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Regular Article| Volume 67, ISSUE 2, P169-178, February 01, 1997

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Coculture of Endothelial Cells and Smooth Muscle Cells in Bilayer and Conditioned Media Models

DOI:https://doi.org/10.1006/jsre.1996.4978
Coculture of Endothelial Cells and Smooth Muscle Cells in Bilayer and Conditioned Media Models
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      Abstract

      Smooth muscle cell (SMC) growth characteristics are affected by endothelial cells (ECs)in vivoandin vitro.In this study, we compare a bilayer EC/SMC coculture model that allows cell contact with a model of SMCs growing in media continuously conditioned by ECs, but without physical contact. Bovine aortic SMCs were plated on one side of a 13-μm-thick, semipermeable membrane. Three models were compared: (1) SMCs cultured alone (with no cells on the opposite side of the membrane, O/SMC); (2) SMCs cultured with ECs on the opposite side of the membrane in a bilayer coculture system that allows physical contact between ECs and SMCs (EC/SMC); and (3) SMCs cultured in media continuously conditioned by adjacent ECs, without contact (conditioned media, CM). After cultures were established, SMCs were harvested at 7 and 14 days after plating (n= 5 cultures/day/group). SMC DNA and protein content and [3H]thymidine incorporation were measured in each group. On Days 7 and 14 after plating, ECs in both the EC/SMC and CM models stimulated SMC proliferation >50% compared to O/SMC controls (P< 0.05). SMC density was similar for the EC/SMC and CM models at Day 7, but SMC density was higher in the EC/SMC group at Day 14 in culture (P< 0.05). At Day 7, protein synthesis was similar in the three groups, but by Day 14, SMCs in the EC/SMC group had produced significantly less cellular protein/DNA than SMCs in the CM group (P< 0.05), which in turn had less protein/DNA than the control (O/SMC) group (P< 0.05). SMCs in the EC/SMC and CM groups retained a thin, spindle shape with filamentous projections, compared to the hypertrophic appearance of SMCs in the absence of ECs. Electron microscopy revealed projections from SMCs which traversed the pores in the coculture membrane and made intimate contact with ECs. The degree of EC/SMC contact increased from 7 to 14 days (P< 0.05). Compared to SMCs alone, ECs in bilayer coculture or conditioned media altered SMCs growth characteristics similarly after 7 days in culture. By 14 days, however, the bilayer coculture had a significantly greater effect on SMC density and protein synthesis. The bilayer model is unique in terms of luminal/abluminal orientation of the cells, the proximity of the cell layers, and the presence of physical cell contact. Since the bilayer model amplifies the effect of ECs on SMCs, it may be more useful than conditioned media to study EC–SMC interactions.
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      References

      REFERENCES

        • Davies P.F.
        Biology of disease: Vascular cell interactions with special reference to the pathogenesis of atherosclerosis.
        Lab. Invest. 1986; 55: 5
        • Zarins C.K.
        • Zatina M.A.
        • Giddens D.P.
        • Ku D.N.
        • Glagov S.
        Shear stress regulation of artery lumen diameter in experimental atherogenesis.
        J. Vasc. Surg. 1987; 5: 413
        • Kamiya A.
        • Togawa T.
        Adaptive regulation of wall shear stress to flow change in the canine carotid artery.
        Am. J. Physiol. 1980; 239
        • Langille B.L.
        • O'Donnell F.
        Reductions in arterial diameter produced by chronic decreases in blood flow are endothelium-dependent.
        Science. 1986; 231: 405
        • Melkumyants A.M.
        • Balashov S.A.
        • Khayutin V.M.
        Endothelium dependent control of arterial diameter by blood viscosity.
        Cardiovasc. Res. 1989; 23: 741
        • Davies M.G.
        • Klyachkin M.L.
        • Dalen H.
        • Svendsen E.
        • Hagen P.
        Regression of intimal hyperplasia with restoration of endothelium-dependent relaxing factor-mediated relaxation in experimental vein grafts.
        Surgery. 1993; 114: 258
        • Chamley-Campbell J.H.
        • Campbell G.R.
        What controls smooth muscle phenotype?.
        Atherosclerosis. 1981; 40: 347
        • Campbell G.R.
        • Campbell J.H.
        Phenotypic modulation of smooth muscle cells in primary culture.
        Vascular Smooth Muscle in Culture. CRC Press, Boca Raton1987 (p. 39–)
        • Spagnoli L.G.
        • Villaschi S.
        • Neri L.
        • Palmieri G.
        Gap junctions in myo-endothelial bridges of rabbit carotid arteries.
        Experientia. 1982; 38: 124
        • Zerwes H.-G.
        • Risau W.
        Polarized secretion of a platelet-derived growth factor-like chemotactic factor by endothelial cells in vitro.
        J. Cell Biol. 1987; 105: 2037
        • Muller W.A.
        • Gimbrone Jr, M.A.
        Plasmalemmal proteins of cultured vascular endothelial cells exhibit apical–basal polarity: Analysis by surface-selective iodination.
        J. Cell Biol. 1986; 103: 2389
        • Roberts R.L.
        • Sandra A.
        Apical–basal membrane polarity of membrane phosphatases in isolated capillary endothelium: Alteration in ultrastructural localisation under culture conditions.
        J. Anat. 1993; 182: 339
        • Stins M.F.
        • Maxfield F.R.
        • Goldberg I.J.
        Polarized binding of lipoprotein lipase to endothelial cells: Implications for its physiological actions.
        Arterioscler. Thromb. 1992; 12: 1437
        • Wagner O.F.
        • Christ G.
        • Wojta J.
        • Vierhapper H.
        • Parzer S.
        • Nowotny P.J.
        • Schneider B.
        • Waldhausl W.
        • Binder B.R.
        Polar secretion of endothelin-1 by cultured endothelial cells.
        J. Biol. Chem. 1992; 267: 16066
        • Fillinger M.F.
        • O'Connor S.E.
        • Wagner R.J.
        • Cronenwett J.L.
        The effect of endothelial cell coculture on smooth muscle cell proliferation.
        J. Vasc. Surg. 1993; 17: 1058
        • Graham D.J.
        • Alexander J.J.
        • Miguel R.
        Aortic endothelial and smooth muscle cell co-culture: An in vitro model of the arterial wall.
        J. Invest. Surg. 1991; 4: 487
        • Gajdusek C.
        • DiCorleto P.E.
        • Ross R.
        • Schwartz S.M.
        An endothelial cell-derived growth factor.
        J. Cell. Biol. 1980; 85: 467
        • Davies P.F.
        • Kerr C.Co-
        cultivation of vascular endothelium and smooth muscle cells using microcarrier techniques.
        Exp. Cell Res. 1982; 141: 455
        • Castellot J.
        • Addonizio M.L.
        • Rosenberg R.
        • Karovsky M.J.
        Cultured endothelial cells produce a heparin like inhibitor of smooth muscle cell growth.
        J. Cell. Biol. 1981; 90: 372
        • Van Buul-Wortelboer M.F.
        • Brinkman H.J.M.
        • Dingeman K.P.
        • DeGroot P.G.
        • van Aken W.G.
        • van Mourik J.A.
        Reconstruction of the vascular wall in vitro: A novel model to study interaction between endothelial and smooth muscle cells.
        Exp. Cell Res. 1986; 162: 151
        • Powell R.J.
        • Cronenwett J.L.
        • Fillinger M.F.
        • Wagner R.J.
        Effect of endothelial cells and TGF-β1.
        J. Vasc. Surg. 1994; 20: 787
        • Smith P.K.
        • Krohn R.I.
        • Hermanson G.T.
        • Mallia A.K.
        • Gartner F.H.
        • Provenzano M.D.
        • Fujimoto E.K.
        Measurement of protein using bicinchoninic acid.
        Anal. Biochem. 1985; 150: 76
        • Cesarone C.F.
        • Bolognesi C.
        • Santi L.
        Improved microfluorometric DNA determination in biological material using 33258 Hoechst.
        Anal. Biochem. 1979; 100: 188
        • Brunk C.F.
        • Jones K.C.
        • James T.W.
        Assay for nanogram quantities of DNA in cellular homogenates.
        Anal. Biochem. 1979; 92: 497
        • Merrilees M.J.
        • Scott L.
        Interaction of aortic endothelial and smooth muscle cells in culture: Effect on glycosaminoglycan levels.
        Atherosclerosis. 1981; 39: 147
        • Ganz P.
        • Davies P.F.
        • Leopold J.A.
        • Gimbrone Jr., M.A.
        • Alexander R.W.
        Short- and long-term interactions of endothelium and vascular smooth muscle in co-culture: Effects on cyclic GMP production.
        Proc. Natl. Acad. Sci. USA. 1986; 83: 3552
        • Koo E.W.Y.
        • Gotlieb A.I.
        Endothelial stimulation of intimal cell proliferation in a porcine aortic organ culture.
        J. Pathol. 1989; 134: 497
        • Allen K.E.
        • Varty K.
        • Jones L.
        • Sayers R.D.
        • Bell P.R.F.
        • London N.J.M.
        Human venous endothelium can promote intimal hyperplasia in a paracrine manner.
        J. Vasc. Surg. 1994; 19: 577
        • Schwartz S.M.
        • Reidy M.A.
        Common mechanisms of proliferation of smooth muscle in atherosclerosis and hypertension.
        Hum. Pathol. 1987; 18: 240
        • Clowes A.W.
        • Reidy M.A.
        Mechanisms of arterial graft failure: the role of cellular proliferation.
        Ann. NY Acad. Sci. 1987; 516: 673
        • Hofstra L.
        • Tordoir J.
        • Kitslaar P.
        • Hoeks A.
        • Daemen M.
        Enhanced cellular proliferation in intact stenotic lesions derived from human arteriovenous fistulas and peripheral bypass grafts.
        Circulation. 1996; 94: 1283
        • Saunders K.B.
        • D'Amore P.A.
        An in vitro model for cell–cell interactions.
        In Vitro Cell Dev. Biol. 1992; 28A: 521
        • Jones P.A.
        Construction of an artificial blood vessel wall from cultured endothelial and smooth muscle cells.
        Proc. Natl. Acad. Sci. USA. 1979; 76: 1882
        • Fallier-Becker P.
        • Betz E.
        • Wolburg-Buchholz K.
        • Fotev Z.
        Fibromuscular proliferates induced in vitro using a trans-filter culture system.
        Res. Exp. Med. 1991; 191: 11
        • Hajjar D.P.
        • Marcus A.J.
        • Hajjar K.A.
        Interactions of arterial cells: studies in the mechanisms of endothelial cell modulation of cholesterol metabolism in co-cultured smooth muscle cells.
        J. Biol. Chem. 1987; 262: 6976

      Article info

      Footnotes

      ☆J. H. CampbellG. R. Campbell, Eds.

      Identification

      DOI: https://doi.org/10.1006/jsre.1996.4978

      Copyright

      © 1997 Academic Press. Published by Elsevier Inc. All rights reserved.

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