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Binding and endocytosis of heparin-gold conjugates by the fenestrated endothelium of the rat choriocapillaris

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Heparin-gold probes were used to localize regions of heparin binding on the luminal surface of the diaphragmed-fenestrated endothelium of the rat choriocapillaris. Structures of endothelial cells were unlabeled when rats were kept on ice and perfused with solutions at 4° C. When the heparin-gold quantity was doubled, only a few heparin-gold particles marked some diaphragms spanning fenestrae, vesicles and channels, parajunctional regions of the plasmalemma, and coated pits. With solutions at 4° C, but the animals kept at room temperature, all of these structures in the endothelial cells were labeled. This binding was not altered by the perfusion of low levels of unlabeled heparin, but was eliminated following high levels of unlabeled heparin, and by digestion with trypsin and pronase. At 37° C, heparin was localized to the above structures and, in addition, was internalized into coated vesicles, endosomes, and multivesicular bodies, but not other types of lysosomes. Some particles were found in tubules adjacent to the Golgi stacks. Heparin-gold was also transported to the abluminal front of the endothelium by vesicles. A desulfated, non-anticoagulant, fraction of heparin bound to gold was localized to the endothelium in the same manner. These results demonstrate receptors for heparin on the surface of a fenestrated endothelium. Furthermore, they show the pathway of endocytosis and transport of heparin. The possible roles of heparin in the function of the endothelium is discussed.

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  1. Bitter T, Muir HM (1962) A modified uronic acid carbazole reaction. Anal Biochem 4:330–334

  2. Carr J (1979) The anti-inflammatory action of heparin as an antagonist to histamine, bradykinin and prostaglandin El. Thromb Res 16:507–516

  3. Castellot JJ, Wong K, Herman B, Albertini DF, Wright TC, Hoover RL, Karnovsky MJ (1983) Binding and internalization of heparin by vascular smooth muscle cells. J Cell Biol 97:431 a

  4. Castellot JJ, Wong K, Herman B, Hoover RL, Albertini DF, Wright TC, Caleb BL, Karnovsky MJ (1985) Binding and internalization of heparin by vascular smooth muscle cells. J Cell Physiol 124:13–20

  5. Dougherty TF, Dolowitz DA (1964) Physiologic actions of heparin not related to blood clotting. Am J Cardiol 14:18–24

  6. Engelberg H (1985) Heparin, heparin fractions, and the atherosclerotic process. Semin Thromb Hemost 11:48–55

  7. Essner E, Gordon SR (1983) Observations on the permeability of the choriocapillaris of the eye. Cell Tissue Res 231:571–577

  8. Fabian I, Bleiberg I, Aronson M (1978) Increased uptake and desulfation of heparin by mouse macrophages in the presence of cations. Biochim Biophys Acta 544:69–76

  9. Farquhar MG (1978) Recovery of surface membrane in anterior pituitary cells. Variations in traffic detected with anionic and cationic ferritin. J Cell Biol 77:R35-R42

  10. Farquhar MG, Palade GE (1981) The Golgi apparatus (complex)-(1954–1981)-from artifact to center stage. J Cell Biol 91:77S-103S

  11. Florey HW, Poole JCF, Meek GA (1959) Endothelial cells and “cement” lines. J Pathol Bact 77:625–636

  12. Folkman J, Langer R, Linhardt RJ, Haudenschild C, Taylor S (1983) Angiogenesis inhibition and tumor regression caused by heparin or heparin fragment in the presence of cortisone. Science 221:719–725

  13. Geisow MJ, Beaven GH, D'Arcy Hart P, Young MR (1980) Site of action of a polyanion inhibitor of phagosome-lysosome fusion in cultured macrophages. Exp Cell Res 126:159–165

  14. Geoghegan WD, Ackerman GA (1977) Adsorption of horseradish peroxidase, ovomucoid and anti-immunoglobulin to colloidal gold for the indirect detection of concanavalin A, wheat germ agglutinin and goat anti-human immunoglobulin G on cell surfaces at the electron microscopic level: a new method, theory and application. J Histochem Cytochem 25:1187–1200

  15. Geuze HJ, Slot JW, Strous GJAM, Lodish HF, Schwartz AL (1983) Intracellular site of asialoglycoprotein receptor-ligand uncoupling: double-label immunoelectron microscopy during receptor-medicated endocytosis. Cell 32:277–286

  16. Glimelius B, Busch C, Hook M (1978) Binding of heparin on the surface of cultured human endothelial cells. Thromb Res 12:773–782

  17. Goldstein JL, Anderson RGW, Brown MS (1979) Coated pits, coated vesicles, and receptor-mediated endocytosis. Nature 279:679–685

  18. Hart TK, Pino RM (1986) Capillary permeability in the pancreas and colon: restriction of exogenous and endogenous molecules by fenestrated endothelia. Am J Anat 175:49–58

  19. Henkind P (1978) Ocular neovascularization. Am J. Ophthal 85:287–301

  20. Hiebert LM, Jacques LB (1976) The observation of heparin on endothelium after injection. Thromb Res 8:195–204

  21. Hoover RL, Rosenberg RD, Haering W, Karnovsky MJ (1980) Inhibition of rat arterial smooth muscle cell proliferation by heparin. II. In vitro studies. Circ Res 47:578–583

  22. Hynes RO, Yamada KM (1982) Fibronectin: multifunctional modular glycoproteins. J Cell Biol 95:369–377

  23. Inoue Y, Nagasawa K (1976) Selective N-desulfation of heparin with dimethylsulfoxide containg water or methanol. Carbohydr Res 46:87–95

