Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Robo4 stabilizes the vascular network by inhibiting pathologic angiogenesis and endothelial hyperpermeability

A Corrigendum to this article was published on 01 May 2008

This article has been updated


The angiogenic sprout has been compared to the growing axon, and indeed, many proteins direct pathfinding by both structures1. The Roundabout (Robo) proteins are guidance receptors with well-established functions in the nervous system2,3; however, their role in the mammalian vasculature remains ill defined4,5,6,7,8. Here we show that an endothelial-specific Robo, Robo4, maintains vascular integrity. Activation of Robo4 by Slit2 inhibits vascular endothelial growth factor (VEGF)-165–induced migration, tube formation and permeability in vitro and VEGF-165–stimulated vascular leak in vivo by blocking Src family kinase activation. In mouse models of retinal and choroidal vascular disease, Slit2 inhibited angiogenesis and vascular leak, whereas deletion of Robo4 enhanced these pathologic processes. Our results define a previously unknown function for Robo receptors in stabilizing the vasculature and suggest that activating Robo4 may have broad therapeutic application in diseases characterized by excessive angiogenesis and/or vascular leak.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Robo4 expression is endothelial specific and stalk-cell centric.
Figure 2: Robo4 signaling inhibits VEGF-165–induced migration, tube formation, permeability and SFK activation.
Figure 3: Slit2 blocks oxygen-induced retinopathy in a Robo4-dependent manner.
Figure 4: Robo4 signaling inhibits pathologic angiogenesis.

Change history

  • 18 April 2008

    Nat. Med. 14, 448–453 (2008); published online 16 March 2008; corrected after print 18 April 2008. In the version of this article initially published, the affiliation of Rebecca A. Stockton was incorrect. Her correct affiliation is affiliation 5: the Department of Medicine, University of California, San Diego, 9500 Gilman Dr., La Jolla, California 92093-0726, USA. The error has been corrected in the HTML and PDF versions of the article.


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

    Article  CAS  Google Scholar 

  2. Kidd, T. et al. Roundabout controls axon crossing of the CNS midline and defines a novel subfamily of evolutionarily conserved guidance receptors. Cell 92, 205–215 (1998).

    Article  CAS  Google Scholar 

  3. Long, H. et al. Conserved roles for Slit and Robo proteins in midline commissural axon guidance. Neuron 42, 213–223 (2004).

    Article  CAS  Google Scholar 

  4. Park, K.W. et al. Robo4 is a vascular-specific receptor that inhibits endothelial migration. Dev. Biol. 261, 251–267 (2003).

    Article  CAS  Google Scholar 

  5. Wang, B. et al. Induction of tumor angiogenesis by Slit-Robo signaling and inhibition of cancer growth by blocking Robo activity. Cancer Cell 4, 19–29 (2003).

    Article  Google Scholar 

  6. Suchting, S., Heal, P., Tahtis, K., Stewart, L.M. & Bicknell, R. Soluble Robo4 receptor inhibits in vivo angiogenesis and endothelial cell migration. FASEB J. 19, 121–123 (2005).

    Article  CAS  Google Scholar 

  7. Seth, P. et al. Magic roundabout, a tumor endothelial marker: expression and signaling. Biochem. Biophys. Res. Commun. 332, 533–541 (2005).

    Article  CAS  Google Scholar 

  8. Kaur, S. et al. Robo4 signaling in endothelial cells implies attraction guidance mechanisms. J. Biol. Chem. 281, 11347–11356 (2006).

    Article  CAS  Google Scholar 

  9. Battye, R., Stevens, A., Perry, R.L. & Jacobs, J.R. Repellent signaling by Slit requires the leucine-rich repeats. J. Neurosci. 21, 4290–4298 (2001).

    Article  CAS  Google Scholar 

  10. Howitt, J.A., Clout, N.J. & Hohenester, E. Binding site for Robo receptors revealed by dissection of the leucine-rich repeat region of Slit. EMBO J. 23, 4406–4412 (2004).

    Article  CAS  Google Scholar 

  11. Smith, L.E. et al. Oxygen-induced retinopathy in the mouse. Invest. Ophthalmol. Vis. Sci. 35, 101–111 (1994).

    CAS  PubMed  Google Scholar 

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

    Article  CAS  Google Scholar 

  13. Uemura, A., Kusuhara, S., Katsuta, H. & Nishikawa, S. Angiogenesis in the mouse retina: a model system for experimental manipulation. Exp. Cell Res. 312, 676–683 (2006).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  15. Cross, M.J., Dixelius, J., Matsumoto, T. & Claesson-Welsh, L. VEGF-receptor signal transduction. Trends Biochem. Sci. 28, 488–494 (2003).

