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VEGF Signaling pp 177–188Cite as

The Embryonic Mouse Hindbrain and Postnatal Retina as In Vivo Models to Study Angiogenesis

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1332))

Abstract

Angiogenesis, the growth of new blood vessels from preexisting ones, is a fundamental process for organ development, exercise-induced muscle growth, and wound healing, but is also associated with different diseases such as cancer and neovascular eye disease. Accordingly, elucidating the molecular and cellular mechanisms of angiogenesis has the potential to identify new therapeutic targets to stimulate new vessel formation in ischemic tissues or inhibit pathological vessel growth in disease. This chapter describes the mouse embryo hindbrain and postnatal retina as models to study physiological angiogenesis and provides detailed protocols for tissue dissection, sample staining, and analysis.

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References

  1. Ruhrberg C, Gerhardt H, Golding M et al (2002) Spatially restricted patterning cues provided by heparin-binding VEGF-A control blood vessel branching morphogenesis. Genes Dev 16:2684–2698

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  2. Gerhardt H, Ruhrberg C, Abramsson A et al (2004) Neuropilin-1 is required for endothelial tip cell guidance in the developing central nervous system. Dev Dyn 231:503–509

    Article  CAS  PubMed  Google Scholar 

  3. Fantin A, Vieira JM, Gestri G et al (2010) Tissue macrophages act as cellular chaperones for vascular anastomosis downstream of VEGF-mediated endothelial tip cell induction. Blood 116:829–840

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  4. Fantin A, Vieira JM, Plein A et al (2013) NRP1 acts cell autonomously in endothelium to promote tip cell function during sprouting angiogenesis. Blood 121:2352–2362

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  5. Graupera M, Guillermet-Guibert J, Foukas LC et al (2008) Angiogenesis selectively requires the p110alpha isoform of PI3K to control endothelial cell migration. Nature 453:662–666

    Article  CAS  PubMed  Google Scholar 

  6. Lu X, Le Noble F, Yuan L et al (2004) The netrin receptor UNC5B mediates guidance events controlling morphogenesis of the vascular system. Nature 432:179–186

    Article  CAS  PubMed  Google Scholar 

  7. Tammela T, Zarkada G, Nurmi H et al (2011) VEGFR-3 controls tip to stalk conversion at vessel fusion sites by reinforcing Notch signalling. Nat Cell Biol 13:1202–1213

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  8. Fantin A, Schwarz Q, Davidson K et al (2011) The cytoplasmic domain of neuropilin 1 is dispensable for angiogenesis, but promotes the spatial separation of retinal arteries and veins. Development 138:4185–4191

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  9. Benedito R, Roca C, Sorensen I et al (2009) The notch ligands Dll4 and Jagged1 have opposing effects on angiogenesis. Cell 137:1124–1135

    Article  CAS  PubMed  Google Scholar 

  10. Gerhardt H, Golding M, Fruttiger M et al (2003) VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia. J Cell Biol 161:1163–1177

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  11. Wang Y, Nakayama M, Pitulescu ME et al (2010) Ephrin-B2 controls VEGF-induced angiogenesis and lymphangiogenesis. Nature 465:483–486

    Article  CAS  PubMed  Google Scholar 

  12. Hellstrom M, Phng LK, Hofmann JJ et al (2007) Dll4 signalling through Notch1 regulates formation of tip cells during angiogenesis. Nature 445:776–780

    Article  PubMed  Google Scholar 

  13. Bar T (1983) Patterns of vascularization in the developing cerebral cortex. Ciba Found Symp 100:20–36

    CAS  PubMed  Google Scholar 

  14. Vieira JM, Schwarz Q, Ruhrberg C (2007) Selective requirements for NRP1 ligands during neurovascular patterning. Development 134:1833–1843

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  15. Fantin A, Vieira JM, Plein A et al (2013) The embryonic mouse hindbrain as a qualitative and quantitative model for studying the molecular and cellular mechanisms of angiogenesis. Nat Protoc 8:418–429

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  16. Breier G, Albrecht U, Sterrer S et al (1992) Expression of vascular endothelial growth factor during embryonic angiogenesis and endothelial cell differentiation. Development 114:521–532

