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. 1986 Dec 1;103(6):2389–2402. doi: 10.1083/jcb.103.6.2389

Plasmalemmal proteins of cultured vascular endothelial cells exhibit apical-basal polarity: analysis by surface-selective iodination

PMCID: PMC2114601  PMID: 3782302

Abstract

Vascular endothelium in vivo appears to function as a polarized epithelium. To determine whether cellular polarity exists at the level of the plasma membrane, we have examined cultured endothelial monolayers for evidence of differential distribution of externally disposed plasmalemmal proteins at apical and basal cell surfaces. Lactoperoxidase beads were used to selectively label the apical surfaces of confluent endothelial monolayers, the total surfaces of nonenzymatically resuspended cells, and the basal surfaces of monolayers inverted on poly-L-lysine-coated coverslips, while maintaining greater than 98% viability in all samples. Comparison of the SDS PAGE radioiodination patterns obtained for each surface revealed a number of specific bands markedly enriched on either apical or basal surface. This polarized distribution involved membrane- associated as well as integral membrane proteins and was observed in several strains of bovine aortic endothelial cells, as well as in both primary and passaged human umbilical vein endothelial cells. In contrast, two morphologically nonpolarized cell types, bovine aortic smooth muscle and mouse peritoneal macrophages, did not display differential localization of integral membrane proteins. Polarized distribution of integral membrane proteins was established before the formation of a confluent monolayer. When inverted (basal-side-up) monolayers were returned to culture, the apical-side-up pattern was reexpressed within a few days. These results demonstrate that cell surface-selective expression of plasmalemmal proteins is an intrinsic property of viable endothelial cells in vitro. This apical/basal asymmetry of membrane structure may provide a basis for polarized endothelial functions in vivo.

