Skip to main content
SpringerLink
Log in

Relationship between pinocytic rate and uptake of transferrin by suspended rat hepatocytes

  • Original Articles
  • Published:
Biology of Metals Aims and scope Submit manuscript

Summary

The aim of this study was to compare quantitatively the capacity to transcytose (i.e. to uptake and release) transferrin (Tf) with the pinocytic activity of suspended adult rat hepatocytes. An oligodisperse preparation of131I-polyvinylpyrrolidone (PVP;M r 36000) was used to measure the inward and outward aspects of the pinocytic process in separate experiments. Cell association of rat125I-Tf was measured at Tf concentrations approaching physiological, where59Fe uptake obeyed first-order kinetics. Release studies with both PVP and Tf were carried out under conditions which minimized the probability ofde novo endocytosis of a molecule already released. Sets of experimental points representing cell-associated radioactivities were converted into continuous algebraic functions by fitting with two-term (release studies) or three-term (uptake studies) exponential equations. Transport of PVP and Tf through the cells was computed from these equations by deconvolution. This analysis showed that, under the present experimental conditions, the fractional transcytosis rates of Tf and PVP by hepatocytes were in the ratio of I:0.77. These values imply that, in the physiological range of Tf concentrations, about 75% of the Fe taken up by hepatocytes may be due to a pinocytic mechanism (fluid-phase or mixed). Inclusion of chloroquine (1 mM) in the suspending medium, both in uptake and release experiments, resulted in more PVP and Tf passing through the cells, while Fe uptake was reduced. It is suggested that the base probably exerted its enhancing effect on transcytosis by shunting the subcellular transport of PVP and Tf to the outward leg through a shorter circuit.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

BSA:

bovine serum albumin

HBSS:

Hank's balanced salt solution

Hepes:

4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid

MEM:

minimal essential medium

PVP:

polyvinylpyrrolidone

Tf:

transferrin

References

  • Baker E, McArdle HJ, Morgan EH (1985) Transferrin-cell interactions: studies with erythroid, placental and hepatic cells. In: Spik G, Montreuil J, Crichton RR, Mazurier J (eds) Proteins of iron storage and transport. Elsevier, Amsterdam, pp 131–142

    Google Scholar 

  • Besterman JM, Low RB (1983) Endocytosis: a review of mechanisms and plasma membrane dynamics. Biochem J 210:l-13

    Google Scholar 

  • Besterman JM, Airhart JA, Woolworth RC, Low RB (1981) Endocytosis of pinocytosed fluid in cultured cells: kinetic evi dence for rapid turnover and compartmentation. J Cell Biol 91:716–727

    PubMed  Google Scholar 

  • Bomford AB, Munro HN (1985) Transferrin and its receptor: their roles in cell function. Hepatology 5:870–875

    PubMed  Google Scholar 

  • Casey KA, Maurey KM, Storrie B (1986) Characterization of early compartments in fluid phase pinocytosis: a cell fractionation study. J Cell Sci 83:119–133

    PubMed  Google Scholar 

  • Cole ES, Glass J (1983) Transferrin binding and iron uptake in mouse hepatocytes. Biochim Biophys Acta 762:102–110

    PubMed  Google Scholar 

  • Dean RT, Jessup W, Roberts CR (1984) Effects of exogenous amines on mammalian cells, with particular reference to membrane flow. Biochern J 217:27–40

    Google Scholar 

  • De Duve C, De Barsy T, Poole B, Trouet A, Tulkens P, Van Hoof F (1974) Lysosomotropic agents. Biochem Pharmacol 23:2495–2531

    PubMed  Google Scholar 

  • Duncan R, Pratten MK, Cable HC, Ringsdorf H, Lloyd JB (1981) Effect of molecular size of125I-Labelled poly(vinylpyrrolidone) on its pinocytosis by rat visceral yolk sacs and rat peritoneal macrophages. Biochem J 196:49–55

    PubMed  Google Scholar 

  • Hasilik A, Neufeld EF (1980) Biosynthesis of lysosomal enzymes in fibroblasts. Synthesis as precursors of higher molecular weight. J Biol Chem 255:4937–4945

