Summary
Heterotypic cell-cell interactions appear to be involved in the control of development and function in a wide variety of tissues. In the vasculature, endothelial cells and mural cells (smooth muscle cells or pericytes) make frequent contacts, suggesting a role for intercellular interactions in the regulation of vascular growth and function. We have previously grown endothelial cells and mural cells together in mixed cultures and found that heterocellular contact led to endothelial growth inhibition. However, this mixed culture system does not lend itself to the examination of the effects of contact on the phenotype of the individual cell types. We have therefore developed a co-culture system in which cells can be co-cultured across a porous membrane, permitting intercellular contact while maintaining pure cell populations. Co-culture of endothelial cells and smooth muscle cells across membranes with pore sizes of 0.02, 0.4, 0.6, and 0.8µm maintained the two cell types as homogeneous populations, whereas smooth muscle cells migrated across the membrane through pores of 2.0µm. Vascular cell co-culture across membranes with 0.8-µm pores resulted the inhibition of endothelial cell proliferation and the generation of conditioned media which inhibited endothelial cell growth. The arrangement of the cells in this co-culture system mimics thein vivo orientation of vascular cells in which mural cells are separated from the abluminal surface of the endothelium by a fenestrated internal elastic lamina or basement membrane. Because this co-culture system maintains separable populations of cells in contact or close proximity allowing for biochemical and molecular analyses of pure populations, it should prove useful for the study of cell-cell interactions in a variety of systems.
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References
Albelda, S. M.; Sampson, P. M.; Haselton, F. R., et al. Permeability characteristics of cultured endothelial monolayers. J. Appl. Physiol. 64:308–322; 1988.
Antonelli-Orlidge, A.; Saunders, K. B.; Smith, S. R., et al. An activated form of TGF-β is produced by co-cultures of endothelial cells and pericytes. Proc. Natl. Acad. Sci. USA 86:4544–4548; 1989.
Arthur, F. E.; Shivers, R. R.; Bowman, P. D. Astrocyte-mediated induction of tight junctions in brain capillary endothelium: an efficientin vitro model. Dev. Brain Res. 36:155–159; 1987.
Barrett, T. B.; Gajdusek, C. M.; Schwartz, S. M., et al. Expression of thesis gene by endothelial cells in culture andin vivo. Proc. Natl. Acad. Sci. USA 81:6772–6774; 1984.
Beck, D. W.; Roberts, R. L.; Olson, J. J. Glial cells influence membrane-associated enzyme activity at the blood-brain barrier. Brain Res. 381:131–137; 1986.
Bently, D.; Caudy, M. Pioneer axons lose directed growth after selective killing of guidepost cells. Nature 304:62–65; 1983.
Bruns, R. R.; Palade, G. E. Studies on blood capillaries. I. General organization of blood capillaries in muscle. J. Cell Biol. 37:244–276; 1968.
Carlson, E. C. Fenestrated subendothelial basement membranes in human retinal capillaries. Invest. Ophthalmol. Vis. Sci. 30:1923–1932; 1989.
Chamley, J. H.; Groschel-Stewart, U.; Campbell, G. R., et al. Distinctions between smooth muscle, fibroblasts and endothelial cells in culture by the use of fluoresceinated antibodies against smooth muscle actin. Cell Tissue Res. 177:445–457; 1977.
Crocker, D. J.; Murad, T. M.; Greer, J. C. Role of the pericyte in wound healing. An ultrastructural study. Exp. Mol. Pathol. 13:51–65; 1970.
Davies, P. F.; Truskey, G. A.; Warren, H. B., et al. Metabolic cooperation between vascular endothelial and smooth muscle cells in co-culture: changes in low density lipoprotein metabolism. J. Cell Biol. 101:871–879; 1985.
DeBault, L. E.; Cancilla, P. A.γ-GTP in isolated brain endothelial cells: induction by glial cellsin vitro. Science 207:653–654; 1980.
Dehouck, M. P.; Meresse, S.; Delorme, P., et al. An easier, reproducible, and mass-production method to study the blood-brain barrierin vitro. J. Neurochem. 54:1798–1801; 1990.
