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Ultrastructure of cultured hepatocytes from fat- and cholesterol-fed rats

  • R. L. Moses
  • Joseph B. Delcarpio
  • Monica Malone McNeal
  • Roger A. Davis
Article

Summary

Hepatocytes were enzymatically dissociated from the livers of rats maintained for eight weeks on diets of (1) standard rat chow, (2) chow plus 20% olive oil, and (3) chow plus 20% olive oil and 2% cholesterol. Cells were cultured for up to one week and characterized by phase contrast light microscopy and transmission electron microscopy. Cultured hepatocytes derived from the cholesterol-fed rats (3) were distinguishable by phase contrast light microscopy from those derived from chow-or chow plus olive oil-fed (1 and 2) cells, since they contained numerous refractile cytoplasmic structures. Transmission electron microscopy demonstrated that these structures were lipid droplets. Hepatocytes were observed to accumulate non-membrane bound cytoplasmic lipid droplets in the order control-fed <olive oil-fed <cholesterol-fed. In addition to varying quantitatively among the feeding groups, lipid droplets also varied in size and osmiophilia as a function of the feeding regimen. Other than the differential accumulation of lipid droplets, the ultrastructure of the cultured hepatocytes was similar among cells derived from the various feeding groups.

Key words

Hepatocyte Tissue culture Electron microscopy Fatty liver Lipid 

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References

  1. Baglio CG, Farber E (1965) Reversal by adenine of the ethionine induced lipid accumulation in the endoplasmic reticulum of the rat liver. J Cell Biol 27:591–601PubMedCentralCrossRefPubMedGoogle Scholar
  2. Bernaert D, Wanson JC, Drochmans P, Popowski A (1977) Effect of insulin on ultrastructure and glycogenesis in primary cultures of adult rat hepatocytes. J Cell Biol 74:878–900PubMedCentralCrossRefPubMedGoogle Scholar
  3. Berry MN, Friend DS (1970) High-yield preparation of isolated rat liver parenchymal cells. J Cell Biol 43:506–520CrossRefGoogle Scholar
  4. Blouin A, Bolender RP, Weibel ER (1977) Distribution of organelles and membranes between hepatocytes and nonhepatocytes in the rat liver parenchyma. J Cell Biol 72:441–455CrossRefPubMedGoogle Scholar
  5. Capuzzi DM, Margolis S (1971) Metabolic studies in isolated rat liver cells. II. Biosynthesis of serum low density lipoproteins and its regulation. Lipids 6:609–616PubMedGoogle Scholar
  6. Chapman GS, Jones AL, Meyer UA, Bissell DM (1973) Parenchymal cells from adult rat liver in nonproliferating monolayer culture. II. Ultrastructural studies. J Cell Biol 59:735–747PubMedCentralCrossRefPubMedGoogle Scholar
  7. Claude A (1970) Growth and differentiation of cytoplasmic membranes in the course of lipoprotein granule synthesis. J Cell Biol 47:745–766PubMedCentralCrossRefPubMedGoogle Scholar
  8. Coleman R, Bell RM (1978) Evidence that biosynthesis of phosphatidylethanolamine, phosphatidylcholine, and triacylglycerol occurs on the cytoplasmic side of microsomal vesicles. J Cell Biol 76:245–253PubMedCentralCrossRefPubMedGoogle Scholar
  9. Davis RA, Showalter R, Kern F (1978) Reversal by triton WR-1339 of ethynyloestradiol-induced hepatic cholesterol esterification. Biochem J 174:45–51PubMedCentralCrossRefPubMedGoogle Scholar
  10. Davis RA, Engelhorn SC, Pangburn SH, Weinstein DB (1979a) Very low density lipoprotein synthesis and secretion by cultured rat hepatocytes. J Biol Chem 254:2010–2016PubMedGoogle Scholar
  11. Davis RA, Englehorn SC, Weinstein DB, Steinberg D (1979b) Very low density lipoprotein secretion by cultured rat hepatocytes: Inhibition by albumin and other macromolecules. J Biol Chem 255:2039–2045Google Scholar
  12. Davis RA, McNeal M, Moses RL (1982) Intrahepatic assembly of very low density lipoprotein: Competition by cholesterol esters for the hydrophobic core. J Biol Chem 257:2634–2640PubMedGoogle Scholar
  13. DeBrabander M, Wanson JC, Mosselmans R, Geuens G, Drochmans P (1978) Effects of anti-microtubular compounds on monolayer cultures of adult rat hepatocytes. Biol Cellul 31:127–140Google Scholar
  14. Deckelbaum RJ, Shipley GG, Small DM, Lee RS, George PK (1975) Thermal transitions in human plasma low density lipoproteins. Science 190:392–394CrossRefPubMedGoogle Scholar
  15. Drevon CA, Weinstein DB, Steinberg D (1980) Regulation of cholesterol esterification and biosynthesis in monolayer cultures of normal rat hepatocytes. J Biol Chem 255:9128–9137PubMedGoogle Scholar
  16. Drochmans P, Wanson JC, Mosselmans R (1975) Isolation and subfractionatin on ficol gradients of adult rat hepatocytes. J Cell Biol 66:1–22CrossRefPubMedGoogle Scholar
  17. Elshourbagy NA, Barwick JL, Guzelian PS (1981) Induction of cytochrome P-450 by pregnenolone-16-carbonitrile in primary monolayer cultures of adult rat hepatocytes and in cell-free translation system. J Biol Chem 256:6060–6068PubMedGoogle Scholar
  18. French SW, Davies PL (1975) Ultrastructural localization of actin like filaments in rat hepatocytes. Gastroent 68:765–774Google Scholar
  19. Goldstein AL, Johnson PR (1980) Primary cultures of fetal hepatocytes from the genetically obese Zucker rat: Characterization and total lipogenesis, in Vitro 16:288–296CrossRefGoogle Scholar
  20. Haagsman HP, Van Golde MG (1981) Synthesis and secretion of very low density lipoproteins by isolated rat hepatocytes in suspension: Role of diacylglycerol acyltransferase. Arch Biochem Biophys 208:395–402CrossRefPubMedGoogle Scholar
  21. Hamilton RL, Regen DN, Gray ME, LeQuire VS (1967) Lipid transport in liver. I. Electron microscopic identification of very low density lipoproteins in perfused rat liver. Lab Invest 16:305–319PubMedGoogle Scholar
  22. Iseri OA, Lieber CS, Gottleib LS (1966) The ultrastructure of fatty liver induced by prolonged ethanol ingestion. Am J Pathol 48:535–55PubMedCentralPubMedGoogle Scholar
  23. Iype PT (1971) Cultures from adult rat liver cells. I. Establishment of monolayer cultures from normal liver. J Cell Phys 78:281–288CrossRefGoogle Scholar
  24. Jones AL, Fawcett DW (1966) Hypertrophy of the agranular endoplasmic reticulum in hamster liver induced by phenobarbital. J Histochem Cytochem 14:215–232CrossRefPubMedGoogle Scholar
  25. Jones AL, Ruderman NB, Herrera MG (1967) Electron microscopic and biochemical study of lipoprotein synthesis in isolated perfused rat liver. J Lip Res 8:429–446Google Scholar
  26. Kroon PA, (1981) The order-disorder transition of the core cholesteryl esters of human plasma low denisty lipoprotein. J Biol Chem 256:5332–5238PubMedGoogle Scholar
  27. Lee SS, Ho KJ (1975) Cholesterol fatty liver. Arch Pathol 99:301–306PubMedGoogle Scholar
  28. Leffert HL, Paul D (1973) Serum dependent growth of primary cultured differentiated fetal hepatocytes in arginine deficient medium. J Cell Phys 81:113–124CrossRefGoogle Scholar
  29. Lieber CS, DeCarli LM (1970) Quantitative relationships between the amount of dietary fat and severity of alcoholic fatty liver. Am J Clin Nutr 23:474–478PubMedGoogle Scholar
  30. Loud AV (1968) A quantitative sterological description of the ultrastructure of normal rat liver parenchymal cells. J Cell Biol 37:27–46PubMedCentralCrossRefPubMedGoogle Scholar
  31. Mahley RW, Gray ME, Hamilton RL, Lequire VS (1968) Lipid transport in liver. II. Electron microscopic and biochemical studies of alterations in lipoprotein transport induced by cortisone in the rabbit. Lab Invest 19:358–369PubMedGoogle Scholar
  32. Mahley RW, Hamilton RL, Lequire VS (1969) Characterization of lipoprotein particles isolated from the Golgi apparatus of rat liver. J Lip Res 10:433–439Google Scholar
  33. Montesano R, Mira-Moser F, Stefan Y, Perrelet A, Orci L (1978) Tight junctions in fetal rat liver expiants grown in vitro. J Ultrastruct Res 64:182–190CrossRefPubMedGoogle Scholar
  34. Morre DJ, Merritt WD, Lembi CA (1971) Connections between mitochondria and endoplasmic reticulum in rat liver and onion stem. Protoplasma 73:43–49CrossRefPubMedGoogle Scholar
  35. Moses RL, Davis RA (1980) Cell morphology and very low density lipoprotein secretion by cultured hepatocytes derived from fat-fed rats. J Cell Biol 87:305aGoogle Scholar
  36. Moses RL, Davis RA (1981) Morphology of cultured hepatocytes from fat-fed rats. Anat Rec 199:177ACrossRefGoogle Scholar
  37. Novikoff AB, Roheim PS, Quintana N (1966) Changes in rat liver cells induced by orotic acid feeding. Lab Invest 15:27–49Google Scholar
  38. Ontko JA (1973) Effects of ethanol on the metabolism of free fatty acids in isolated liver cells. J Lip Res 14:78–86Google Scholar
  39. Parks HF (1967) An experimental study of microscopic and submicroscopic lipid inclusions in hepatic cells of the mouse. Am J Anat 120:253–280CrossRefGoogle Scholar
  40. Ridout JH, Lucas CC, Patterson JN, Best CH (1954) Changes in chemical composition during the development of “cholesterol fatty livers”. Biochem J 58:297–301PubMedCentralCrossRefPubMedGoogle Scholar
  41. Schanne FAX, Zuker AH, Farber JL (1981) Alcohol-dependent liver cell necrosis in vitro: A new model. Science (US) 212:338–340Google Scholar
  42. Stein O, Stein Y (1965) Fine structure of the ethanol induced fatty liver in the rat. Isr J Med Sci 1:378–388Google Scholar
  43. Stein O, Stein Y (1966) Electron microscopic autoradiography of3H glycerol labelled lipid in ethanol induced fatty liver. Exp Cell Res 42:198–201CrossRefPubMedGoogle Scholar
  44. Stein O, Stein Y (1967) Lipid synthesis, intracellular transport, storage, and secretion. I. Electron microscopic radioautographic study of liver after injection of tritrated palmitate or glycerol in fasted and ethanol treated rats. J Cell Biol 33:319–339PubMedCentralCrossRefPubMedGoogle Scholar
  45. Tarlow DM, Watkins PA, Reed RE, Miller RS, Zwergel EE, Lane MD (1977) Lipogenesis and the synthesis and secretion of very low density lipoprotein by avian liver cells in nonproliferating monolayer culture. J Cell Biol 73:332–353CrossRefPubMedGoogle Scholar
  46. Wanson JC, Drochmans P, Mosselmans R, Ronveaux MF (1977) Adult heaptocytes in primary monolayer culture. J Cell Biol 74:858–877CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 1982

Authors and Affiliations

  • R. L. Moses
    • 1
  • Joseph B. Delcarpio
    • 1
  • Monica Malone McNeal
    • 2
  • Roger A. Davis
    • 2
  1. 1.Department of AnatomyDepartment of Physiology Louisiana State University Medical CenterNew OrleansUSA
  2. 2.Cell Biology UnitDepartment of Physiology Louisiana State University Medical CenterNew OrleansUSA

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