes, Lipids. 16: 240 (1981).PubMedCrossRefGoogle Scholar
  16. 16.
    P.H. Fishman, R.M. Bradley, M.S. Brown, J.R. Faust and J.L. Goldstein, Similar content of phospholipids and gangliosides in normal and homozygous familial hypercholesterolemia fibroblasts, J. Lipid Res.. 19: 304 (1978).PubMedGoogle Scholar
  17. 17.
    M.W. Spence, J.T.R. Clarke and H.W. Cook, Pathways of sphingomyelin metabolism in cultured fibroblasts from normal and sphingomyelin lipidosis subjects, J. Biol. Chem., 258: 8595 (1983).PubMedGoogle Scholar
  18. 18.
    H.J. Kayden, L. Hatam and N.G. Beratis, Regulation of 3-hydroxy-3-methylglutaryl Coenzyme A reductase activity and the esterification of cholesterol in human long term lymphoid cell lines, Biochemistry 15: 521 (1976).PubMedCrossRefGoogle Scholar
  19. 19.
    Y.K. Ho, M.S. Brown, H.J. Kayden and J.L. Goldstein, Binding, internalization, and hydrolysis of low density lipoprotein in long-term lymphoid cell lines from a normal subject and a patient with homozygous familial hypercholesterolemia, J. Exp. Med.. 144: 444 (1976).PubMedCrossRefGoogle Scholar
  20. 20.
    A.I. Leikin, M. Mihovilovic and A.M. Scanu, High density lipoproteins influence cholesterol homeostasis in cultured virus-transformed human lymphoblastoid cells, J. Biol. Chem.. 257: 14280 (1982).PubMedGoogle Scholar
  21. 21.
    R.L. Jackson, J.D. Morrisett and A.M. Gotto, Lipoprotein structure and metabolism, Physiol. Rev.. 56: 259 (1976).PubMedGoogle Scholar
  22. 22.
    A.L. Beaudet and A.A. Manschreck, Metabolism of sphingomyelin by intact cultured fibroblasts: differentiation of Niemann-Pick disease types A and B, Biochem. Biophvs. Res. Commun.. 105: 14 (1982).CrossRefGoogle Scholar
  23. 23.
    T. Kudoh, M.A. Velkoff and D.A. Wenger, Uptake and metabolism of radioactively labeled sphingomyelin in cultured skin fibroblasts from controls and patients with Niemann-Pick disease and other lysosomal storage diseases, Biochim. Biophvs. Acta. 754: 82 (1983).Google Scholar
  24. 24.
    M.T. Vanier, R. Rousson, I. Garcia, G. Bailloud, M. C. Juge, A. Revol and P. Louisot, Biochemical studies in Niemann-Pick disease. III, Clin. Genet.. 27: 20 (1985).PubMedCrossRefGoogle Scholar
  25. 25.
    S.L. Sutrina and W.W. Chen, Lysosomal involvement in cellular turnover of plasma membrane sphingomyelin, Biochim. Biophys. Acta, 793: 169 (1984).PubMedGoogle Scholar

Copyright information

© Plenum Press, New York 1988

Authors and Affiliations

  • Thierry Levade
    • 1
    • 2
  • Robert Salvayre
    • 1
    • 2
  • Arlette Maret
    • 1
    • 2
  • Louis Douste-Blazy
    • 1
    • 2
  1. 1.INSERM U.101France
  2. 2.Laboratoire de Biochimie, Faculté de Médecine PurpanToulouse CedexFrance

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