Skip to main content
SpringerLink
Log in

Vasculogenesis and Angiogenesis in VEGF Receptor-1 Deficient Mice

  • Protocol
Book cover VEGF Signaling

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

Abstract

Vascular endothelial growth factor receptor-1 (VEGFR-1)/Flt-1 is a transmembrane tyrosine kinase receptor for VEGF-A, VEGF-B, and placental growth factor (PlGF). VEGFR-1 is an enigmatic molecule whose precise role in postnatal angiogenesis remains controversial. Although many postnatal and adult studies have been performed by manipulating VEGFR-1 ligands, including competitive binding by truncated VEGFR-1 protein, neutralization by antibodies, or specific ligand overexpression or knockout, much less is known at the level of the receptor per se, especially in vivo. Perplexingly, while VEGFR-1 negatively regulates endothelial cell differentiation during development, it has been implied in promoting angiogenesis under certain conditions in adult tissues, especially in tumors and ischemic tissues. Additionally, it is unclear how VEGFR-1 is involved in vascular maturation and maintenance of vascular quiescence in adult tissues. To facilitate further investigation, we generated a conditional knockout mouse line for VEGFR-1 and characterized angiogenesis in postnatal and adult mice, including angiogenesis in ischemic myocardium. These methods are briefly outlined in this chapter. We also discuss these findings in the context of the interplay between VEGF family members and their receptors, and summarize various mouse models in the VEGF pathway.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Shibuya M, Yamaguchi S, Yamane A et al (1990) Nucleotide sequence and expression of a novel human receptor-type tyrosine kinase gene (flt) closely related to the fms family. Oncogene 5:519–524

    CAS  PubMed  Google Scholar 

  2. de Vries C, Escobedo JA, Ueno H et al (1992) The fms-like tyrosine kinase, a receptor for vascular endothelial growth factor. Science 255:989–991

    Article  PubMed  Google Scholar 

  3. Dumont DJ, Fong GH, Puri MC et al (1995) Vascularization of the mouse embryo: a study of flk-1, tek, tie, and vascular endothelial growth factor expression during development. Dev Dyn 203:80–92

    Article  CAS  PubMed  Google Scholar 

  4. Kendall RL, Thomas KA (1993) Inhibition of vascular endothelial cell growth factor activity by an endogenously encoded soluble receptor. Proc Natl Acad Sci U S A 90:10705–10709

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  5. Millauer B, Wizigmann-Voos S, Schnurch H et al (1993) High affinity VEGF binding and developmental expression suggest Flk-1 as a major regulator of vasculogenesis and angiogenesis. Cell 72:835–846

    Article  CAS  PubMed  Google Scholar 

  6. Waltenberger J, Claesson-Welsh L, Siegbahn A et al (1994) Different signal transduction properties of KDR and Flt1, two receptors for vascular endothelial growth factor. J Biol Chem 269:26988–26995

    CAS  PubMed  Google Scholar 

  7. Watanabe Y, Lee SW, Detmar M et al (1997) Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) delays and induces escape from senescence in human dermal microvascular endothelial cells. Oncogene 14:2025–2032

    Article  CAS  PubMed  Google Scholar 

  8. Clauss M, Weich H, Breier G et al (1996) The vascular endothelial growth factor receptor Flt-1 mediates biological activities. Implications for a functional role of placenta growth factor in monocyte activation and chemotaxis. J Biol Chem 271:17629–17634

    Article  CAS  PubMed  Google Scholar 

  9. Gerber HP, McMurtrey A, Kowalski J et al (1998) Vascular endothelial growth factor regulates endothelial cell survival through the phosphatidylinositol 3′-kinase/Akt signal transduction pathway. Requirement for Flk-1/KDR activation. J Biol Chem 273:30336–30343

    Article  CAS  PubMed  Google Scholar 

  10. Fong GH, Rossant J, Gertsenstein M et al (1995) Role of the Flt-1 receptor tyrosine kinase in regulating the assembly of vascular endothelium. Nature 376:66–70

    Article  CAS  PubMed  Google Scholar 

  11. Fong GH, Zhang L, Bryce DM et al (1999) Increased hemangioblast commitment, not vascular disorganization, is the primary defect in flt-1 knock-out mice. Development 126:3015–3025

    CAS  PubMed  Google Scholar 

  12. Carmeliet P, Ferreira V, Breier G et al (1996) Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele. Nature 380:435–439

    Article  CAS  PubMed  Google Scholar 

  13. Ferrara N, Carver-Moore K, Chen H et al (1996) Heterozygous embryonic lethality induced by targeted inactivation of the VEGF gene. Nature 380:439–442

    Article  CAS  PubMed  Google Scholar 

  14. Shalaby F, Rossant J, Yamaguchi TP et al (1995) Failure of blood-island formation and vasculogenesis in Flk-1-deficient mice. Nature 376:62–66

    Article  CAS  PubMed  Google Scholar 

  15. Shalaby F, Ho J, Stanford WL et al (1997) A requirement for Flk1 in primitive and definitive hematopoiesis and vasculogenesis. Cell 89:981–990

    Article  CAS  PubMed  Google Scholar 

  16. Takashima S, Kitakaze M, Asakura M et al (2002) Targeting of both mouse neuropilin-1 and neuropilin-2 genes severely impairs developmental yolk sac and embryonic angiogenesis. Proc Natl Acad Sci U S A 99:3657–3662

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  17. Bellomo D, Headrick JP, Silins GU et al (2000) Mice lacking the vascular endothelial growth factor-B gene (Vegfb) have smaller hearts, dysfunctional coronary vasculature, and impaired recovery from cardiac ischemia. Circ Res 86:E29–E35

    Article  CAS  PubMed  Google Scholar 

  18. Aase K, von Euler G, Li X et al (2001) Vascular endothelial growth factor-B-deficient mice display an atrial conduction defect. Circulation 104:358–364

    Article  CAS  PubMed  Google Scholar 

  19. Carmeliet P, Moons L, Luttun A et al (2001) Synergism between vascular endothelial growth factor and placental growth factor contributes to angiogenesis and plasma extravasation in pathological conditions. Nat Med 7:575–583

    Article  CAS  PubMed  Google Scholar 

  20. Hiratsuka S, Minowa O, Kuno J et al (1998) Flt-1 lacking the tyrosine kinase domain is sufficient for normal development and angiogenesis in mice. Proc Natl Acad Sci U S A 95:9349–9354

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. Hiratsuka S, Nakao K, Nakamura K et al (2005) Membrane fixation of vascular endothelial growth factor receptor 1 ligand-binding domain is important for vasculogenesis and angiogenesis in mice. Mol Cell Biol 25:346–354

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  22. Kearney JB, Kappas NC, Ellerstrom C et al (2004) The VEGF receptor flt-1 (VEGFR-1) is a positive modulator of vascular sprout formation and branching morphogenesis. Blood 103:4527–4535

    Article  CAS  PubMed  Google Scholar 

  23. Luttun A, Tjwa M, Moons L et al (2002) Revascularization of ischemic tissues by PlGF treatment, and inhibition of tumor angiogenesis, arthritis and atherosclerosis by anti-Flt1. Nat Med 8:831–840

    CAS  PubMed  Google Scholar 

  24. Ho VC, Duan LJ, Cronin C et al (2012) Elevated vascular endothelial growth factor receptor-2 abundance contributes to increased angiogenesis in vascular endothelial growth factor receptor-1-deficient mice. Circulation 126:41–752

    Article  Google Scholar 

  25. Shizuya H, Birren B, Kim UJ et al (1992) Cloning and stable maintenance of 300-kilobase-pair fragments of human DNA in Escherichia coli using an F-factor-based vector. Proc Natl Acad Sci U S A 89:8794–8797

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  26. Liu P, Jenkins NA, Copeland NG (2003) A highly efficient recombineering-based method for generating conditional knockout mutations. Genome Res 13:476–484

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  27. Joyner AL (2000) Gene targeting: a practical approach, vol xviii. Oxford University Press, Oxford, p 293

    Google Scholar 

  28. George SH, Gertsenstein M, Vintersten K et al (2007) Developmental and adult phenotyping directly from mutant embryonic stem cells. Proc Natl Acad Sci U S A 104:4455–4460

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  29. Vintersten K, Monetti C, Gertsenstein M et al (2004) Mouse in red: red fluorescent protein expression in mouse ES cells, embryos, and adult animals. Genesis 40:241–246

