Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1973 Jul 1;58(1):152–171. doi: 10.1083/jcb.58.1.152

ISOLATION AND CHARACTERIZATION OF LAMELLAR BODIES AND TUBULAR MYELIN FROM RAT LUNG HOMOGENATES

Joan Gil 1, Oscar K Reiss 1
PMCID: PMC2109018  PMID: 4726305

Abstract

Three surface-active fractions which differ in their morphology have been isolated from rat lung homogenates by ultracentrifugation in a discontinuous sucrose density gradient. In order of increasing density, the fractions consisted, as shown by electron microscopy, primarily of common myelin figures, lamellar bodies, and tubular myelin figures. The lipid of all three fractions contained approximately 94% polar lipids and 2% cholesterol. In the case of the common myelin figures and the lamellar bodies, the polar lipids consisted of 73% phosphatidylcholines, 9% phosphatidylserines and inositols, and 8% phosphatidylethanolamines. In the case of the tubular myelin figures, the respective percentages were 58, 19, and 5. Over 90% of the fatty acids of the lecithins of all three fractions were saturated. Electrophoresis of the proteins of the fractions in sodium dodecyl sulfate or Triton X-100 revealed that the lamellar bodies and the tubular myelin figures differed in the mobilities of their proteins. The common myelin figures, however, contained proteins from both of the other fractions. These data indicate that, whereas the lipids of the extracellular, alveolar surfactant(s) originate in the lamellar bodies, the proteins arise from another source. It is further postulated that the tubular myelin figures represent a liquid crystalline state of the alveolar surface-active lipoproteins.

Full Text

The Full Text of this article is available as a PDF (1.8 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. ABEL L. L., LEVY B. B., BRODIE B. B., KENDALL F. E. A simplified method for the estimation of total cholesterol in serum and demonstration of its specificity. J Biol Chem. 1952 Mar;195(1):357–366. [PubMed] [Google Scholar]
  2. Askin F. B., Kuhn C. The cellular origin of pulmonary surfactant. Lab Invest. 1971 Sep;25(3):260–268. [PubMed] [Google Scholar]
  3. BALIS J. U., CONEN P. E. THE ROLE OF ALVEOLAR INCLUSION BODIES IN THE DEVELOPING LUNG. Lab Invest. 1964 Oct;13:1215–1229. [PubMed] [Google Scholar]
  4. BARRON E. J., HANAHAN D. J. Observations on the silicic acid chromatography of the neutral lipides of rat liver, beef liver, and yeast. J Biol Chem. 1958 Mar;231(1):493–503. [PubMed] [Google Scholar]
  5. BLIGH E. G., DYER W. J. A rapid method of total lipid extraction and purification. Can J Biochem Physiol. 1959 Aug;37(8):911–917. doi: 10.1139/o59-099. [DOI] [PubMed] [Google Scholar]
  6. BROWN E. S. ISOLATION AND ASSAY OF DIPALMITYL LECITHIN IN LUNG EXTRACTS. Am J Physiol. 1964 Aug;207:402–406. doi: 10.1152/ajplegacy.1964.207.2.402. [DOI] [PubMed] [Google Scholar]
  7. BUCKINGHAM S., AVERY M. E. Time of appearance of lung surfactant in the foetal mouse. Nature. 1962 Feb 17;193:688–689. doi: 10.1038/193688a0. [DOI] [PubMed] [Google Scholar]
  8. BUCKINGHAM S., MCNARY W. F., Jr, SOMMERS S. C. PULMONARY ALVEOLAR CELL INCLUSIONS: THEIR DEVELOPMENT IN THE RAT. Science. 1964 Sep 11;145(3637):1192–1193. doi: 10.1126/science.145.3637.1192. [DOI] [PubMed] [Google Scholar]
  9. Balis J. U., Shelley S. A., McCue M. J., Rappaport E. S. Mechanisms of damage to the lung surfactant system. Ultrastructure and quantitation of normal and in vitro inactivated lung surfactant. Exp Mol Pathol. 1971 Apr;14(2):243–262. doi: 10.1016/0014-4800(71)90069-4. [DOI] [PubMed] [Google Scholar]
  10. Buckingham S., Heinemann H. O., Sommers S. C., McNary W. F. Phospholipid synthesis in the large pulmonary alveolar cell. Its relation to lung surfactants. Am J Pathol. 1966 Jun;48(6):1027–1041. [PMC free article] [PubMed] [Google Scholar]
