Abstract
In vitro models mimicking capillary sprouting are important tools to investigate the tumor angiogenesis, developmental blood vessel formation, and pathophysiological remodeling processes of the capillary system in the adult. With this focus, in 1998 Korff et al. introduced endothelial cell (EC) spheroids as a three-dimensional in vitro model resembling angiogenic responses and sprouting behavior [1]. As such, EC spheroids are capable of giving rise to capillary-like sprouts which are relatively close to the physiologically and genetically programmed arrangement of endothelial cells in vessels. Co-culture spheroids consisting of endothelial cells and smooth muscle cells form a spheroidal core composed of smooth muscle cells and an outer monolayer of endothelial cells, similar to the physiological architecture of larger blood vessels. In practise, a defined number of endothelial cells are cultured in a round-bottom well plate or in “hanging drops” to allow the formation and arrangement of the spheroidal three-dimensional structure. Subsequently, they are harvested and embedded in a collagen gel to allow outgrowth of endothelial cell sprouts originating from each spheroid. To evaluate the pro- or antiangiogenic impact of a cytokine or compound, the number and length of sprouts is determined.
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References
Korff T, Augustin HG (1998) Integration of endothelial cells in multicellular spheroids prevents apoptosis and induces differentiation. J Cell Biol 143(5):1341–1352
Garlanda C, Dejana E (1997) Heterogeneity of endothelial cells. Specific markers. Arterioscler Thromb Vasc Biol 17(7):1193–1202
Wolburg H, Neuhaus J, Kniesel U et al (1994) Modulation of tight junction structure in blood-brain barrier endothelial cells. Effects of tissue culture, second messengers and cocultured astrocytes. J Cell Sci 107(Pt 5):1347–1357
Krause DS, Fackler MJ, Civin CI et al (1996) CD34: structure, biology, and clinical utility. Blood 87(1):1–13
Delia D, Lampugnani MG, Resnati M et al (1993) CD34 expression is regulated reciprocally with adhesion molecules in vascular endothelial cells in vitro. Blood 81(4):1001–1008
DeLisser HM, Newman PJ, Albelda SM (1994) Molecular and functional aspects of PECAM-1/CD31. Immunol Today 15(10):490–495
Baldwin HS, Shen HM, Yan HC et al (1994) Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31): alternatively spliced, functionally distinct isoforms expressed during mammalian cardiovascular development. Development 120(9):2539–2553
Korff T, Augustin HG (1999) Tensional forces in fibrillar extracellular matrices control directional capillary sprouting. J Cell Sci 112(Pt 19):3249–3258
Jakobsson L, Franco CA, Bentley K et al (2010) Endothelial cells dynamically compete for the tip cell position during angiogenic sprouting. Nat Cell Biol 12(10):943–953
Siemerink MJ, Klaassen I, Van Noorden CJ et al (2013) Endothelial tip cells in ocular angiogenesis: potential target for anti-angiogenesis therapy. J Histochem Cytochem 61(2):101–115
Siemerink MJ, Klaassen I, Vogels IM et al (2012) CD34 marks angiogenic tip cells in human vascular endothelial cell cultures. Angiogenesis 15(1):151–163
Laib AM, Bartol A, Alajati A et al (2009) Spheroid-based human endothelial cell microvessel formation in vivo. Nat Protoc 4(8):1202–1215. doi:10.1038/nprot.2009.96
Korff T, Kimmina S, Martiny-Baron G et al (2001) Blood vessel maturation in a 3-dimensional spheroidal coculture model: direct contact with smooth muscle cells regulates endothelial cell quiescence and abrogates VEGF responsiveness. FASEB J 15(2):447–457
Maisonpierre PC, Suri C, Jones PF et al (1997) Angiopoietin-2, a natural antagonist for Tie2 that disrupts in vivo angiogenesis. Science 277(5322):55–60
Hoeben A, Landuyt B, Highley MS et al (2004) Vascular endothelial growth factor and angiogenesis. Pharmacol Rev 56(4):549–580
Flamme I, Frolich T, Risau W (1997) Molecular mechanisms of vasculogenesis and embryonic angiogenesis. J Cell Physiol 173(2):206–210
Presta M, Dell’Era P, Mitola S et al (2005) Fibroblast growth factor/fibroblast growth factor receptor system in angiogenesis. Cytokine Growth Factor Rev 16(2):159–178
Battegay EJ, Rupp J, Iruela-Arispe L et al (1994) PDGF-BB modulates endothelial proliferation and angiogenesis in vitro via PDGF beta-receptors. J Cell Biol 125(4):917–928
Xue Y, Lim S, Yang Y et al (2012) PDGF-BB modulates hematopoiesis and tumor angiogenesis by inducing erythropoietin production in stromal cells. Nat Med 18(1):100–110
Alajati A, Laib AM, Weber H et al (2008) Spheroid-based engineering of a human vasculature in mice. Nat Methods 5(5):439–44521
Nakatsu MN, Sainson RC, Aoto JN et al (2003) Angiogenic sprouting and capillary lumen formation modeled by human umbilical vein endothelial cells (HUVEC) in fibrin gels: the role of fibroblasts and Angiopoietin-1. Microvasc Res 66(2):102–11222
Vernon RB, Angello JC, Iruela-Arispe ML et al (1992) Reorganization of basement membrane matrices by cellular traction promotes the formation of cellular networks in vitro. Lab Invest 66(5):536–547
Grant D, Cid M, Kibbey MC et al (1992) Extracellular matrix-cell interaction: matrigel and complex cellular pattern formation. Lab Invest 67(6):805–806
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Pfisterer, L., Korff, T. (2016). Spheroid-Based In Vitro Angiogenesis Model. In: Martin, S., Hewett, P. (eds) Angiogenesis Protocols. Methods in Molecular Biology, vol 1430. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3628-1_11
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DOI: https://doi.org/10.1007/978-1-4939-3628-1_11
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