    PubMed  Google Scholar 

  • Hoffenberg R, Gordon AH, Black EG, Louis LN (1970) Plasma protein catabolism by the perfused rat liver. The effect of alteration of albumin concentration and dietary protein depletion. Biochem J 118:401–404

    PubMed  Google Scholar 

  • Holmes JM, Morgan EH (1989) Uptake and distribution of transferrin and iron in perfused, iron-deficient rat liver. Am J Physiol 256:GI022-GIO27

    Google Scholar 

  • Hornick CA, Jones AL, Renaud G, Hradek G, Havel RJ (1984) Effect of chloroquine on low-density lipoprotein catabolic pathway in rat hepatocytes. Am J Physiol 246:GI87–GI94

    Google Scholar 

  • Jacques PJ (1973) Endocytosis. In: Dingle JT, Fell HB (eds) Lysosomes in biology and pathology, vol. 2. North-Holland, Amsterdam, pp 395–420

    Google Scholar 

  • Knutson VP, Ronnett GV, Lane MD (1985) The effects of cycloheximide and chloroquine on insulin receptor metabolism. J Biol Chem 260:14180–14188

    PubMed  Google Scholar 

  • Mellman I, Fuchs R, Helenius A (1986) Acidification of the endocytic and exocytic pathways. Annu Rev Biochem 55:663–700

    PubMed  Google Scholar 

  • Moore HP, Gumbiner B, Kelly RB (1983) Chloroquine diverts ACTH from a regulated to a constitutive secretory pathway. Nature 302:434–436

    PubMed  Google Scholar 

  • Nelder JA, Mead R (1965) A simplex method for function minimization. Comput J 7:308–313

    Google Scholar 

  • Ose L, Ose T, Reinertsen R, Berg T (1980) Fluid endocytosis in isolated rat parenchyma) and non-parenchyma) liver cells. Exp Cell Res 126:109–119

    PubMed  Google Scholar 

  • Page MA, Baker E, Morgan EH (1984) Transferrin and iron uptake by rat hepatocytes in culture. Am I Physiol 246:G26–G33

    Google Scholar 

  • Rao K, van Renswoude J, Kempf C, Klausner RD (1983) Separation of Fe3+ from transferrin in endocytosis: role of the acidic endosome. FEBS Lett 160:213–216

    PubMed  Google Scholar 

  • Regoeczi E (1976) Labelled polyvinylpyrrolidone as an in vivo indicator of reticuloendothelial activity. Br J Exp Pathol 57:431–442

    PubMed  Google Scholar 

  • Regoeczi E, Debanne MT, Hatton MWC, Koj A (1978) Elimination of asialofetuin and asialoorosomucoid by the intact rat. Quantitative aspects of the hepatic clearance mechanism. Biochim Biophys Acta 541:372–384

    PubMed  Google Scholar 

  • Rudolph JR (1988) The interaction of rat transferrin with the liver with special reference to the glycan of transferrin. Doctoral thesis, McMaster University, Hamilton, Ontario, Canada, pp 295–296

    Google Scholar 

  • Rudolph JR, Regoeczi E (1988) Interaction of rat asialotransferrin with adult rat hepatocytes: its relevance for iron uptake and protein degradation. J Cell Physiol 135:539–544

    PubMed  Google Scholar 

  • Rudolph JR, Regoeczi E, Chindemi PA, Debanne MT (1986) Preferential hepatic uptake of iron from rat asialotransferrin: possible engagement of two receptors. Am J Physiol 251:G398–G404

    PubMed  Google Scholar 

  • Rudolph JR, Regoeczi E, Southward S (1988) Quantification of rat hepatocyte transferrin receptors with poly- and monoclonal antibodies and protein A. Histochemistry 88:187–192

    PubMed  Google Scholar 

  • Ryan BF, Joiner BL, Ryan TA Jr (1985) Minitab, 2nd edn, Duxbury Press, Boston, pp 193–217