  11. CAMPICHE M. [Lamellar inclusions of the alveolar cells in the lung of the young rat. Relation between ultrastructure and fixation]. J Ultrastruct Res. 1960 Feb;3:302–312. doi: 10.1016/s0022-5320(60)80016-0. [DOI] [PubMed] [Google Scholar]
  12. Chapman D., Fluck D. J. Physical studies of phospholipids. 3. Electron microscope studies of some pure fully saturated 2,3-diacyl-DL-phosphatidyl-ethanolamines and phosphatidyl-cholines. J Cell Biol. 1966 Jul;30(1):1–11. doi: 10.1083/jcb.30.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. FLEISCHER S., ROUSER G. LIPIDS OF SUBCELLULAR PARTICLES. J Am Oil Chem Soc. 1965 Jul;42:588–607. doi: 10.1007/BF02541295. [DOI] [PubMed] [Google Scholar]
  14. Faulkner C. S., 2nd The role of the granular pneumonocyte in surfactant metabolism an autoradiographic study. Arch Pathol. 1969 May;87(5):521–525. [PubMed] [Google Scholar]
  15. Finley T. N., Pratt S. A., Ladman A. J., Brewer L., McKay M. B. Morphological and lipid analysis of the alveolar lining material in dog lung. J Lipid Res. 1968 May;9(3):357–365. [PubMed] [Google Scholar]
  16. Frosolono M. F., Charms B. L., Pawlowski R., Slivka S. Isolation, characterization, and surface chemistry of a surface-active fraction from dog lung. J Lipid Res. 1970 Sep;11(5):439–457. [PubMed] [Google Scholar]
  17. Gacad G., Dickie K., Massaro D. Protein synthesis in lung: influence of starvation on amino acid incorporation into protein. J Appl Physiol. 1972 Sep;33(3):381–384. doi: 10.1152/jappl.1972.33.3.381. [DOI] [PubMed] [Google Scholar]
  18. Gil J. Ultrastructure of lung fixed under physiologically defined conditions. Arch Intern Med. 1971 May;127(5):896–902. [PubMed] [Google Scholar]
  19. Gil J., Weibel E. R. Extracellular lining of bronchioles after perfusion-fixation of rat lungs for electron microscopy. Anat Rec. 1971 Feb;169(2):185–199. doi: 10.1002/ar.1091690205. [DOI] [PubMed] [Google Scholar]
  20. Gil J., Weibel E. R. Improvements in demonstration of lining layer of lung alveoli by electron microscopy. Respir Physiol. 1969 Dec;8(1):13–36. doi: 10.1016/0034-5687(69)90042-5. [DOI] [PubMed] [Google Scholar]
  21. Greenfield L. J., Kimmell G. O. Application of pneumatic techniques to surface tension determinations. The surfactometer. J Surg Res. 1967 Jun;7(6):276–279. doi: 10.1016/0022-4804(67)90066-2. [DOI] [PubMed] [Google Scholar]
  22. Harrison G. A., Weibel J. The membranous component of alveolar exudate. J Ultrastruct Res. 1968 Aug;24(3):334–342. doi: 10.1016/s0022-5320(68)90069-5. [DOI] [PubMed] [Google Scholar]
  23. Hirsch J. G., Fedorko M. E. Ultrastructure of human leukocytes after simultaneous fixation with glutaraldehyde and osmium tetroxide and "postfixation" in uranyl acetate. J Cell Biol. 1968 Sep;38(3):615–627. doi: 10.1083/jcb.38.3.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Hoffman L. Isolation of inclusion bodies from rabbit lung parenchyma. J Cell Physiol. 1972 Feb;79(1):65–72. doi: 10.1002/jcp.1040790107. [DOI] [PubMed] [Google Scholar]
  25. KLAUS M. H., CLEMENTS J. A., HAVEL R. J. Composition of surface-active material isolated from beef lung. Proc Natl Acad Sci U S A. 1961 Nov 15;47:1858–1859. doi: 10.1073/pnas.47.11.1858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. KLAUS M. H., CLEMENTS J. A., HAVEL R. J. Composition of surface-active material isolated from beef lung. Proc Natl Acad Sci U S A. 1961 Nov 15;47:1858–1859. doi: 10.1073/pnas.47.11.1858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. KLAUS M., REISS O. K., TO OLEY W. H., PIEL C., CLEMENTS J. A. Alveolar epithelial cell mitochondria as source of the surface-active lung lining. Science. 1962 Sep 7;137(3532):750–751. doi: 10.1126/science.137.3532.750. [DOI] [PubMed] [Google Scholar]
  28. Kikkawa Y. Morphology of alveolar lining layer. Anat Rec. 1970 Aug;167(4):389–400. doi: 10.1002/ar.1091670403. [DOI] [PubMed] [Google Scholar]