  24. Kjellen L, Oldberg A, Rubin K, Hook M (1977) Binding of heparin and heparan sulfate to rat liver cells. Biochem Biophys Res Comm 74:126–133

  25. Livne E, Oliver C (1986) Internalization of cationized ferritin by isolated pancreatic acinar cells. J Histochem Cytochem 34:167–176

  26. Luft JH (1961) Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol 9:409–414

  27. Maciag T, Mehlman T, Friesel R (1984) Heparin binds endothelial cell growth factor, the principal endothelial cell mitogen in bovine brain. Science 225:932–934

  28. Muhlpfordt H (1982) The preparation of colloidal gold particles using tannic acid as an additional reducing agent. Experientia 38:1127–1128

  29. Muller LI, Jacks TJ (1975) Rapid chemical dehydration of samples for electron microscopic examinations. J Histochem Cytochem 23:107–110

  30. Novikoff AB, Novikoff PM (1977) Cytochemical contributions to differentiating GERL from the Golgi apparatus. Histochem J 9:525–551

  31. Pasten IH, Willingham MC (1981) Journey to the center of the cell: role of receptosome. Science 214:504–509

  32. Pelikan P, Gimbrone MA, Cotran RS (1979) Distribution and movement of anionic cell surface sites in cultured human vascular endothelial cells. Atherosclerosis 32:69–80

  33. Pino RM (1984) Heparin and heparan sulfate domains on a fenestrated capillary. Abstracts of the Third Intl Symp of the Biol of the Vascular Endothelial Cell, Cambridge, MA: 32

  34. Pino RM (1985a) Heparin and the rat choriocapillary endothelium. Invest Ophthal Vis Sci 26[Suppl]327

  35. Pino RM (1985b) Restriction to endogenous plasma proteins by a fenestrated capillary endothelium: an ultrastructural immunocytochemical study of the choriocapillary endothelium. Am J Anat 172:279–289

  36. Pino RM (1986a) The cell surface of a restrictive fenestrated endothelium. I. Distribution of lectin-receptor monosaccharides on the choriocapillaris. Cell Tissue Res 243:145–155

  37. Pino RM (1986b) The cell surface of a restrictive fenestrated capillary endothelium. II. Dynamics of cationic ferritin binding and the identification of heparin and heparan sulfate domains on the choriocapillaris. Cell Tissue Res 243:157–164

  38. Pino RM (1986c) Restriction to exogenous transtheyretin (prealbumin) by the endothelium of the rat choriocapillaris. Am J Anat 177:63–70

  39. Pino RM, Essner E (1980) Structure and permeability to ferritin of the choriocapillary endothelium of the rat eye. Cell Tissue Res 208:21–27

  40. Pino RM, Essner E (1981) Permeability of rat choriocapillaris to hemeproteins. Restriction of tracers by a fenestrated endothelium. J Histochem Cytochem 29:281–290

  41. Reynolds ES (1963) The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol 17:155–159

  42. Roth J, Bendayan M, Orci L (1978) Ultrastructural localization of intracellular antigens by the use of a protein A-gold complex. J Histochem Cytochem 26:1074–1081

  43. Ryser H J-P, Morad N, Shein W-C (1983) Heparin interaction with cultured cells: possible role of fibronectin in uncoupling surface binding and endocytosis. Cell Biol Int Rep 7:923–930

  44. Samuels PB, Webster DR (1952) The role of venous endothelium in the inception of thrombosis. Ann Surg 136:422–438

  45. Shanberge JN, Kambayashi J, Nakagawa M (1976) The interaction of platelets with a tritium-labeled heparin. Thromb Res 9:595–609

  46. Shen W-C, Ryser H J-P (1981) Poly (L-lysine) has different membrane-transport and drug-carrier properties when complexed with heparin. Proc Natl Acad Sci USA 78:7589–7593

  47. Shing Y, Folkman J, Sullivan R, Butterfield C, Murray J, Klagsbrun M (1984) Heparin affinity: purification of a tumor-derived capillary endothelial cell growth factor. Science 223:1296–1299

  48. Simionescu M, Simionescu N, Silbert JE, Palade GE (1984) Differentiated microdomains on the luminal surface of the capillary endothelium. II. Partial characterization of their anionic sites. J Cell Biol 90:614–621

  49. Skutelsky E, Danon D (1976) Redistribution of surface anionic sites on the luminal front of blood vessel endothelium after interaction with polycationic ligand. J Cell Biol 71:232–241

  50. Van Deurs B, von Bulow F, Moller M (1981) Vesicular transport of cationized ferritin by the epithelium of the rat choroid plexus. J Cell Biol 89:131–139

  51. Wright TC, Johnstone TV, Castellot JJ, Karnovsky MJ (1985) Inhibition of rat cervical epithelial cell growth by heparin and its reversal by EGF. J Cell Physiol 125:499–506

  52. Wolff JR (1977) Ultrastructure of the terminal vascular bed as related to function. In: Kaley G, Altura BM (eds) Microcirculation, Vol. I. University Park Press, Baltimore, pp 95–130

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Correspondence to Dr. Richard M. Pino.

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Pino, R.M. Binding and endocytosis of heparin-gold conjugates by the fenestrated endothelium of the rat choriocapillaris. Cell Tissue Res. 250, 257–266 (1987). https://doi.org/10.1007/BF00219070

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Key words

  • Choriocapillaris
  • Endocytosis
  • Endothelium
  • Eye
  • Glycosaminoglycan
  • Heparin
  • Receptors
  • Rat