    Google Scholar 

  • Scharschmidt BF, Lake JR, Renner EL, Licko V, Van Dyke RW (1986) Fluid phase endocytosis by cultured rat hepatocytes and perfused rat liver: implications for plasma membrane turnover and vesicular trafficking of fluid phase markers. Proc Natl Acad Sci USA 83:9488–9492

    PubMed  Google Scholar 

  • Schwartz AL, Bolognesi A, Fridovich SE (1984) Recycling of the asialoglycoprotein receptor and the effect of lysosomotropic amines in hepatoma cells. J Cell Biol 98:732–738

    PubMed  Google Scholar 

  • Shipley RA, Clarke RE (1972) Tracer methods for in vivo kinetics. Academic Press, New York, pp 205–208

    Google Scholar 

  • Sibille JC, Octave JN, Schneider YJ, Trouet A, Crichton RR (1982) Transferrin protein and iron uptake by cultured hepatocytes. FEBS Lett 150:365–369

    PubMed  Google Scholar 

  • Spady DK, Turley SD, Dietschy JM (1985) Receptor-independent low density lipoprotein transport in the rat in vivo. J Clin Invest 76:1113–1122

    PubMed  Google Scholar 

  • Stenseth K, Thyberg J (1989) Monensin and chloroquine inhibit transfer to lysosomes of endocytosed macromolecules in cultured mouse peritoneal macrophages. Eur J Cell Biol 49:326–333

    PubMed  Google Scholar 

  • Swanson JA, Yirinec BD, Silverstein SC (1985) Phorbol esters and horseradish peroxidase stimulate pinocytosis and redirect the flow of pinocytosed fluid in macrophages. J Cell Biol 100:851–859

    PubMed  Google Scholar 

  • Thorstensen K, Romslo I (1984) Albumin prevents nonspecific transferrin binding and iron uptake by isolated hepatocytes. Biochim Biophys Acta 804:393–397

    PubMed  Google Scholar 

  • Thorstensen K, Romslo I (1990) The role of transferrin in the mechanism of cellular iron uptake. Biochem J 271:1–10

    PubMed  Google Scholar 

  • Tolleshaug H, Berg T (1979) Chloroquine reduces the number of asialoglycoprotein receptors in the hepatocyte plasma membrane. Biochem Pharmacol 28:2919–2922

    PubMed  Google Scholar 

  • Townsend RR, Wall DA, Hubbard AL, Lee YC (1984) Rapid release of galactose-terminated ligands after endocytosis by hepatic parenchymal cells: evidence for a role of carbohydrate structure in the release of internalized ligand from receptor. Proc Natl Acad Sci USA 81:466–470

    PubMed  Google Scholar 

  • Trinder D, Morgan E, Baker E (1986) The mechanisms of iron uptake by fetal rat hepatocytes in culture. Hepatology 6:852–858

    PubMed  Google Scholar 

  • Wakshull E, Cooper JL, Wharton W (1985) Chloroquine allows the secretion of internalized125I-epidermal growth factor from fibroblasts. J Cell Physiol 125:215–222

    PubMed  Google Scholar 

  • Ward JH (1987) The structure, function, and regulation of transferrin receptor. Invest Radiol 22:74–83

    PubMed  Google Scholar 

  • Young SP, Aisen P (1981) Transferrin receptors and the uptake and release of iron by isolated hepatocytes. Hepatology 1:114–119

    PubMed  Google Scholar 

  • Zijderhand-Bleekemolen JE, Schwartz AL, Slot JW, Strous GJ, Geuze HJ (1987) Ligand- and weak-base-induced redistribution of asialoglycoprotein receptors in hepatoma cells. J Cell Biol 104:1647–1654

    PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Pathology, McMaster University Health Sciences Centre, Room 4N66, L8N 3Z5, Hamilton, Ontario, Canada

    John R. Rudolph & Erwin Regoeczi

Authors
  1. John R. Rudolph
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Erwin Regoeczi
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Rudolph, J.R., Regoeczi, E. Relationship between pinocytic rate and uptake of transferrin by suspended rat hepatocytes. Biol Metals 4, 166–172 (1991). https://doi.org/10.1007/BF01141309

Download citation

  • Received:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF01141309

Key words

Search

Navigation