  29. Kikkawa Y., Motoyama E. K., Gluck L. Study of the lungs of fetal and newborn rabbits. Morphologic, biochemical, and surface physical development. Am J Pathol. 1968 Jan;52(1):177–210. [PMC free article] [PubMed] [Google Scholar]
  30. Kistler G. S., Caldwell P. R., Weibel E. R. Development of fine structural damage to alveolar and capillary lining cells in oxygen-poisoned rat lungs. J Cell Biol. 1967 Mar;32(3):605–628. doi: 10.1083/jcb.32.3.605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kuhn C., 3rd A comparison of freeze-substitution with other methods for preservation of the pulmonary alveolar lining layer. Am J Anat. 1972 Apr;133(4):495–507. doi: 10.1002/aja.1001330410. [DOI] [PubMed] [Google Scholar]
  32. Kuhn C., 3rd Cytochemistry of pulmonary alveolar epithelial cells. Am J Pathol. 1968 Nov;53(5):809–833. [PMC free article] [PubMed] [Google Scholar]
  33. LUFT J. H. Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol. 1961 Feb;9:409–414. doi: 10.1083/jcb.9.2.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Leeson T. S., Leeson C. R. Osmiophilic lamellated bodies and associated material in lung alveolar spaces. J Cell Biol. 1966 Mar;28(3):577–581. doi: 10.1083/jcb.28.3.577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. MACKENZIE C. G., MACKENZIE J. B., REISS O. K. REGULATION OF CELL LIPID METABOLISM AND ACCUMULATION. 3. THE LIPID CONTENT OF MAMMALIAN CELLS AND THE RESPONSE TO THE LIPOGENIC ACTIVITY OF RABBIT SERUM. Exp Cell Res. 1964 Dec;36:533–547. doi: 10.1016/0014-4827(64)90310-6. [DOI] [PubMed] [Google Scholar]
  36. Massaro D., Simon M. R., Steinkamp H. Metabolic factors affecting protein synthesis by lung in vitro. J Appl Physiol. 1971 Jan;30(1):1–6. doi: 10.1152/jappl.1971.30.1.1. [DOI] [PubMed] [Google Scholar]
  37. Massaro D., Weiss H., Simon M. R. Protein synthesis and secretion by lung. Am Rev Respir Dis. 1970 Feb;101(2):198–206. doi: 10.1164/arrd.1970.101.2.198. [DOI] [PubMed] [Google Scholar]
  38. Massaro G. D., Massaro D. Granular pneumocytes. Electron microscopic radioautographic evidence of intracellular protein transport. Am Rev Respir Dis. 1972 Jun;105(6):927–931. doi: 10.1164/arrd.1972.105.6.927. [DOI] [PubMed] [Google Scholar]
  39. Meban C. Localization of phosphatidic acid phosphatase activity in granular pneumonocytes. J Cell Biol. 1972 Apr;53(1):249–252. doi: 10.1083/jcb.53.1.249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Noack W. Das elektronenmikroskopische Bild des Lungenepithels von Rattenembryonen vom Tag 16 bis zur Geburt. Acta Anat (Basel) 1971;79(4):445–465. [PubMed] [Google Scholar]
  41. O'Hare K. H., Newman J. K., Vatter A. E., Reiss O. K. Esterases in developing an adult rat lung. II. An electrophoretic analysis. J Histochem Cytochem. 1971 Feb;19(2):116–123. doi: 10.1177/19.2.116. [DOI] [PubMed] [Google Scholar]
  42. O'Hare K. H., Reiss O. K., Vatter A. E. Esterases in developing an adult rat lung. I. Biochemical and electron microscopic observations. J Histochem Cytochem. 1971 Feb;19(2):97–115. doi: 10.1177/19.2.97. [DOI] [PubMed] [Google Scholar]
  43. OYAMA V. I., EAGLE H. Measurement of cell growth in tissue culture with a phenol reagent (folin-ciocalteau). Proc Soc Exp Biol Med. 1956 Feb;91(2):305–307. doi: 10.3181/00379727-91-22245. [DOI] [PubMed] [Google Scholar]
  44. PATTLE R. E. SURFACE LINING OF LUNG ALVEOLI. Physiol Rev. 1965 Jan;45:48–79. doi: 10.1152/physrev.1965.45.1.48. [DOI] [PubMed] [Google Scholar]
  45. Pfleger R. C., Henderson R. F., Waide J. Phosphatidyl glycerol--a major component of pulmonary surfactant. Chem Phys Lipids. 1972 Jul;9(1):51–68. doi: 10.1016/0009-3084(72)90033-3. [DOI] [PubMed] [Google Scholar]
