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Thromb Haemost 1988; 60(02): 240-246
DOI: 10.1055/s-0038-1647038
Original Article
Schattauer GmbH Stuttgart

Characterization of an In Vitro Model to Study the Permeability of Human Arterial Endothelial Cell Monolayers

Erna G Langeler
The Gaubius Institute TNO, Leiden, The Netherlands
,
Victor W M van Hinsbergh
The Gaubius Institute TNO, Leiden, The Netherlands
› Author Affiliations
Further Information

Publication History

Received 11 April 1988

Accepted after revision 09 June 1988

Publication Date:
28 June 2018 (online)

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Summary

A model has been developed to study the transport of fluid and macromolecules through human arterial umbilical cord endothelial cell monolayers in vitro. Cells were cultured on fibronectin- coated polycarbonate filters and formed within a few days a tight monolayer, with an electrical resistance of 17 ± 4 Ohm · cm2. The cells were connected by close cell contacts with tight junctions. The passáge-rate of horse radish peroxidase (HRP) through these filters was 20-40 fold lower than through filters without an endothelial monolayer. The continuous presence of 10% human serum was needed to maintain the electrical resistance of the monolayer and its barrier function towards macromolecules. Chelation of extracellular calcium resulted in an increased permeability and a decreased electrical resistance of the monolayer. This process was reversible by re-addition of calcium ions to the cells. The permeation rate of dextrans of various molecular weights (9-480 kD) was inversely related to the molecular mass of the molecule. No difference was measured between the passage rate of dextran of 480 kD and dextran of 2,000 kD. Incubation of the endothelial cell monolayer with 2-deoxy-D-glucose resulted in a decreased permeability but it had no effect on electrical resistance. This suggests that the passage-process is energy- dependent.

Fluid permeation through the endothelial cell monolayer on filters was measured in a perfusion chamber under 20 mmHg hydrostatic pressure. It was decreased by the presence of serum proteins and responded reversibly on the chelation and readdition of extracellular calcium ions.