    Article  CAS  PubMed  Google Scholar 

  30. Sonin D, Zhou SY, Cronin C et al (2008) Role of P2X purinergic receptors in the rescue of ischemic heart failure. Am J Physiol Heart Circ Physiol 295:H1191–H1197

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  31. Gerber HP, Hillan KJ, Ryan AM et al (1999) VEGF is required for growth and survival in neonatal mice. Development 126:1149–1159

    CAS  PubMed  Google Scholar 

  32. Damert A, Miquerol L, Gertsenstein M et al (2002) Insufficient VEGFA activity in yolk sac endoderm compromises haematopoietic and endothelial differentiation. Development 129:1881–1892

    CAS  PubMed  Google Scholar 

  33. Stalmans I, Ng YS, Rohan R et al (2002) Arteriolar and venular patterning in retinas of mice selectively expressing VEGF isoforms. J Clin Invest 109:327–336

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  34. Lee S, Chen TT, Barber CL et al (2007) Autocrine VEGF signaling is required for vascular homeostasis. Cell 130:691–703

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  35. Jin J, Sison K, Li C et al (2012) Soluble FLT1 binds lipid microdomains in podocytes to control cell morphology and glomerular barrier function. Cell 151:384–399

    Article  CAS  PubMed  Google Scholar 

  36. Sakurai Y, Ohgimoto K, Kataoka Y et al (2005) Essential role of Flk-1 (VEGF receptor 2) tyrosine residue 1173 in vasculogenesis in mice. Proc Natl Acad Sci U S A 102:1076–1081

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  37. Benedito R, Rocha SF, Woeste M et al (2012) Notch-dependent VEGFR3 upregulation allows angiogenesis without VEGF-VEGFR2 signalling. Nature 484:110–114

    Article  CAS  PubMed  Google Scholar 

  38. Dumont DJ, Jussila L, Taipale J et al (1998) Cardiovascular failure in mouse embryos deficient in VEGF receptor-3. Science 282:946–949

    Article  CAS  PubMed  Google Scholar 

  39. 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 

  40. Hagberg CE, Falkevall A, Wang X et al (2010) Vascular endothelial growth factor B controls endothelial fatty acid uptake. Nature 464:917–921

    Article  CAS  PubMed  Google Scholar 

  41. Kitsukawa T, Shimizu M, Sanbo M et al (1997) Neuropilin-semaphorin III/D-mediated chemorepulsive signals play a crucial role in peripheral nerve projection in mice. Neuron 19:995–1005

    Article  CAS  PubMed  Google Scholar 

  42. 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 

  43. 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 

  44. Giger RJ, Cloutier JF, Sahay A et al (2000) Neuropilin-2 is required in vivo for selective axon guidance responses to secreted semaphorins. Neuron 25:29–41

    Article  CAS  PubMed  Google Scholar 

  45. Walz A, Rodriguez I, Mombaerts P (2002) Aberrant sensory innervation of the olfactory bulb in neuropilin-2 mutant mice. J Neurosci 22:4025–4035

    CAS  PubMed  Google Scholar 

  46. Karkkainen MJ, Haiko P, Sainio K et al (2004) Vascular endothelial growth factor C is required for sprouting of the first lymphatic vessels from embryonic veins. Nat Immunol 5:74–80

    Article  CAS  PubMed  Google Scholar 

  47. Baldwin ME, Halford MM, Roufail S et al (2005) Vascular endothelial growth factor D is dispensable for development of the lymphatic system. Mol Cell Biol 25:2441–2449

    Article  PubMed Central  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Cell Biology, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, CT, 06030-3501, USA

    Vivienne C. Ho & Guo-Hua Fong

  2. Center for Vascular Biology, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, CT, 06030-3501, USA

    Vivienne C. Ho & Guo-Hua Fong

Authors
  1. Vivienne C. Ho
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Guo-Hua Fong
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Guo-Hua Fong .

Editor information

Editors and Affiliations

  1. Caversham, Reading, United Kingdom

    Lorna Fiedler

Rights and permissions

Reprints and permissions

Copyright information

© 2015 Springer Science+Business Media New York

About this protocol

Cite this protocol

Ho, V.C., Fong, GH. (2015). Vasculogenesis and Angiogenesis in VEGF Receptor-1 Deficient Mice. 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_12

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-2917-7_12

  • 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

Publish with us

Policies and ethics

Search

Navigation