  46. REYNOLDS E. S. The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol. 1963 Apr;17:208–212. doi: 10.1083/jcb.17.1.208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. ROUSER G., KRITCHEVSKY G., GALLI C., HELLER D. DETERMINATION OF POLAR LIPIDS: QUANTITATIVE COLUMN AND THIN-LAYER CHROMATOGRAPHY. J Am Oil Chem Soc. 1965 Mar;42:215–227. doi: 10.1007/BF02541135. [DOI] [PubMed] [Google Scholar]
  48. Reiss O. K. Studies of lung metabolism. I. Isolation and properties of subcellular fractions from rabbit lung. J Cell Biol. 1966 Jul;30(1):45–57. doi: 10.1083/jcb.30.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Rouser G., Siakotos A. N., Fleischer S. Quantitative analysis of phospholipids by thin-layer chromatography and phosphorus analysis of spots. Lipids. 1966 Jan;1(1):85–86. doi: 10.1007/BF02668129. [DOI] [PubMed] [Google Scholar]
  50. STOECKENIUS W. An electron microscope study of myelin figures. J Biophys Biochem Cytol. 1959 May 25;5(3):491–500. doi: 10.1083/jcb.5.3.491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. STOECKENIUS W., MAHR S. C. STUDIES ON THE REACTION OF OSMIUM TETROXIDE WITH LIPIDS AND RELATED COMPOUNDS. Lab Invest. 1965 Jun;14:1196–1207. [PubMed] [Google Scholar]
  52. STOECKENIUS W. Some electron microscopical observations on liquid-crystalline phases in lipid-water systems. J Cell Biol. 1962 Feb;12:221–229. doi: 10.1083/jcb.12.2.221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Singh J., Wasserman A. R. Detection of aggregation and non-destructive disaggregation of membranous proteins using polyacrylamide gel electrophoresis with non-ionic detergents. Biochim Biophys Acta. 1970 Nov 17;221(2):379–382. doi: 10.1016/0005-2795(70)90281-3. [DOI] [PubMed] [Google Scholar]
  54. Sorokin S P. A morphologic and cytochemical study on the great alveolar cell. J Histochem Cytochem. 1966 Dec;14(12):884–897. doi: 10.1177/14.12.884. [DOI] [PubMed] [Google Scholar]
  55. Stossel T. P., Mason R. J., Pollard T. D., Vaughan M. Isolation and properties of phagocytic vesicles. II. Alveolar macrophages. J Clin Invest. 1972 Mar;51(3):604–614. doi: 10.1172/JCI106850. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Sun C. N. Lattice structures and osmiophilic bodies in the developing respiratory tissue of rats. J Ultrastruct Res. 1966 Jun;15(3):380–388. doi: 10.1016/s0022-5320(66)80114-4. [DOI] [PubMed] [Google Scholar]
  57. Trump B. F., Bulger R. E. New ultrastructural characteristics of cells fixed in a glutaraldehyde-osmium tetroxide mixture. Lab Invest. 1966 Jan;15(1 Pt 2):368–379. [PubMed] [Google Scholar]
  58. Untersee P., Gil J., Weibel E. R. Visualization of extracellular lining layer of lung alveoli by freeze-etching. Respir Physiol. 1971 Nov;13(2):171–185. doi: 10.1016/0034-5687(71)90088-0. [DOI] [PubMed] [Google Scholar]
  59. Vatter A. E., Reiss O. K., Newman J. K., Lindquist K., Groeneboer E. Enzymes of the lung. I. Detection of esterase with a new cytochemical method. J Cell Biol. 1968 Jul;38(1):80–98. doi: 10.1083/jcb.38.1.80. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
  61. Weibel E. R., Gil J. Electron microscopic demonstration of an extracellular duplex lining layer of alveoli. Respir Physiol. 1968 Jan;4(1):42–57. doi: 10.1016/0034-5687(68)90006-6. [DOI] [PubMed] [Google Scholar]
  62. Weibel E. R., Kistler G. S., Töndury G. A stereologic electron microscope study of "tubular myelin figures" in alveolar fluids of rat lungs. Z Zellforsch Mikrosk Anat. 1966;69:418–427. doi: 10.1007/BF00406293. [DOI] [PubMed] [Google Scholar]
  63. Williams C. H., Vail W. J., Harris R. A., Green D. E., Valdivia E. The isolation and characterization of the lamellar body of bovine lung. Prep Biochem. 1971 Jan;1(1):37–45. doi: 10.1080/00327487108081928. [DOI] [PubMed] [Google Scholar]
  64. Young S. L., Tierney D. F. Dipalmitoyl lecithin secretion and metabolism by the rat lung. Am J Physiol. 1972 Jun;222(6):1539–1544. doi: 10.1152/ajplegacy.1972.222.6.1539. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES