Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1979 Oct 1;83(1):47–64. doi: 10.1083/jcb.83.1.47

An electron microscope autoradiographic study of the carbohydrate recognition systems in rat liver. I. Distribution of 125I-ligands among the liver cell types

PMCID: PMC2110443  PMID: 511941

Abstract

Electron microscope autoradiography was used to study the cellular localization of seven glycoproteins rapidly cleared from the circulating plasma of rats and taken up by the liver. 1 and 15 min after intravenous administration of the 125I-glycoproteins, livers were fixed in situ by perfusion and processed for autoradiography. Autoradiographic grains in the developed sections were found to represent the intact 125I-ligand. A quantitative analysis of the distribution and concentration (density) of autoradiographic grains over the three major cell types of the liver was then performed. Three molecules, asialo-fetuin, asialo-orosomucoid, and lactosaminated RNase A dimer, the oligosaccharide chains of which terminate in galactose residues, were bound and internalized almost exclusively (greater than 90%) by hepatocytes. Conversely, four molecules, the oligosaccharide chains of which terminate in either N-acetyl-glucosamine (agalacto- orosomucoid) or mannose (ahexosamino-orosomucoid, preputial beta- glucuronidase, and mannobiosaminated RNase A dimer), were specifically bound and internalized by cells lining the blood sinusoids--that is, by Kupffer cells and endothelial cells. Endothelial cells were two to six times more active (on a cell volume basis) than were Kupffer cells in the internalization of these four 125I-ligands. Mannose and N- acetylglucosamine-terminated glycoproteins competed with each other for uptake into either endothelial cells or Kupffer cells, indicating that a single system recognized mannose or N-acetyl-glucosamine residues. Finally, agalacto-orosomucoid and ahexosamino-orosomucoid were also associated with hepatocytes, but competition experiments utilizing excess asialo-orosomucoid demonstrated that residual galactosyl residues were responsible for this association.

Full Text

The Full Text of this article is available as a PDF (1.7 MB).

Selected References

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

  1. Achord D. T., Brot F. E., Bell C. E., Sly W. S. Human beta-glucuronidase: in vivo clearance and in vitro uptake by a glycoprotein recognition system on reticuloendothelial cells. Cell. 1978 Sep;15(1):269–278. doi: 10.1016/0092-8674(78)90102-2. [DOI] [PubMed] [Google Scholar]
  2. Achord D., Brot F., Gonzalez-Noriega A., Sly W., Stahl P. Human beta-glucuronidase. II. Fate of infused human placental beta-glucuronidase in the rat. Pediatr Res. 1977 Jul;11(7):816–822. doi: 10.1203/00006450-197707000-00008. [DOI] [PubMed] [Google Scholar]
  3. Ashwell G., Morell A. G. The role of surface carbohydrates in the hepatic recognition and transport of circulating glycoproteins. Adv Enzymol Relat Areas Mol Biol. 1974;41(0):99–128. doi: 10.1002/9780470122860.ch3. [DOI] [PubMed] [Google Scholar]
  4. Baynes J. W., Wold F. Effect of glycosylation on the in vivo circulating half-life of ribonuclease. J Biol Chem. 1976 Oct 10;251(19):6016–6024. [PubMed] [Google Scholar]
  5. Bergeron J. J., Sikstrom R., Hand A. R., Posner B. I. Binding and uptake of 125I-insulin into rat liver hepatocytes and endothelium. An in vivo radioautographic study. J Cell Biol. 1979 Feb;80(2):427–443. doi: 10.1083/jcb.80.2.427. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brown T. L., Henderson L. A., Thorpe S. R., Baynes J. W. The effect of alpha-mannose-terminal oligosaccharides on the survival of glycoproteins in the circulation. Rapid uptake and catabolism of bovine pancreatic ribonuclease B by nonparenchymal cells of rat liver. Arch Biochem Biophys. 1978 Jun;188(2):418–428. doi: 10.1016/s0003-9861(78)80026-5. [DOI] [PubMed] [Google Scholar]
  7. Buys C. H., Dejong A. S., Bouma J. M., Gruber M. Rapid uptake by liver sinusoidal cells of serum albumin modified with retention of its compact conformation. Biochim Biophys Acta. 1975 May 5;392(1):95–100. doi: 10.1016/0304-4165(75)90169-5. [DOI] [PubMed] [Google Scholar]
  8. Buys C. H., Elferink M. G., Bouma J. M., Gruber M., Nieuwenhuis P. Proteolysis of formaldehyde-treated albumin in Kupffer cells and its inhibition by suramin. J Reticuloendothel Soc. 1973 Aug;14(2):209–223. [PubMed] [Google Scholar]
  9. Fahimi H. D. The fine structural localization of endogenous and exogenous peroxidase activity in Kupffer cells of rat liver. J Cell Biol. 1970 Oct;47(1):247–262. doi: 10.1083/jcb.47.1.247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Farquhar M. G., Palade G. E. Cell junctions in amphibian skin. J Cell Biol. 1965 Jul;26(1):263–291. doi: 10.1083/jcb.26.1.263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fertuck H. C., Salpeter M. M. Sensitivity in electron microscope autoradiography for 125I. J Histochem Cytochem. 1974 Feb;22(2):80–87. doi: 10.1177/22.2.80. [DOI] [PubMed] [Google Scholar]
  12. Furbish F. S., Steer C. J., Barranger J. A., Jones E. A., Brady R. O. The uptake of native and desialylated glucocerebrosidase by rat hepatocytes and Kupffer cells. Biochem Biophys Res Commun. 1978 Apr 14;81(3):1047–1053. doi: 10.1016/0006-291x(78)91456-0. [DOI] [PubMed] [Google Scholar]
  13. Greengard O., Federman M., Knox W. E. Cytomorphometry of developing rat liver and its application to enzymic differentiation. J Cell Biol. 1972 Feb;52(2):261–272. doi: 10.1083/jcb.52.2.261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gregoriadis G., Morell A. G., Sternlieb I., Scheinberg I. H. Catabolism of desialylated ceruloplasmin in the liver. J Biol Chem. 1970 Nov 10;245(21):5833–5837. [PubMed] [Google Scholar]
  15. Himeno M., Ohara H., Arakawa Y. Beta-glucuronidase of rat preputial gland. Crystallization, properties, carbohydrate composition, and subunits. J Biochem. 1975 Feb;77(2):427–438. doi: 10.1093/oxfordjournals.jbchem.a130742. [DOI] [PubMed] [Google Scholar]
  16. Hubbard A. L., Cohn Z. A. Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells. J Cell Biol. 1975 Feb;64(2):438–460. doi: 10.1083/jcb.64.2.438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hubbard A. L., Cohn Z. A. The enzymatic iodination of the red cell membrane. J Cell Biol. 1972 Nov;55(2):390–405. doi: 10.1083/jcb.55.2.390. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hubbard A. L., Stukenbrok H. An electron microscope autoradiographic study of the carbohydrate recognition systems in rat liver. II. Intracellular fates of the 125I-ligands. J Cell Biol. 1979 Oct;83(1):65–81. doi: 10.1083/jcb.83.1.65. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hudgin R. L., Pricer W. E., Jr, Ashwell G., Stockert R. J., Morell A. G. The isolation and properties of a rabbit liver binding protein specific for asialoglycoproteins. J Biol Chem. 1974 Sep 10;249(17):5536–5543. [PubMed] [Google Scholar]
  20. Kawasaki T., Ashwell G. Carbohydrate structure of glycopeptides isolated from an hepatic membrane-binding protein specific for asialoglycoproteins. J Biol Chem. 1976 Sep 10;251(17):5292–5299. [PubMed] [Google Scholar]
  21. Kawasaki T., Ashwell G. Chemical and physical properties of an hepatic membrane protein that specifically binds asialoglycoproteins. J Biol Chem. 1976 Mar 10;251(5):1296–1302. [PubMed] [Google Scholar]
  22. Kawasaki T., Ashwell G. Isolation and characterization of an avian hepatic binding protein specific for N-acetylglucosamine-terminated glycoproteins. J Biol Chem. 1977 Sep 25;252(18):6536–6543. [PubMed] [Google Scholar]
  23. Kawasaki T., Etoh R., Yamashina I. Isolation and characterization of a mannan-binding protein from rabbit liver. Biochem Biophys Res Commun. 1978 Apr 14;81(3):1018–1024. doi: 10.1016/0006-291x(78)91452-3. [DOI] [PubMed] [Google Scholar]
  24. 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]
  25. LaBadie J. H., Chapman K. P., Aronson N. N., Jr Glycoprotein catabolism in rat liver: Lysosomal digestion of iodinated asialo-fetuin. Biochem J. 1975 Nov;152(2):271–279. doi: 10.1042/bj1520271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Morell A. G., Gregoriadis G., Scheinberg I. H., Hickman J., Ashwell G. The role of sialic acid in determining the survival of glycoproteins in the circulation. J Biol Chem. 1971 Mar 10;246(5):1461–1467. [PubMed] [Google Scholar]
  27. Morell A. G., Irvine R. A., Sternlieb I., Scheinberg I. H., Ashwell G. Physical and chemical studies on ceruloplasmin. V. Metabolic studies on sialic acid-free ceruloplasmin in vivo. J Biol Chem. 1968 Jan 10;243(1):155–159. [PubMed] [Google Scholar]
  28. NOVIKOFF A. B., ESSNER E. The liver cell. Some new approaches to its study. Am J Med. 1960 Jul;29:102–131. doi: 10.1016/0002-9343(60)90011-5. [DOI] [PubMed] [Google Scholar]
  29. Naito M., Wisse E. Filtration effect of endothelial fenestrations on chylomicron transport in neonatal rat liver sinusoids. Cell Tissue Res. 1978 Jul 10;190(3):371–382. doi: 10.1007/BF00219553. [DOI] [PubMed] [Google Scholar]
  30. Nilsson M., Berg T. Uptake and degradation of formaldehyde-treated 125I-labelled human serum albumin in rat liver cells in vivo and in vitro. Biochim Biophys Acta. 1977 Mar 29;497(1):171–182. doi: 10.1016/0304-4165(77)90150-7. [DOI] [PubMed] [Google Scholar]
  31. Paulson J. C., Prieels J. P., Glasgow L. R., Hill R. L. Sialyl- and fucosyltransferases in the biosynthesis of asparaginyl-linked oligosaccharides in glycoproteins. Mutually exclusive glycosylation by beta-galactoside alpha2 goes to 6 sialyltransferase and N-acetylglucosaminide alpha1 goes to 3 fucosyltransferase. J Biol Chem. 1978 Aug 25;253(16):5617–5624. [PubMed] [Google Scholar]
  32. Prieels J. P., Pizzo S. V., Glasgow L. R., Paulson J. C., Hill R. L. Hepatic receptor that specifically binds oligosaccharides containing fucosyl alpha1 leads to 3 N-acetylglucosamine linkages. Proc Natl Acad Sci U S A. 1978 May;75(5):2215–2219. doi: 10.1073/pnas.75.5.2215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Salpeter M. M., Fertuck H. C., Salpeter E. E. Resolution in electron microscope autoradiography. III. Iodine-125, the effect of heavy metal staining, and a reassessment of critical parameters. J Cell Biol. 1977 Jan;72(1):161–173. doi: 10.1083/jcb.72.1.161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Schlesinger P. H., Doebber T. W., Mandell B. F., White R., DeSchryver C., Rodman J. S., Miller M. J., Stahl P. Plasma clearance of glycoproteins with terminal mannose and N-acetylglucosamine by liver non-parenchymal cells. Studies with beta-glucuronidase, N-acetyl-beta-D-glucosaminidase, ribonuclease B and agalacto-orosomucoid. Biochem J. 1978 Oct 15;176(1):103–109. doi: 10.1042/bj1760103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Silverstein S. C., Steinman R. M., Cohn Z. A. Endocytosis. Annu Rev Biochem. 1977;46:669–722. doi: 10.1146/annurev.bi.46.070177.003321. [DOI] [PubMed] [Google Scholar]
  36. Stahl P. D., Touster O. Beta-glucuronidase of rat liver lysosomes. Purification, properties, subunits. J Biol Chem. 1971 Sep 10;246(17):5398–5406. [PubMed] [Google Scholar]
  37. Stahl P., Rodman J. S., Schlesinger P. Clearance of lysosomal hydrolases following intravenous infusion. Kinetic and competition experiments with beta-glucuronidase and N-acetyl-beta-D-glucosaminidase. Arch Biochem Biophys. 1976 Dec;177(2):594–605. doi: 10.1016/0003-9861(76)90471-9. [DOI] [PubMed] [Google Scholar]
  38. Stahl P., Schlesinger P. H., Rodman J. S., Doebber T. Recognition of lysosomal glycosidases in vivo inhibited by modified glycoproteins. Nature. 1976 Nov 4;264(5581):86–88. doi: 10.1038/264086a0. [DOI] [PubMed] [Google Scholar]
  39. Stahl P., Six H., Rodman J. S., Schlesinger P., Tulsiani D. R., Touster O. Evidence for specific recognition sites mediating clearance of lysosomal enzymes in vivo. Proc Natl Acad Sci U S A. 1976 Nov;73(11):4045–4049. doi: 10.1073/pnas.73.11.4045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Stockert R. J., Morell A. G., Scheinberg I. H. The existence of a second route for the transfer of certain glycoproteins from the circulation into the liver. Biochem Biophys Res Commun. 1976 Feb 9;68(3):988–993. doi: 10.1016/0006-291x(76)91243-2. [DOI] [PubMed] [Google Scholar]
  41. THORBECKE G. J., MAURER P. H., BENACERRAF B. The affinity of the reticulo-endothelial system for various modified serum proteins. Br J Exp Pathol. 1960 Apr;41:190–197. [PMC free article] [PubMed] [Google Scholar]
  42. Tanabe T., Pricer W. E., Jr, Ashwell G. Subcellular membrane topology and turnover of a rat hepatic binding protein specific for asialoglycoproteins. J Biol Chem. 1979 Feb 25;254(4):1038–1043. [PubMed] [Google Scholar]
  43. Tolleshaug H., Berg T., Nilsson M., Norum K. R. Uptake and degradation of 125I-labelled asialo-fetuin by isolated rat hepatocytes. Biochim Biophys Acta. 1977 Aug 25;499(1):73–84. doi: 10.1016/0304-4165(77)90230-6. [DOI] [PubMed] [Google Scholar]
  44. Tulsiani D. R., Keller R. K., Touster O. The preparation and chemical composition of the multiple forms of beta-glucuronidase from the female rat preputial gland. J Biol Chem. 1975 Jun 25;250(12):4770–4776. [PubMed] [Google Scholar]
  45. VENABLE J. H., COGGESHALL R. A SIMPLIFIED LEAD CITRATE STAIN FOR USE IN ELECTRON MICROSCOPY. J Cell Biol. 1965 May;25:407–408. doi: 10.1083/jcb.25.2.407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Van Lenten L., Ashwell G. Studies on the chemical and enzymatic modification of glycoproteins. A general method for the tritiation of sialic acid-containing glycoproteins. J Biol Chem. 1971 Mar 25;246(6):1889–1894. [PubMed] [Google Scholar]
  47. WARREN L. The thiobarbituric acid assay of sialic acids. J Biol Chem. 1959 Aug;234(8):1971–1975. [PubMed] [Google Scholar]
  48. Widmann J. J., Cotran R. S., Fahimi H. D. Mononuclear phagocytes (Kupffer cells) and endothelial cells. Identification of two functional cell types in rat liver sinusoids by endogenous peroxidase activity. J Cell Biol. 1972 Jan;52(1):159–170. doi: 10.1083/jcb.52.1.159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Wilson G. Effect of reductive lactosamination on the hepatic uptake of bovine pancreatic ribonuclease A dimer. J Biol Chem. 1978 Apr 10;253(7):2070–2072. [PubMed] [Google Scholar]
  50. Wisse E. An electron microscopic study of the fenestrated endothelial lining of rat liver sinusoids. J Ultrastruct Res. 1970 Apr;31(1):125–150. doi: 10.1016/s0022-5320(70)90150-4. [DOI] [PubMed] [Google Scholar]
  51. Wisse E. An ultrastructural characterization of the endothelial cell in the rat liver sinusoid under normal and various experimental conditions, as a contribution to the distinction between endothelial and Kupffer cells. J Ultrastruct Res. 1972 Mar;38(5):528–562. doi: 10.1016/0022-5320(72)90089-5. [DOI] [PubMed] [Google Scholar]
  52. Wisse E. Kupffer cell reactions in rat liver under various conditions as observed in the electron microscope. J Ultrastruct Res. 1974 Mar;46(3):499–520. doi: 10.1016/s0022-5320(74)90070-7. [DOI] [PubMed] [Google Scholar]
  53. Wisse E. Observations on the fine structure and peroxidase cytochemistry of normal rat liver Kupffer cells. J Ultrastruct Res. 1974 Mar;46(3):393–426. doi: 10.1016/s0022-5320(74)90064-1. [DOI] [PubMed] [Google Scholar]

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

RESOURCES