Summary
Morphological heterogeneity has not been documented for cultured endothelial cells isolated from the microvascular bed of any organ. As the corpus luteum depends on a rich microvascularization, endothelial cells were dislodged from developing corpora lutea by mechanical dissection followed either by collagenase digestion or by no digestion. Cell separation was carried out by Percoll density centrifugation. Although the yield of intact cells was higher with collagenase treatment than without, successful endothelial cell cultures were only established when cells remained untreated. Viewed by light microscopy after an average lag phase of 10 days, five different phenotypes of endothelial cells were found under similar simple culture conditions: isomorphic epithelioid, polymorphic epithelioid, spindle-shaped, round, and phase-dense phenotypes. Monolayers appeared within 2–4 weeks. After an additional period of 2–4 weeks, tubular forms with a specific pattern were noted for types 1–3, the so-called pseudotubular forms for type 4, and none for type 5. Cell types differed in their cytochemical and immunocytochemical responses. Examined by SEM, type 1 displayed a more conspicuous surface anatomy than type 2. Types 3–5 demonstrated striking cell processes that were characteristic of each type. Tubular forms of types 1 and 2 showed cell borders and a marked increase in surface specializations, whereas tubular forms of type 3 lacked detectable cell borders in the absence of a striking surface anatomy. Pseudotubular forms of type 4 developed no particular spatial organization. Thus, for the first time, morphological evidence is provided that different endothelial cell types are obtained from diverse segments of the microvascular bed.
Similar content being viewed by others
References
Anders E, Alles J-U, Delvos U- Pötzsch B, Preissner KT, Müller-Berghaus G (1987) Microvascular endothelial cells from human omental tissue: modified method for long-term cultivation and new aspects of characterization. Microvas Res 34:239–249
Becker CG, Harpel PC (1976) 46-1 on human vascular endothelium. J Exp Med 144:1–9
Bensch KG, Davison PM, Karasek MA (1983) Factors controlling the in vitro growth pattern of human microvascular endothelial cells. J Ultrastruct Res 82:76–89
Björntorp P, Hansson GK, Jonasson L, Pettersson P, Sypniewska G (1983) Isolation and characterization of endothelial cells from the epididymal fat pad of the rat. J Lipid Res 24:105–112
Bottaro D, Shepro D, Hechtman HB (1986) Heterogeneity of intimal and microvessel endothelial cell barriers in vitro. Microvasc Res 32:389–398
Bowman PD, Betz AL, Ar D, Wolinsky JS, Penney JB, Shivers RR, Goldstein GW (1981) Primary culture of capillary endothelium from rat brain. In Vitro 17:353–362
Carson MP, Haudenschild CC (1986) Microvascular endothelium and pericytes: high yield, low passage cultures. In Vitro Cell Dev Biol 22:344–354
Chung-Welch N, Shepro D, Dunham B, Hechtman HB (1988) Prostacyclin and prostaglandin E2 secretions by bovine pulmonary microvessel endothelial cells are altered by changes in culture conditions. J Cell Physiol 135:224–234
Cotta-Pereira G, Sage H, Bornstein P, Ross R, Schwartz S (1980) Studies of morphologically atypical (“sprouting”) cultures of bovine aortic endothelial cells. Growth characteristics and connective tissue protein synthesis. J Cell Physiol 102:183–191
Davison PM, Karasek MA (1981) Human dermal microvascular endothelial cells in vitro: effect of cyclic AMP on cellular morphology and proliferation rate. J Cell Physiol 106:253–258
Diglio CA, Grammas P, Giacomelli F, Wiener J (1982) Primary culture of rat cerebral microvascular endothelial cells. Lab Invest 46:554–563
Duthu GS, Smith JR (1980) In vitro proliferation and lifespan of bovine aorta endothelial cells: effect of culture conditions and fibroblast growth factor. J Cell Physiol 103:385–392
Edanaga M (1974) A scanning electron microscope study on the endothelium of the vessels. I. Fine structure of the endothelial surface of aorta and some other arteries in normal rabbits. Arch Histol J 37:1–14
Feder J, Marasa JC, Olander JV (1983) The formation of capillarylike tubes by calf aortic endothelial cells grown in vitro. J Cell Physiol 116:1–6
Folkman J, Haudenschild CC (1980) Angiogenesis in vitro. Nature 288:551–556
Folkman J, Haudenschild CC, Zetter BR (1979) Long-term culture of capillary endothelial cells. Proc Natl Acad Sci USA 76:5217–5221
Fujita T, Tanaka K, Okunaga J (1981) SEM atlas of cells and tissues. Igaku-Shoin, Tokyo New York
Gerritsen ME (1987) Functional heterogenity of vascular endothelial cells. Biochem Pharmacol 36:2701–2711
Gitlin JD, D'Amore PA (1983) Culture of retinal capillary cells using selective growth media. Microvasc Res 26:74–80
Gospodarowicz D, Ill C (1980) Extracellular matrix and control of proliferation of vascular endothelial cells. J Clin Invest 65:1351–1364
Gospodarowicz D, Lui G-M (1981) Effect of substrata and fibroblast growth factor on the proliferation in vitro of bovine aortic endothelial cells. J Cell Physiol 109:69–81
Gospodarowicz D, Thakral KK (1978) Production of a corpus luteum angiogenic factor responsible for proliferation of capillaries and neovascularization of the corpus luteum. Proc Natl Acad Sci USA 75:847–851
Gospodarowicz D, Moran J, Braun D, Birdwell C (1976) Clonal growth of bovine vascular endothelial cells: fibroblast growth factor as a survival agent. Proc Natl Acad Sci USA 73:4120–4124
Gospodarowicz D, Massoglia S, Cheng J, Fujii DK (1986) Effect of fibroblast growth factor and lipoproteins on the proliferation on endothelial cells derived from bovine adrenal cortex, brain cortex, and corpus luteum capillaries. J Cell Physiol 127:121–136
Greenburg G, Vlodavsky I, Foidart JM, Gospodarowicz D (1980) Conditioned medium from endothelial cell cultures can restore the normal phenotypic expression of vascular endothelium maintained in vitro in the absence of fibroblast growth factor. J Cell Physiol 103:333–347
Höckel M, Sasse J, Wissler JH (1987) Purified monocyte-derived angiogenic substance (angiotropin) stimulates migration, phenotypic changes, and “tube formation” but not proliferation of capillary endothelial cells in vitro. J Cell Physiol 133:1–13
Höflinger H (1948) Das Ovar des Rindes in den verschiedenen Lebensperioden unter besonderer Berücksichtigung seiner Feinstruktur. Acta Anat Suppl 5
Ireland JJ, Murphee RL, Coulson PB (1980) Accuracy of predicting stages of bovine estrous cycle by gross appearance of the corpus luteum. J Dairy Sci 63:155–160
Jaffe EA, Nachman RL, Becker CG, Minick CR (1973) Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria. J Clin Invest 52:2745–2756
Johannisson E, Redard M (1984) Culture of human endothelial cells derived from capillaries of the decidual tissuc. Acta Obstet Gynecol Scand 63:27–36
Kanzaki H, Okamura H, Okuda Y, Takenaka A, Morimoto K, Nishimura T (1982) Scanning electron microscopy study of rabbit ovarian follicle microvasculature using resin injection corrosion casts. J Anat 134:697–704
Kern P, Knedler A, Eckel RH (1983) Isolation and culture of microvascular endothelium from human adipose tissue. J Clin Invest 71:1822–1829
Koos RD (1986) Stimulation of endothelial cell proliferation by rat granulosa cell-conditioned medium. Endocrinology 119:481–489
Kumar S, West DC, Ager A (1987) Heterogeneity in endothelial cells from large vessels and microvessels. Differentiation 36:57–70