    Article  CAS  Google Scholar 

  16. Eliceiri, B.P. et al. Src-mediated coupling of focal adhesion kinase to integrin alpha(v)beta5 in vascular endothelial growth factor signaling. J. Cell Biol. 157, 149–160 (2002).

    Article  CAS  Google Scholar 

  17. Eliceiri, B.P. et al. Selective requirement for Src kinases during VEGF-induced angiogenesis and vascular permeability. Mol. Cell 4, 915–924 (1999).

    Article  CAS  Google Scholar 

  18. Gavard, J. & Gutkind, J.S. VEGF controls endothelial-cell permeability by promoting the β-arrestin-dependent endocytosis of VE-cadherin. Nat. Cell Biol. 8, 1223–1234 (2006).

    Article  CAS  Google Scholar 

  19. Garrett, T.A., Van Buul, J.D. & Burridge, K. VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2. Exp. Cell Res. 313, 3285–3297 (2007).

    Article  CAS  Google Scholar 

  20. Yuan, W. et al. The mouse SLIT family: secreted ligands for ROBO expressed in patterns that suggest a role in morphogenesis and axon guidance. Dev. Biol. 212, 290–306 (1999).

    Article  CAS  Google Scholar 

  21. Morlot, C. et al. Structural insights into the Slit-Robo complex. Proc. Natl. Acad. Sci. USA 104, 14923–14928 (2007).

    Article  CAS  Google Scholar 

  22. Patel, K. et al. Slit proteins are not dominant chemorepellents for olfactory tract and spinal motor axons. Development 128, 5031–5037 (2001).

    CAS  PubMed  Google Scholar 

  23. Brown, D.M. et al. Ranibizumab versus verteporfin for neovascular age-related macular degeneration. N. Engl. J. Med. 355, 1432–1444 (2006).

    Article  CAS  Google Scholar 

  24. Rosenfeld, P.J. et al. Ranibizumab for neovascular age-related macular degeneration. N. Engl. J. Med. 355, 1419–1431 (2006).

    Article  CAS  Google Scholar 

  25. Ozaki, H. et al. Blockade of vascular endothelial cell growth factor receptor signaling is sufficient to completely prevent retinal neovascularization. Am. J. Pathol. 156, 697–707 (2000).

    Article  CAS  Google Scholar 

  26. Werdich, X.Q., McCollum, G.W., Rajaratnam, V.S. & Penn, J.S. Variable oxygen and retinal VEGF levels: correlation with incidence and severity of pathology in a rat model of oxygen-induced retinopathy. Exp. Eye Res. 79, 623–630 (2004).

    Article  CAS  Google Scholar 

  27. Plump, A.S. et al. Slit1 and Slit2 cooperate to prevent premature midline crossing of retinal axons in the mouse visual system. Neuron 33, 219–232 (2002).

    Article  CAS  Google Scholar 

  28. Lima e Silva, R. et al. Suppression and regression of choroidal neovascularization by polyamine analogues. Invest. Ophthalmol. Vis. Sci. 46, 3323–3330 (2005).

    Article  Google Scholar 

  29. Xu, Q., Qaum, T. & Adamis, A.P. Sensitive blood-retinal barrier breakdown quantitation using Evans blue. Invest. Ophthalmol. Vis. Sci. 42, 789–794 (2001).

    CAS  PubMed  Google Scholar 

Download references


We thank K. Thomas and J. Wythe for critical reading of the manuscript, L. Sorensen-Brunhart and W. Zhu for technical assistance and D. Lim for expert graphical assistance. This work was funded by grants from the US National Cancer Institute Multidisciplinary Cancer Research Training Program (N.R.L.); Cancer Research–UK (H.G.); the National Eye Institute (K.Z.); National Heart, Lung and Blood Institute (M.H.G. and D.Y.L.); and National Institute of Arthritis and Musculoskeletal and Skin Diseases (M.H.G.), the H.A. and Edna Benning Foundation, the Juvenile Diabetes Research Foundation, the American Heart Association, the Burroughs Wellcome Fund and the Flight Attendants Medical Research Institute (D.Y.L.).

Author information

Authors and Affiliations


Corresponding authors

Correspondence to Kang Zhang or Dean Y Li.

Ethics declarations

Competing interests

C.A.J., N.R.L., H.C., K.W.P., J.D.W., W.S., F.L.-L., A.F., K.Z. and D.Y.L. are or were previously employed by the University of Utah, which has filed intellectual property surrounding the therapeutic uses of targeting Robo4 and with the intent to license this body of intellectual property for commercialization.

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–10 and Supplementary Table 1 (PDF 7932 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Jones, C., London, N., Chen, H. et al. Robo4 stabilizes the vascular network by inhibiting pathologic angiogenesis and endothelial hyperpermeability. Nat Med 14, 448–453 (2008).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing