Abstract
Diapedesis of leukocytes across endothelial barriers is generally believed to require the opening of endothelial tight junctions. At the blood-brain barrier (BBB), endothelial cells are interconnected by complex tight junctions. Here, we show by serial section conventional electron microscopy that during experimental autoimmune encephalomyelitis mononuclear cells traverse cerebral microvessels by a transcellular pathway, leaving the endothelial tight junctions intact. Cerebral endothelial cells were found to form filopodia-like membrane protrusions on their luminal aspect, thus embracing the mononuclear cells and forming cup-like structures, and eventually pores, through which the traversing cell could reach the abluminal side. At the abluminal side endothelial cell protrusions surrounding a migrating inflammatory cell were found to be progressively lined with basal lamina, suggesting a change from luminal to abluminal membrane characteristics of endothelial cell membranes during inflammatory cell diapedesis. Morphological evidence for the involvement of tight junctions in the diapedesis of mononuclear cells across the inflamed BBB could not be obtained in any case. Taken together, the presence of morphologically intact tight junctions and our novel finding of the presence of a basal lamina on both sides of abluminal endothelial cell protrusions surrounding migrating inflammatory cells suggests that during experimental autoimmune encephalomyelitis diapedesis of mononuclear cells occurs via a transendothelial process.
Similar content being viewed by others
References
Astrom KE (1968) Migration of lymphocytes through the endothelium of venules in experimental allergic neuritis. Experientia 24:589–590
Bobik R, Dabrowski Z (1995) Emperipolesis of marrow cells within megakaryocytes in the bone marrow of sublethally irradiated mice. Ann Hematol 70:91–95
Butcher EC (1991) Leukocyte-endothelial cell recognition: three (or more) steps to specificity and diversity. Cell 67:1033–1036
Butcher EC, Williams M, Youngman K, Rott L, Briskin M (1999) Lymphocyte trafficking and regional immunity. Adv Immunol 72:209–253
Carman CV, Jun C-D, Salas A, Springer TA (2003) Endothelial cells proactively form microvilli-like membrane projections upon intercellular adhesion molecule 1 engagement of leukocyte LFA-1. J Immunol 171:6135–6144
Claudio L, Raine CS, Brosnan C (1995) Evidence of persistent blood-brain barrier abnormalities in chronic-progressive multiple sclerosis. Acta Neuropathol 90:228–238
Corada M, Mariotti M, Thurston G, Smith K, Kunkel R, Brockhaus M, Lampugnani MG, Martin-Padura I, Stoppacciaro A, Ruco L, McDonald DM, Ward PA, Dejana E (1999) Vascular endothelial-cadherin is an important determinant of microvascular integrity in vivo. Proc Natl Acad Sci USA 96:9815–9820
Del Maschio A, De Luigi A, Martin-Padura I, Brockhaus M, Bartfai T, Fruscella P, Adorini L, Martino G, Furlan R, De Simoni MG, Dejana E (1999) Leukocyte recruitment in the cerebrospinal fluid of mice with experimental meningitis is inhibited by an antibody to junctional adhesion molecule (JAM). J Exp Med 190:1351–1356
Ebnet K, Vestweber D (1999) Molecular mechanisms that control leukocyte extravasation: the selectins and the chemokines. Histochem Cell Biol 112:1–23
Engelhardt B, Vestweber D, Hallmann R, Schulz M (1997) E- and P-selectin are not involved in the recruitment of inflammatory cells across the blood-brain barrier in experimental autoimmune encephalomyelitis. Blood 90:4459–4472
Engelhardt B, Laschinger M, Schulz M, Samulowitz U, Vestweber D, Hoch G (1998) The development of experimental autoimmune encephalomyelitis in the mouse requires alpha4-integrin but not alpha4beta7-integrin. J Clin Invest 102:2096–2105
Feng D, Nagy JA, Pyne K, Dvorak HF, Dvorak AM (1998) Neutrophils emigrate from venules by a transendothelial cell pathway in response to FMLP. J Exp Med 187:903–915
Feng D, Nagy JA, Dvorak HF, Dvorak AM (2002) Ultrastructural studies define soluble macromolecular, particulate, and cellular transendothelial cell pathways in venules, lymphatic vessels, and tumor-associated microvessels in man and animals. Microsc Res Tech 57:289–326
Gowans JL, Knight EJ (1964) The route of recirculation of lymphocytes in the rat. Proc R Soc Lond B Biol 159:257–282
Gotsch U, Borges E, Bosse R, Böggemeyer E, Simon M, Mossmann H, Vestweber D (1997) VE-cadherin antibody accelerates neutrophil recruitment in vivo. J Cell Sci 110:583–588
Greenwood J, Howes R, Lightman S (1994) The blood-retinal barrier in experimental autoimmune uveoretinitis-leukocyte interactions and functional damage. Lab Invest 70:39–52
Greenwood J, Wang Y, Calder VL (1995) Lymphocyte adhesion and transendothelial migration in the central nervous system: the role of LFA-1, ICAM-1, VLA-4 and VCAM-1. Immunology 86:408–415
Humble JG, Jayne WHW, Pulvertaft RJV (1956) Biological interaction between lymphocytes and other cells. Br J Haematol 2:283–294