Keywords

Endothelial cell monolayer - Passage of macromolecules
 

  • References

  • 1 Bennett HS, Luft JJ, Hampton JC. Morphological classification of vertebrate blood capillaries.. Am J Physiol 1959; 196: 381-390
    CrossrefPubMedGoogle Scholar
  • 2 Renkin EM. Capillary transport of macromolecules: pores and other endothelial pathways.. J Appl Physiol 1985; 58: 315-325
    CrossrefPubMedGoogle Scholar
  • 3 Curry FRE. Determinants of capillary permeability: A review of mechanisms based on single capillary studies in the frog.. Circ Res 1986; 59: 367-380
    CrossrefPubMedGoogle Scholar
  • 4 Majno G, Shea SM, Leventhal M. Endothelial contractions induced by histamine-type mediators.. J Cell Biol 1969; 42: 647-672
    CrossrefPubMedGoogle Scholar
  • 5 Joris I, Majno G, Ryan GB. Endothelial contraction in vivo: a study of the rat mesentery. Virchows Arch Abt Zellpathol 1972; 12: 73-88
    PubMedGoogle Scholar
  • 6 Bruns RR, Palade GE. Studies on blood capillaries. II. Transport of ferritin molecules across the wall of muscle capillaries.. J Cell Biol 1968; 37: 277-299
    CrossrefPubMedGoogle Scholar
  • 7 Clough C, Michel CC. The role of vesicles in the transport of ferritin through frog endothelium.. J Physiol 1981; 315: 127-142
    CrossrefPubMedGoogle Scholar
  • 8 Simionescu N. Transcytosis and traffic of membranes in the endothelial cell.. In: International Cell Biology 1980-1981 Schweiger HG. (ed) Springer-Verlag; Berlin : 1981. pp 657-672
    Google Scholar
  • 9 King GL, Johnson SM. Receptor-mediated transport of insulin across endothelial cells.. Science 1985; 227: 1583-1586
    CrossrefPubMedGoogle Scholar
  • 10 Hashida R, Anamizu C, Kimura J, Ohkuma S, Yoshida Y, Takano T. Transcellular transport of lipoprotein through arteriel endothelial cells in monolayer culture.. Cell Structure Function 1986; 11: 31-42
    CrossrefPubMedGoogle Scholar
  • 11 Simionescu M, Simionescu N. Isolation of endothelial cells from the heart microvasculature.. Microvasc Res 1978; 16: 426-452
    CrossrefPubMedGoogle Scholar
  • 12 Renkin EM. Multiple pathway of capillary permeability.. Circ Res 1977; 41: 735-743
    CrossrefPubMedGoogle Scholar
  • 13 Navab M, Hough GP, Berliner JA, Frank JA, Fogelman AM, Haberland ME, Edwards PA. Rabbit beta-migrating very low density lipoprotein increases endothelial macromolecular transport without altering electrical resistance.. J Clin Invest 1986; 78: 389-397
    CrossrefPubMedGoogle Scholar
  • 14 Cooper JA, Del VecchioPJ, Minnear FL, Burhop KE, Selig WM, Garcia JG N, Malik AB. Measurement of albumin permeability across endothelial monolayers in vitro.. J Appl Physiol 1987; 62: 1076-1083
    CrossrefPubMedGoogle Scholar
  • 15 Olesen SP, Crone C. Electrical resistance of muscle capillary endothelium.. Biophys J 1983; 42: 31-41
    CrossrefPubMedGoogle Scholar
  • 16 Hodson SA, Wigham CG. Paracellular ionic and transcellular water diffusions across rabbit corneal endothelium.. J Physiol 1987; 385: 89-96
    CrossrefPubMedGoogle Scholar
  • 17 Siflinger-Birnboim A, Del VecchioPJ, Cooper JA, Blumenstock FA, Shepard JM, Malik AB. Molecular sieving characteristics of the cultured endothelial monolayer.. J Cell Physiol 1987; 132: 111-117
    CrossrefPubMedGoogle Scholar
  • 18 Hashida R, Anamizu C, Yagyu-Mizuno Y, Ohkuma S, Takano T. Transcellular transport of fluorescein dextran through an arterial endothelial cell monolayer.. Cell Structure Function 1986; 11: 343-349
    CrossrefPubMedGoogle Scholar
  • 19 Kern DF, Malik AB. Microvascular albumin permeability in isolated perfused lung: effects of EDTA.. J Appl Physiol 1985; 58: 372-375
    CrossrefPubMedGoogle Scholar
  • 20 Shasby DM, Shasby SS. Effects of calcium on transendothelial albumin transfer and electrical resistance.. J Appl Physiol 1986; 60: 71-79
    CrossrefPubMedGoogle Scholar
  • 21 Baetscher M, Brune K. An in vitro system for measuring endothelial permeability under hydrostatic pressure.. Exp Cell Res 1983; 148: 541-547
    CrossrefPubMedGoogle Scholar
  • 22 Harlan JM, Schwartz BR, Reidy MA, Schwartz SM, Ochs HD, Harker LA. Activated neutrophils disrupt endothelial monolayer integrity by an oxygen radical-independent mechanism. Lab Invest 1985; 52: 141-150
    PubMedGoogle Scholar
  • 23 Maciag T, Cerundolo J, Ilsley S, Kelley PR, Forand R. An endothelial cell growth factor from bovine hypothalamus: identification and partial characterization.. Proc Natl Acad Sci USA 1979; 76: 5674-5678
    CrossrefPubMedGoogle Scholar
  • 24 Jaffe EA, Nachman RL, Becker CG, Minick CR. Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria.. J Clin Invest 1973; 52: 2745-2756
    CrossrefPubMedGoogle Scholar
  • 25 Van Hinsbergh VWM, Mommaas-Kienhuis AM, Weinstein R, Maciag T. Propagation and morphologic phenotypes of human umbilical cord artery endothelial cells. Eur J Cell Biol 1986; 42: 101-110
    PubMedGoogle Scholar
  • 26 Scott Jr TA, Melvin EH. Determination of dextran with anthrone.. Anal Chem 1953; 25: 1656-1661
    CrossrefPubMedGoogle Scholar
  • 27 Simionescu M, Simionescu N, Palade GE. Segmental differentiations of cell junctions in the vascular endothelium.. J Cell Biol 1976; 68: 705-723
    CrossrefPubMedGoogle Scholar
  • 28 Schneeberger EE, Lynch RD. Tight junctions. Their structure, composition, and function.. Circ Res 1984; 55: 723-733
    CrossrefPubMedGoogle Scholar
  • 29 Powell D. Barrier function of epithelia.. Am J Physiol 1981; 241G: 275-288
    PubMedGoogle Scholar
  • 30 Territo M, Berliner JA, Fogelman AM. Effect of monocyte migration on low density lipoprotein transport across aortic endothelial cell monolayers.. J Clin Invest 1984; 74: 2279-2284
    CrossrefPubMedGoogle Scholar
  • 31 Mason JC, Curry FE, Michel CC. The effects of proteins upon the filtration coefficient of individually perfused from mesenteric capillaries.. Microvasc Res 1977; 13: 185-202
    CrossrefPubMedGoogle Scholar
  • 32 Levick JR, Michel CC. The effect of bovine albumin on the permeability of frog mesenteric capillaries. Q J Exp Physiol 1973; 58: 87-97
    PubMedGoogle Scholar
  • 33 Danielli JF. Capillary permeability and oedema in the perfused frog.. J Physiol 1940; 98: 109-129
    CrossrefPubMedGoogle Scholar
  • 34 Pappenheimer JR, Soto-Rivera A. Effective osmotic pressure of the plasma proteins and other quantities associated with the capillary circulation in the hind-limbs of cats and dogs.. Am J Physiol 1948; 152: 471-491
    PubMedGoogle Scholar
  • 35 Michel CC, Philips ME, Turner RM. The effects of native and modified bovine serum albumin on the permeability of frog mesenteric capillaires.. J Physiol 1985; 360: 333-346
    CrossrefPubMedGoogle Scholar
  • 36 Schneeberger EE, Hamelin M. Interaction of serum proteins with lung endothelial glycocalyx: its effect on endothelial permeability.. Am J Physiol 1984; 16H: 206-217
    PubMedGoogle Scholar
  • 37 Curry FE, Michel CC. A fiber matrix theory of capillary permeability.. Microvasc Res 1980; 20: 96-99
    CrossrefPubMedGoogle Scholar
  • 38 Chambers R, Zweifach BW. Capillary endothelial cement in relation to permeability.. J Cell Comp Physiol 1940; 15: 255-272
    CrossrefPubMedGoogle Scholar
  • 39 Nicolaysen G. Intravascular concentrations of calcium and magnesium ions and edema formation in isolated lungs.. Acta Physiol Scand 1971; 81: 325-339
    CrossrefPubMedGoogle Scholar
  • 40 Grotte G. Passage of dextran molecules across the blood-lymph barrier.. Acta Chir Scand 1956; 211: 01-84
    PubMedGoogle Scholar
  • 41 Simionescu N. Cellular aspects of transcapillary exchange. Physiol Rev 1983; 63: 1536-1579
    CrossrefPubMedGoogle Scholar
  • 42 Carter RD, Joyner WL, Renkin EM. Effects of histamine and some other substances on molecular selectivity of the capillary wall to plasma proteins and dextran.. Microvasc Res 1974; 7: 31-48
    CrossrefPubMedGoogle Scholar