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Selected References

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  1. Bartles J. R., Braiterman L. T., Hubbard A. L. Endogenous and exogenous domain markers of the rat hepatocyte plasma membrane. J Cell Biol. 1985 Apr;100(4):1126–1138. doi: 10.1083/jcb.100.4.1126. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bevilacqua M. P., Pober J. S., Majeau G. R., Cotran R. S., Gimbrone M. A., Jr Interleukin 1 (IL-1) induces biosynthesis and cell surface expression of procoagulant activity in human vascular endothelial cells. J Exp Med. 1984 Aug 1;160(2):618–623. doi: 10.1084/jem.160.2.618. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bevilacqua M. P., Pober J. S., Wheeler M. E., Cotran R. S., Gimbrone M. A., Jr Interleukin 1 acts on cultured human vascular endothelium to increase the adhesion of polymorphonuclear leukocytes, monocytes, and related leukocyte cell lines. J Clin Invest. 1985 Nov;76(5):2003–2011. doi: 10.1172/JCI112200. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bordier C. Phase separation of integral membrane proteins in Triton X-114 solution. J Biol Chem. 1981 Feb 25;256(4):1604–1607. [PubMed] [Google Scholar]
  5. Davies P. F., Kuczera L. Endocytic vesicles and surface invaginations in cultured vascular endothelium: a morphometric comparison. J Histochem Cytochem. 1981 Dec;29(12):1437–1441. doi: 10.1177/29.12.6172467. [DOI] [PubMed] [Google Scholar]
  6. Farquhar M. G., Reid J. J., Daniell L. W. Intracellular transport and packaging of prolactin: a quantitative electron microscope autoradiographic study of mammotrophs dissociated from rat pituitaries. Endocrinology. 1978 Jan;102(1):296–311. doi: 10.1210/endo-102-1-296. [DOI] [PubMed] [Google Scholar]
  7. Gimbrone M. A., Jr Culture of vascular endothelium. Prog Hemost Thromb. 1976;3:1–28. [PubMed] [Google Scholar]
  8. Graham R. C., Jr, Karnovsky M. J. The early stages of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney: ultrastructural cytochemistry by a new technique. J Histochem Cytochem. 1966 Apr;14(4):291–302. doi: 10.1177/14.4.291. [DOI] [PubMed] [Google Scholar]
  9. Harlan J. M. Leukocyte-endothelial interactions. Blood. 1985 Mar;65(3):513–525. [PubMed] [Google Scholar]
  10. Heimark R. L., Schwartz S. M. The role of membrane-membrane interactions in the regulation of endothelial cell growth. J Cell Biol. 1985 Jun;100(6):1934–1940. doi: 10.1083/jcb.100.6.1934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Horvat R., Hovorka A., Dekan G., Poczewski H., Kerjaschki D. Endothelial cell membranes contain podocalyxin--the major sialoprotein of visceral glomerular epithelial cells. J Cell Biol. 1986 Feb;102(2):484–491. doi: 10.1083/jcb.102.2.484. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. King G. L., Johnson S. M. Receptor-mediated transport of insulin across endothelial cells. Science. 1985 Mar 29;227(4694):1583–1586. doi: 10.1126/science.3883490. [DOI] [PubMed] [Google Scholar]
  13. Madri J. A., Williams S. K. Capillary endothelial cell cultures: phenotypic modulation by matrix components. J Cell Biol. 1983 Jul;97(1):153–165. doi: 10.1083/jcb.97.1.153. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Makino S., Reynolds J. A., Tanford C. The binding of deoxycholate and Triton X-100 to proteins. J Biol Chem. 1973 Jul 25;248(14):4926–4932. [PubMed] [Google Scholar]
  15. Muller W. A., Steinman R. M., Cohn Z. A. Intracellular iodination of lysosome membrane for studies of membrane composition and recycling. Methods Enzymol. 1983;98:404–415. doi: 10.1016/0076-6879(83)98168-5. [DOI] [PubMed] [Google Scholar]
  16. Muller W. A., Steinman R. M., Cohn Z. A. The membrane proteins of the vacuolar system I. Analysis of a novel method of intralysosomal iodination. J Cell Biol. 1980 Jul;86(1):292–303. doi: 10.1083/jcb.86.1.292. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Nakache M., Schreiber A. B., Gaub H., McConnell H. M. Heterogeneity of membrane phospholipid mobility in endothelial cells depends on cell substrate. Nature. 1985 Sep 5;317(6032):75–77. doi: 10.1038/317075a0. [DOI] [PubMed] [Google Scholar]
  18. Neville D. M., Jr Molecular weight determination of protein-dodecyl sulfate complexes by gel electrophoresis in a discontinuous buffer system. J Biol Chem. 1971 Oct 25;246(20):6328–6334. [PubMed] [Google Scholar]
  19. Rodriguez Boulan E., Sabatini D. D. Asymmetric budding of viruses in epithelial monlayers: a model system for study of epithelial polarity. Proc Natl Acad Sci U S A. 1978 Oct;75(10):5071–5075. doi: 10.1073/pnas.75.10.5071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rodriguez-Boulan E., Paskiet K. T., Sabatini D. D. Assembly of enveloped viruses in Madin-Darby canine kidney cells: polarized budding from single attached cells and from clusters of cells in suspension. J Cell Biol. 1983 Mar;96(3):866–874. doi: 10.1083/jcb.96.3.866. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Rosenberg R. D., Rosenberg J. S. Natural anticoagulant mechanisms. J Clin Invest. 1984 Jul;74(1):1–6. doi: 10.1172/JCI111389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Ryan U. S., Ryan J. W. Surface properties of pulmonary endothelial cells. Ann N Y Acad Sci. 1983;416:441–456. doi: 10.1111/j.1749-6632.1983.tb35204.x. [DOI] [PubMed] [Google Scholar]
  23. SALPETER M. M., BACHMANN L. AUTORADIOGRAPHY WITH THE ELECTRON MICROSCOPE. A PROCEDURE FOR IMPROVING RESOLUTION, SENSITIVITY, AND CONTRAST. J Cell Biol. 1964 Aug;22:469–477. doi: 10.1083/jcb.22.2.469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Simionescu M., Simionescu N., Palade G. E. Preferential distribution of anionic sites on the basement membrane and the abluminal aspect of the endothelium in fenestrated capillaries. J Cell Biol. 1982 Nov;95(2 Pt 1):425–434. doi: 10.1083/jcb.95.2.425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Simionescu M., Simionescu N., Santoro F., Palade G. E. Differentiated microdomains of the luminal plasmalemma of murine muscle capillaries: segmental variations in young and old animals. J Cell Biol. 1985 May;100(5):1396–1407. doi: 10.1083/jcb.100.5.1396. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Simionescu M., Simionescu N., Silbert J. E., Palade G. E. Differentiated microdomains on the luminal surface of the capillary endothelium. II. Partial characterization of their anionic sites. J Cell Biol. 1981 Sep;90(3):614–621. doi: 10.1083/jcb.90.3.614. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Simionescu N., Simionescu M., Palade G. E. Differentiated microdomains on the luminal surface of the capillary endothelium. I. Preferential distribution of anionic sites. J Cell Biol. 1981 Sep;90(3):605–613. doi: 10.1083/jcb.90.3.605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Simons K., Fuller S. D. Cell surface polarity in epithelia. Annu Rev Cell Biol. 1985;1:243–288. doi: 10.1146/annurev.cb.01.110185.001331. [DOI] [PubMed] [Google Scholar]
  30. Steinman R. M., Cohn Z. A. The interaction of soluble horseradish peroxidase with mouse peritoneal macrophages in vitro. J Cell Biol. 1972 Oct;55(1):186–204. doi: 10.1083/jcb.55.1.186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Steinman R. M., Mellman I. S., Muller W. A., Cohn Z. A. Endocytosis and the recycling of plasma membrane. J Cell Biol. 1983 Jan;96(1):1–27. doi: 10.1083/jcb.96.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Vasile E., Simionescu M., Simionescu N. Visualization of the binding, endocytosis, and transcytosis of low-density lipoprotein in the arterial endothelium in situ. J Cell Biol. 1983 Jun;96(6):1677–1689. doi: 10.1083/jcb.96.6.1677. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vlodavsky I., Johnson L. K., Gospodarowicz D. Appearance in confluent vascular endothelial cell monolayers of a specific cell surface protein (CSP-60) not detected in actively growing endothelial cells or in cell types growing in multiple layers. Proc Natl Acad Sci U S A. 1979 May;76(5):2306–2310. doi: 10.1073/pnas.76.5.2306. [DOI] [PMC free article] [PubMed] [Google Scholar]

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