Molecular Control of Neuro-Vascular Development

Principal Investigator: Isabelle BRUNET, CR1 Inserm

Our laboratory studies vascular and neuronal development, with particular interests in mechanisms that direct patterning and guidance. Specialized endothelial cells (EC) called tip cells located at the extremities of growing capillary sprouts mediate guided vascular patterning. Tip cells exhibit characteristic features, including extension of filopodia that explore the tip cell environment, lack of a lumen and a slow proliferation rate. Capillary sprouting shows morphological similarities to axon guidance. Our research is focused on tip cell formation and guidance of angiogenesis and lymphangiogenesis. We are also closely investigating relashionship between vascular and nervous system and how they developp interactions at a molecular and functionnal level.

Guidance of Angiogenesis and Lymphangiogenesis

Like endothelial tip cells, axonal growth cones extend filopodia that sense and respond to guidance cues provided by soluble, cell- or matrix-bound ligands (Figure 1). We have identified several key molecules regulate capillary and axon guidance, including Robo4, UNC5B and the Neuropilin receptors 1 and 2 (Nrp1, Nrp2) (Koch et al., Dev Cell 2011, Xu et al., J Cell Biol 2010). Nrp1 and Nrp2 regulate guidance of distinct sets of axons to their targets. In the vascular system, Nrp1 and 2 are expressed in distinct compartments, with Nrp1 mainly labeling arteries and Nrp2 veins and lymphatic vessels. Deletion of the genes encoding Nrp1 or Nrp2 specifically affects arterial or lymphatic vessel sprouting, respectively. Despite prominent expression of Nrp2 in lymphatic vessels, Nrp2 expression is absent from valve forming lymphatic EC (LECs). Valves prevent backflow of lymph, and abnormal valve formation leads to lymphedema formation. Lymphatic valves express the Nrp1 receptor and one of the four signal-transducing type A Plexins, PlexinA1. We have shown that Sema3A preferentially binds to lymphatic valves and that valve formation is deficient in mouse mutants for Sema3a, Plexna1 and the Sema3A binding site in Nrp1 (Bouvrée, Brunet et al., Circ. Res. 2012). These experiments reveal essential, non-redundant function for Nrp signaling in lymphatic development: VEGF-C signaling through Nrp2-VEGFR3 directs lymphatic sprouting, while Sema3A-Nrp1 signaling directs valve morphogenesis.

Figure 1. Left image: endothelial tip cell, right image axonal growth cone. Middle: structure of guidance molecules common to the vascular and the nervous system.

Endothelial Plasticity

To identify the factors involved in the endothelial plasticity is important in cardiovascular research specially to improve the morbidity and the cost of arterioveinous bridging. Our current research in avian model showed that vasoconstrictor molecules induced the lost of the endothelial plasticity while vasodilating factors mediated the recovery of the endothelial plasticity. Using pharmacological and molecular approaches, the project will consist to decipher more precisely the different components of the sympathetic nervous system playing a role in the endothelial plasticity. Furthermore, we would like to know the link between the maintenance of the endothelial plasticity and the ontogeny of the peripheral nervous system.

Arterial Innervation

Our as yet unpublished work (Brunet et al.) directly investigates interactions between the nervous and the vascular systems, in particular arterial innervation (Figure 2). Innervation of peripheral resistance arteries by autonomic sympathetic nerves controls blood supply to organs by regulating vascular tone. Despite the fundamental importance of blood flow control and vascular tone, signals controlling the development of sympathetic arterial innervation are currently unknown. We have determined the developmental time window when arterial innervation is initiated in mice, and identified several novel regulators of arterial innervation that we are currently characterizing. Deletion of those regulators in mice models in a cell-type and inducible fashion give rise to hypo or hyper-innervation and we have now models allowing to modifie sympathetic innervation to study the impact of arterial innervation in physiological and pathological conditions such as hypertension.