    CAS  PubMed  Google Scholar 

  17. Raab S, Beck H, Gaumann A et al (2004) Impaired brain angiogenesis and neuronal apoptosis induced by conditional homozygous inactivation of vascular endothelial growth factor. Thromb Haemost 91:595–605

    CAS  PubMed  Google Scholar 

  18. Haigh JJ, Morelli PI, Gerhardt H et al (2003) Cortical and retinal defects caused by dosage-dependent reductions in VEGF-A paracrine signaling. Dev Biol 262:225–241

    Article  CAS  PubMed  Google Scholar 

  19. Farrell CL, Risau W (1994) Normal and abnormal development of the blood-brain barrier. Microsc Res Tech 27:495–506

    Article  CAS  PubMed  Google Scholar 

  20. Carmeliet P, Ng YS, Nuyens D et al (1999) Impaired myocardial angiogenesis and ischemic cardiomyopathy in mice lacking the vascular endothelial growth factor isoforms VEGF164 and VEGF188. Nat Med 5:495–502

    Article  CAS  PubMed  Google Scholar 

  21. Schwarz Q, Gu C, Fujisawa H et al (2004) Vascular endothelial growth factor controls neuronal migration and cooperates with Sema3A to pattern distinct compartments of the facial nerve. Genes Dev 18:2822–2834

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  22. Kawasaki T, Kitsukawa T, Bekku Y et al (1999) A requirement for neuropilin-1 in embryonic vessel formation. Development 126:4895–4902

    CAS  PubMed  Google Scholar 

  23. Jones EA, Yuan L, Breant C et al (2008) Separating genetic and hemodynamic defects in neuropilin 1 knockout embryos. Development 135:2479–2488

    Article  CAS  PubMed  Google Scholar 

  24. Nagy A (2000) Cre recombinase: the universal reagent for genome tailoring. Genesis 26:99–109

    Article  CAS  PubMed  Google Scholar 

  25. Fruttiger M (2007) Development of the retinal vasculature. Angiogenesis 10:77–88

    Article  PubMed  Google Scholar 

  26. Hofmann JJ, Luisa Iruela-Arispe M (2007) Notch expression patterns in the retina: an eye on receptor-ligand distribution during angiogenesis. Gene Expr Patterns 7:461–470

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  27. Carmeliet P, Tessier-Lavigne M (2005) Common mechanisms of nerve and blood vessel wiring. Nature 436:193–200

    Article  CAS  PubMed  Google Scholar 

  28. Hughes S, Chang-Ling T (2000) Roles of endothelial cell migration and apoptosis in vascular remodeling during development of the central nervous system. Microcirculation 7:317–333

    Article  CAS  PubMed  Google Scholar 

  29. Sawamiphak S, Ritter M, Acker-Palmer A (2010) Preparation of retinal explant cultures to study ex vivo tip endothelial cell responses. Nat Protoc 5:1659–1665

    Article  CAS  PubMed  Google Scholar 

  30. Zudaire E, Gambardella L, Kurcz C et al (2011) A computational tool for quantitative analysis of vascular networks. PLoS One 6:e27385

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  31. Pitulescu ME, Schmidt I, Benedito R et al (2010) Inducible gene targeting in the neonatal vasculature and analysis of retinal angiogenesis in mice. Nat Protoc 5:1518–1534

    Article  CAS  PubMed  Google Scholar 

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Acknowledgement

We thank Marcus Fruttiger and Shalini Jadeja for teaching us the retina dissection technique.

Author information

Authors and Affiliations

  1. UCL Institute of Ophthalmology, University College London, 11-43 Bath Street, London, EC1V 9EL, UK

    Alessandro Fantin & Christiana Ruhrberg

Authors
  1. Alessandro Fantin
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  2. Christiana Ruhrberg
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Corresponding author

Correspondence to Alessandro Fantin .

Editor information

Editors and Affiliations

  1. Caversham, Reading, United Kingdom

    Lorna Fiedler

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Fantin, A., Ruhrberg, C. (2015). The Embryonic Mouse Hindbrain and Postnatal Retina as In Vivo Models to Study Angiogenesis. In: Fiedler, L. (eds) VEGF Signaling. Methods in Molecular Biology, vol 1332. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2917-7_13

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  • DOI: https://doi.org/10.1007/978-1-4939-2917-7_13

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2916-0

  • Online ISBN: 978-1-4939-2917-7

  • eBook Packages: Springer Protocols

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