Growth of brain microvessel endothelial cells on collagen gels: Applications to the study of blood-brain barrier physiology and CNS inflammation
Received: 19 October 1993 Accepted: 06 January 1994 DOI:
10.1007/BF02631256 Cite this article as: Biegel, D. & Pachter, J.S. In Vitro Cell Dev Biol - Animal (1994) 30: 581. doi:10.1007/BF02631256 Summary
Brain microvessel endothelial cells (BMEC) exhibit the tendency to migrate through 3.0-
vm pore semipermeable inserts and establish monolayers on both apical and basal filter surfaces. This can potentially lead to complications in accurately assessing a wide variety of physiologic parameters uniquely associated with these cells. To avoid this problem, we have explored growing BMEC on Transwell filters coated with hydrated collagen gels. BMEC seeded on such gels grow as a monolayer until confluency, but do not invade the subendothelial collagen matrix or the underlying support filter. Furthermore, BMEC grown in this manner exhibit biochemical, morphologic, and electrophysiologic properties reflective of the endothelial cells that comprise the blood-brain barrier in vivo. Although the collagen gel acts as an impenetrable barrier to BMEC, and thus ensures the growth of only a single layer of cells, it nevertheless can be infiltrated by monocytes that have been stimulated by a chemotaxin to undergo diapedesis. Thus, growing BMEC on collagen gel-coated Transwells has broad applications for the in vitro study of both blood-brain barrier physiology as well as the mechanisms underlying central nervous system inflammation. Key words endothelial cells brain microvessels blood-brain barrier References
Audus, K. L.; Borchardt, R. T. Characterization of an in vitro blood-brain barrier model system for studying drug transport and metabolism. Pharmacol. Res. 3:81–87; 1986.
Biegel, D.; Pachter, J. S. Attachment of mRNA to the cytoskeletal framework likely represents a physiological binding event. J. Cell. Biochem. 48:98–106; 1992.
Clayson, E. T.; Compans, R. W. Entry of simian virus 40 is restricted to apical surfaces of polarized epithelial cells. Mol. Cell. Biol. 8:3391–3396; 1988.
Crone, C.; Olesen, S. P. Electrical resistance of the brain microvascular endothelium. Brain Res. 241:49–55; 1981.
Finlay, B. B.; Fry, J.; Rock, E. P., et al. Passage of Salmonella through polarized epithelial cells. Role of the host bacterium. J. Cell Sci. Suppl. 11:99–108; 1989.
Hakkert, B. C.; Rentenaar, J. M.; Van Aken, W. G., et al. A three-dimensional model system to study the interactions between human leukocytes and endothelial cells. Eur. J. Immunol. 20:2775–2781; 1990.
Harris, P.; Ralph, P. Human leukemic models of myelomonocytic development: a review of the HL-60 and U937 cell lines. J. Leukocyte Biol. 37:407–418; 1985.
Hoekstra, D.; Kok, J. W. Entry mechanism of enveloped viruses. Implications for fusion of intracellular membranes. Biosci. Rev. 9:273–305; 1989.
Huber, A.; Ellis, S.; Johnson, K. J., et al. Monocyte diapedesis through an in vitro vessel wall construct: inhibition with monoclonal antibodies to thrombospondin. J. Leukocyte Biol. 52:524–528; 1992.
Joo, F. The cerebral microvessels in culture. J. Neurochem. 58:1–17; 1992.
Miller, D. W.; Audus, K. L.; Borchardt, R. T. Application of cultured endothelial cells of the brain microvasculature in the study of the blood-brain barrier. J. Tissue Cult. Methods 14:217–224; 1992.
Muller, W. A.; Weigl, S. A. Monocyte-selective transendothelial migration: dissection of the binding and transmigration phases by an in vitro assay. J. Exp. Med. 176:819–827; 1992.
Morzycki, W.; Sadowska, J.; Issekutz, A. C. Interleukin-1 and tumour necrosis factor
induced polymorphonuclear leukocyte-endothelial cell adhesion and transendothelial migration in vitro: the effect of apical versus basal monolayer stimulation. Immunol. Lett. 25:331–340; 1990.
Patrick, D.; Betts, J.; Frey, E. A., et al. Haemophilus influenza lipopolysaccharide disrupts confluent monolayers of bovine brain endothelial cells via a serum-dependent cytotoxic pathway. J. Infect. Dis. 165:865–872; 1992.
Pardridge, W. M.; Nowlin, D. M.; Choi, T. B., et al. Brain capillary 46,000 dalton protein is cytoplasmic actin and is localized to endothelial plasma membrane. J. Cereb. Blood Flow Metab. 9:675–680; 1989.
Pawlowski, N. A.; Kaplan, G.; Abraham, E., et al. The selective binding and transmigration of monocytes through the junctional complexes of human endothelium. J. Exp. Med. 168:1865–1882; 1988.
Reese, T. S.; Karnovsky, M. J. Fine structural localization of a blood-brain barrier to exogenous peroxidase. J. Cell Biol. 34:207–217; 1967.
Rodriguez, D.; Rodriguez, J.-R.; Pjakian, G. K., et al. Vaccinia virus preferentially enters polarized epithelial cells through the basalateral surface. J. Virol. 65:494–498; 1991.
Rubin, L. L.; Hall, D. E.; Porter, S., et al. A cell culture model of the blood-brain barrier. J. Cell Biol. 115:1725; 1991.
Sahagun, G.; Moore, S. A.; Fabry, Z., et al. Purification of murine endothelial cell cultures by flow cytometry using fluorescein-labeled
agglutinin. Am. J. Pathol. 134:1227–1232; 1989.
Shasby, S. S. Endothelial cells grown on permeable membrane supports. J. Tissue Cult. Methods 14:247–252; 1992.
Sundstrum, C.; Nilsson, K. Establishment and characterization of a human histiocytic lymphoma cell line (U937). Int. J. Cancer 17:565–577; 1976.
Takakura, Y.; Audus, K. L.; Borchardt, R. T. Blood-brain barrier: transport studies in isolated brain capillaries and in cultured brain endothelial cells. Adv. Pharmacol. 22:137–165; 1991.
Tucker, S. P.; Melson, L. R.; Compans, R. W. Migration of polarized epithelial cells through permeable membrane substrates of defined pore size. Eur. J. Cell Biol. 58:280–290; 1992.
Virji, M.; Kayhty, H.; Fergusen, D. J. P., et al. Interactions of
with cultured human endothelial cells. Microb. Pathog. 10:231–245; 1991.
Vorbrodt, A. W.; Lossinsky, A. S.; Wisniewski, H. M. Ultrastructural studies of conconavalin A receptors and 5′ nucleotidase localization in normal and injured mouse cerebral microvasculature. Acta Neuropathol. 63:210–217; 1984.
Wyrick, P. B.; Choong, J.; Davis, C. H., et al. Infect. Immun. 57:2378–2389; 1989.
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