Dunmore, P. J.; Song, S. H.; Roach, M. R. A comparison of the size of fenestrations in the internal elastic lamina of young and old porcine aortas as seen with the scanning electron microscope. Can. J. Physiol. Pharmacol. 68:139–143; 1990.
Folkman, J.; Haudenschild, C.; Zetter, B. R. Long-term culture of capillary endothelial cells. Proc. Natl. Acad. Sci. USA 76:5217–5221; 1979.
Gabriels, J. E.; D’Amore, P. A. Smooth muscle cells (SMC) respond chemotactically to vascular endothelial cells (EC) in an in vitro under agarose migration assay. J. Cell Biol. 115:404a; 1991.
Gimbrone, M. A., Jr. Culture of vascular endothelium. Prog. Hemostasis Thromb. 3:1–28; 1976.
Gimbrone, M. A., Jr. Cotran, R. S.; Folkman, J. Human vascular endothelial cells in culture: growth and DNA synthesis. J. Cell Biol. 60:673–684; 1974.
Gitlin, J. D.; D’Amore, P. A. Culture of retinal capillary cells using selective growth media. Microvasc. Res. 26:74–80; 1983.
Grobstein, C. Trans-filter induction of tubules in mouse metanephrogenic mesenchyme. Exp. Cell Res. 10:424–440; 1956.
Guguen-Guillouzo, C.; Clément, B.; Baffet, C., et al. Maintenance and reversibility of active albumin secretion by adult rat hepatocytes co-cultured with another liver epithelial cell type. Exp. Cell Res. 143:47–54; 1983.
Hansson, G. K.; Schwartz, S. M. Evidence for cell death in the vascular endotheliumin vivo andin vitro. Am. J. Pathol. 112:278–286; 1983.
Haudenschild, C. C.; Dahniser, D.; Folkman, J., et al. Human vascular endothelial cells in culture: lack of response to serum growth factors. Exp. Cell Res. 98:175–182; 1976.
Hisanaga, K.; Sharp, F. R. Marked neurotrophic effects of diffusible substances released from non-target cerebellar cells on thalamic neurons in culture. Dev. Brain Res. 54:151–160; 1990.
Janzer, R. C.; Raff, M. C. Astrocytes induce blood-brain barrier properties in endothelial cells. Science 325:253–257; 1987.
Kédinger, M.; Simon-Assmann, P.; Alexandre, E., et al. Importance of a fibroblastic support forin vitro differentiation of intestinal endodermal cells and for their response to glucocorticoids. Cell Differ. 20:171–182; 1987.
Kierney, P. C.; Dorshkind, K. B lymphocyte precursors and myeloid progenitors survive in diffusion chamber cultures but B cell differentiation requires close association with stromal cells. Blood 70:1418–1424; 1987.
Krueger, G. G.; Jorgensen, C. M.; Bradshaw, B. R., et al. Approach for and assessment of interactive communication via cytokines of cellular components of skin. Dermatologica 179:91S-100S; 1989.
Laterra, J.; Guerin, C.; Goldstein, G. W. Astrocytes induce neural microvascular endothelial cells to form capillary-like structuresin vitro. J. Cell Physiol. 144:204–215; 1990.
Lehtonen, E.; Wartiovaara, J.; Nordling, S., et al. Demonstration of cytoplasmic processes in Millipore filters permitting kidney tubule induction. J. Embryol. Exp. Morphol. 33:187–203; 1975.
Lillien, L.; Raff, M. Analysis of the cell-cell interactions that control type-2 astrocyte developmentin vitro. Neuron 4:525–534; 1990.
Madri, J.; Williams, S. K. Capillary endothelial cell cultures: phenotypic modulation by matrix components. J. Cell Biol. 97:153–165; 1983.
Mazanet, R.; Franzini-Armstrong, C. Scanning electron microscopy of pericytes in rat red muscle. Microvasc. Res. 23:361–369; 1982.