Maciag T, Kadish J, Wilkins L, Stemerman MB, Weinstein R (1982) Organizational behavior of human umbilical vein endothelial cells. J Cell Biol 94:511–520
Madri JA, Williams SK, Wyatt T, Mezzio C (1983) Capillary endothelial cell cultures: phenotypic modulation by matrix components. J Cell Biol 97:153–165
Marynen P, Van der Schueren B, Van Leuven F, Cassiman JJ, Van den Berghe H (1982) Absence of specific binding of receptor-mediated endocytosis, and of secretion of alpha-2-macroglobulin by cultured endothelial cells. Haemostasis 11:210–214
McAuslan BR, Hannan GN, Reilly W, Stewart FHC (1980) Variant endothelial cells. Fibronectin as a transducer of signals for migration and neovascularisation. J Cell Physiol 104:177–186
McAuslan BR, Hannan GN, Reilly W (1982) Signals causing change in morphological phenotype, growth mode, and gene expression of vascular endothelial cells. J Cell Physiol 112:96–106
McGuire PG, Orkin RW (1987) Methods in laboratory investigation. Islation of rat aortic endothelial cells by primary explant techniques and their phenotypic modulation by defined substrata. Lab Invest 57:94–105
McIntosh LC, Muckersie L, Forrester JV (1988) Retinal capillary endothelial cells prefer different substrates for growth and migration. Tissue Cell 20:193–209
Moyer CF, Dennis PA, Majno G, Joris I (1988) Venular endothelium in vitro: isolation and chracterization. In Vitro Cell Dev Biol 24:359–368
Pitas RE, Boyles J, Mahley RW, Bissell DM (1985) Uptake of chemically modified low density lipoproteins in vivo is mediated by specific endothelial cells. J Cell Biol 100:103–117
Redmer DA, Rone JD, Goodman AL (1985) Evidence for a non-steroidal angiotropic factor from the primate corpus luteum: stimulation of endothelial cell migration in vitro. Proc Soc Exp Biol Med 179:136–140
Rupnick MA, Carey A, Williams SK (1988) Phenotypic diversity in cultured cerebral microvascular endothelial cells. In Vitro Cell Dev Biol 24:435–444
Ryan US, Clements E, Habliston D, Ryan JW (1978) Isolation and culture of pulmonary artery endothelial cells. Tissue Cell 10:535–554
Ryan US, Mortara M, Whitaker C (1980) Methods for microcarrier culture of bovine pulmonary artery endothelial cells avoiding the use of enzymes. Tissue Cell 12:619–635
Sato N, Sawasaki Y, Senoo A, Fuse Y, Hirano Y, Goto T (1987) Development of capillary networks from rat microvascular fragments in vitro: the role of myofibroblastic cells. Microvasc Res 33:194–210
Schindelmeiser J, Münstermann D, Witzel H (1987) Histochemical investigations on the localization of the purple acid phosphatase in the bovine spleen. Histochemistry 87:13–19
Schor AM, Schor SL, Allen TD (1983) Effects of culture conditions on the proliferation, morphology and migration of bovine aortic endothelial cells. J Cell Sci 62:267–285
Schwartz SM (1978) Selection and characterization of bovine aortic endothelial cells. In Vitro 14:966–980
Tilton RG, Kilo C, Williamson JR (1979) Pericyte-endothelial relationships in cardiac and skeletal muscle capillaries. Microvasc Res 18:325–335
Vlodavsky I, Johnson LK, Greenburg G, Gospodarowicz D (1979) Vascular endothelial cells maintained in the absence of fibroblast growth factor undergo structural and functional alterations that are incompatible with their in vivo differentiated properties. J Cell Biol 83:468–486
Zetter BR (1981) The endothelial cells of large and small blood vessels. Diabetes 30:24–28
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Spanel-Borowski, K., van der Bosch, J. Different phenotyes of cultured microvessel endothelial cells obtained from bovine corpus luteum. Cell Tissue Res 261, 35–47 (1990). https://doi.org/10.1007/BF00329436
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00329436