Johnson-Léger C, Imhof BA (2003) Forging the endothelium during inflammation: pushing at a half-open door? Cell Tissue Res 314:93–105
Laschinger M, Vajkoczy P, Engelhardt B (2002) Encephalitogenic T cells use LFA-1 during transendothelial migration but not during capture and adhesion in spinal cord microvessels in vivo. Eur J Immunol 32:3598–3606
Lechner F, Sahrbacher U, Suter T, Frei K, Brockhaus M, Koedel U, Fontana A (2000) Antibodies to the junctional adhesion molecule cause disruption of endothelial cells and do not prevent leukocyte influx into the meninges after viral or bacterial infection. J Infect Dis 182:978–982
Lossinski AS, Badmajew V, Robson JA, Moretz RC, Wisniewski H, M (1989) Sites of egress of inflammatory cells and horseradish peroxidase transport across the blood-brain barrier in a murine model of chronic relapsing experimental allergic encephalomyelitis. Acta Neuropathol 78:359–371
Luscinskas FW, Ma S, Nusrat A, Parkos CA, Shaw SK (2002) Leukocyte transendothelial migration: a junctional affair. Semin Immunol 14:105–113
Lyck R, Reiss Y, Gerwin N, Greenwood J, Adamson P, Engelhardt B (2003) T cell interaction with ICAM-1/ICAM-2-double-deficient brain endothelium in vitro: the cytoplasmic tail of endothelial ICAM-1 is necessary for transendothelial migration of T cells. Blood 102:3675–3683
Marchesi VT, Gowans JL (1964) The migration of lymphocytes through the endothelium of venules in lymph nodes: an electron microscope study. Proc R Soc Lond B Biol Sci 159:283–290
Martin R, McFarland HF (1995) Immunological aspects of experimental allergic encephalomyelitis and multiple sclerosis. Crit Rev Clin Lab Sci 32:121–182
McMenamin PG, Forrester JV, Steptoe RJ, Dua HS (1992) Ultrastructural pathology of experimental autoimmune uveitis. Quantitative evidence of activation and possible high endothelial venule-like changes in retinal vascular endothelium. Lab Invest 67:42–55
Muller WA (2003) Leukocyte-endothelial-cell interactions in leukocyte transmigration and the inflammatory response. Trends Immunol 6:327–334
Muller WA, Weigl SA, Deng X, Phillips DM (1993) PECAM-1 is required for transendothelial migration of leukocytes. J Exp Med 178:449–460
Oppenheimer-Marks N, Davis LS, Bogue DT, Ramberg J, Lipsky PE (1991) Differential utilization of ICAM-1 and VCAM-1 during the adhesion and transendothelial migration of human T lymphocytes. J Immunol 147:2913–2921
Raine CS, Cannella B, Duijvestijn AM, Cross AH (1990) Homing to central nervous system vasculature by antigen-specific lymphocytes. II. Lymphocyte/endothelial cell adhesion during the initial stages of autoimmune demyelination. Lab Invest 63:476–489
Reiss Y, Hoch G, Deutsch U, Engelhardt B (1998) T cell interaction with ICAM-1-deficient endothelium in vitro: essential role for ICAM-1 and ICAM-2 in transendothelial migration of T cells. Eur J Immunol 28:3086–3099
Tavassoli M (1986) Modulation of megakaryocyte emperipolesis by phlebotomy: megakaryocytes as a component of marrow-blood barrier. Blood Cells 12:205–216
Vajkoczy P, Laschinger M, Engelhardt B (2001) Alpha4-integrin-VCAM-1 binding mediates G protein-independent capture of encephalitogenic T cell blasts to CNS white matter microvessels. J Clin Invest 108:557–565
Wakelin MW, Sanz MJ, Dewar A, Albelda SM, Larkin SW, Boughton-Smith N, Williams TJ, Nourshargh S (1996) An anti-platelet-endothelial cell adhesion molecule-1 antibody inhibits leukocyte extravasation from mesenteric microvessels in vivo by blocking the passage through the basement membrane. J Exp Med 184:229–239
Wekerle H, Engelhardt B, Risau W, Meyermann R (1990) Passage of lymphocytes across the blood-brain barrier in health and disease. In: Johansson BB, Owman C, Widner H (eds) Pathophysiology of the blood-brain barrier. Elsevier, Amsterdam, pp 439–445
Wekerle H, Engelhardt B, Risau W, Meyermann R (1991) Interaction of T lymphocytes with cerebral endothelial cells in vitro. Brain Pathol 1:107–114
Wolburg H, Wolburg-Buchholz K, Kraus J, Rascher-Eggstein G, Liebner S, Hamm S, Duffner F, Grote E-H, Risau W, Engelhardt B (2003) Localization of claudin-3 in tight junctions of the blood-brain barrier is selectively lost during experimental autoimmune encephalomyelitis and human glioblastoma multiforme. Acta Neuropathol 105:586–592
Wong D, Prameya R, Dorovini-Zis K (1999) In vitro adhesion and migration of T lymphocytes across monolayers of human brain microvessel endothelial cells: regulation by ICAM-1, VCAM-1, E-Selectin and PECAM-1. J Neuropathol Exp Neurol 58:138–152
Acknowledgements
We thank Dietmar Vestweber for continuous support and discussion of the manuscript.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Wolburg, H., Wolburg-Buchholz, K. & Engelhardt, B. Diapedesis of mononuclear cells across cerebral venules during experimental autoimmune encephalomyelitis leaves tight junctions intact. Acta Neuropathol 109, 181–190 (2005). https://doi.org/10.1007/s00401-004-0928-x
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00401-004-0928-x