Mehta, R. P.; Bertram, J. S.; Loewenstein, W. R. Growth inhibition of transformed cells correlates with their junctional communication with normal cells. Cell 44:187–196; 1986.
Melchers, F.; Andersson, J. Factors controlling the B cell cycle. Annu. Rev. Immunol. 4:13–36; 1986.
Milici, A. J.; Furie, M. B.; Carley, W. W. The formation of fenestrations and channels by capillary endotheliumin vitro. Proc. Natl. Acad. Sci. USA 82:6181–6185; 1985.
Munro, J. M.; Cotran, R. S. The pathogenesis of atherosclerosis: atherogenesis and inflammation. Lab. Invest. 58:249–261; 1988.
Orlidge, A.; D’Amore, P. A. Inhibition of capillary endothelial cell growth by pericytes and smooth muscle cells. J. Cell Biol. 105:1455–1462; 1987.
Rakic, P. Contact regulation of neuronal migration. In: Edelman, G. M.; Thiery, J. P., eds. The cell in contact. New York: Wiley Intersciences; 1985:67–91.
Rhodin, J. Ultrastructure of mammalian venous capillaries, venules, and small collecting veins. J. Ultrastruct. Res. 25:425–500; 1968.
Robison, W. G., Jr.; Magata, M.; Tillis, T. N., et al. Aldose reductase and pericyte-endothelial cell contacts in retina and optic nerve. Invest. Ophthalmol. Vis. Sci. 30:2293–2299; 1989.
Ross, R. The smooth muscle cell. II. Growth of smooth muscle in culture and formation of elastic fibers. J. Cell Biol. 50:172–186; 1971.
Sargent, T. D.; Jamrich, M.; Dawid, I. Cell interactions and the control of gene activity during early development ofXenopus laevis. Dev. Biol. 114:238–246; 1986.
Sato, Y.; Rifkin, D. B. Inhibition of endothelial cell movement by pericytes and smooth muscle cells: activation of a latent transforming growth factor-beta 1-like molecule by plasmin during co-culture. J. Cell Biol. 109:309–315; 1989.
Sheridan, J. D.; Larson, D. M. Junctional communication in the peripheral vasculature. In: Pitts, J. D.; Finbow, M. E., ed. The functional integration of cells in animal tissues. British society for cell biology symposium series, vol. 5. England; Cambridge University Press; 1982:263–283.
Shimaoka, S.; Nakamura, T.; Ichihara, A. Stimulation of growth of primary cultured adult rat hepatocytes without growth factors by coculture with nonparenchymal liver cells. Exp. Cell Res. 172:228–242; 1987.
Shivers, R. R.; Arthur, F. E.; Bowman, P. D. Induction of gap junctions and brain endothelium-like tight junctions in cultured bovine endothelial cells: local control of cell specialization. J. Submicrosc. Cytol. Pathol. 20:1–14; 1988.
Spagnoli, L.; Villaschi, S.; Neri, L., et al. Gap junctions in myoendothelial bridges of rabbit carotid arteries. Experientia 38:124–125; 1980.
Spitznas, M.; Reale, E. Fracture faces of fenestrations and junctions of endothelial cells in human choroidal vessels. Invest. Ophthalmol. Vis. Sci. 14:98–107; 1975.
Sweet, E.; Abraham, E. H.; D’Amore, P. A. Functional evidence of gap junctions between capillary endothelial cells and pericytesin vitro. Invest. Ophthalmol. Vis. Sci. 29:109a; 1988.
Ueda, H.; Tres, L. L.; Kierszenbaum, A. L. Culture patterns and sorting of rat Sertoli cell secretory proteins. J. Cell Sci. 89:175–188; 1988.
Wagenvoort, C. A.; Dingemans, K. P. Pulmonary vascular smooth muscle and its interaction with endothelium. Chest 88:200S-202S; 1985.
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Saunders, K.B., D’Amore, P.A. An in vitro model for cell-cell interactions. In Vitro Cell Dev Biol - Animal 28, 521–528 (1992). https://doi.org/10.1007/BF02634136
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DOI: https://doi.org/10.1007/BF02634136