Figure 2. Neurovascular interactions. Postnatal day 2 mouse mesentery stained with the indicated markers for vessels (green) and axons (red). Note preferential alignment of axons with arteries (A) but not veins (V) or lymphatic vessels.

Selected publications

- Minocha, S., Valloton, D., Brunet, I., Eichmann, A., Hornung, J.-P., Lebrand, C., (2015), NG2 glia are required for vessel network formation during embryonic development. Elife 4.

- Aspalter, I.M., Gordon, E., Dubrac, A., Ragab, A., Narloch, J., Vizán, P., Geudens, I., Collins, R.T., Franco, C.A., Abrahams, C.L., Thurston, G., Fruttiger, M., Rosewell, I., Eichmann, A., Gerhardt, H., (2015). Alk1 and Alk5 inhibition by Nrp1 controls vascular sprouting downstream of Notch. Nat Commun 6, 7264.

- Fortuna, V., Pardanaud, L., Brunet, I., Ola, R., Ristori, E., Santoro, M.M., Nicoli, S., and Eichmann, A. (2015). Vascular Mural Cells Promote Noradrenergic Differentiation of Embryonic Sympathetic Neurons. Cell Rep 11, 1786–1796.

- Rama N., Dubrac A., Mathivet T., Ní Chárthaigh R.-A., Genet G., Cristofaro B., Pibouin-Fragner L., Ma L., Eichmann A. & Chédotal A. (2015), Slit2 signaling through Robo1 and Robo2 is required for retinal neovascularization. Nat. Med. 21, 483–491. 

- Eichmann A. & Brunet I. (2014), Arterial innervation in development and disease. Science Translational Medecine 6, 252ps9.

- Greif D.M. and Eichmann A. (2014), Vascular biology: Brain vessels squeezed to death. Nature 508, 50–51.

- Prahst C., Kasaai B., Moraes F., Jahnsen E.D., Larrivee B., Villegas D., Pardanaud L., Pibouin-Fragner L., Zhang F., Zaun H.C., et al. (2014), The Homeobox Transcription Factor H2.0-Like Homeobox Transcription Factor Modulates Yolk Sac Vascular Remodeling in Mouse Embryos. Thromb. Vasc. Biol. 34, 1468–1476.

- Brunet I., Gordon E., Han J., Cristofaro B., Broqueres-You D., Liu C., Bouvrée K., Zhang J., del Toro R., Mathivet T., Larrivée B., Jagu J., Pibouin-Fragner L., Pardanaud L., Machado M.J.C., Kennedy T.E., Zhuang Z., Simons M., Levy B.I., Tessier-Lavigne M., Grenz A., Eltzschig H. & Eichmann A. (2014), Netrin-1 controls sympathetic arterial innervation. Journal of Clinical Investigation 124, 3230–3240.

- Cristofaro B., Shi Y., Faria M., Suchting S., Leroyer A.S., Trindade A., Duarte A., Zovein A.C., Iruela-Arispe M.L., Nih L.R., Kubis N., Henrion D., Loufrani L., Todiras M., Schleifenbaum J., Gollasch M., Zhuang Z.W., Simons M., Eichmann A. & le Noble F. (2013), Dll4-Notch signaling determines the formation of native arterial collateral networks and arterial function in mouse ischemia models. Development 140, 1720-1729

- Bouvrée K.*, Brunet I.*, del Toro R., Gordon E., Prahst C., Cristofaro B., Mathivet T., Xu Y., Soueid J., Fortuna V., Miura N., Aigrot M.S., Maden C.H., Ruhrberg C., Thomas J.L. & Eichmann A. (2012), Semaphorin3A, Neuropilin-1 and PlexinA1 are required for lymphatic valve formation. Circ. Res., 111: 437-445. * equal contribution

- Larrivée B., Prahst C., Gordon E., del Toro R., Mathivet T., Duarte A., Simons M. & Eichmann A. (2012), Alk1 signaling inhibits angiogenesis by cooperating with the Notch pathway. Dev. Cell, 22: 489-500.

- Pardanaud L. & Eichmann A. (2011), Extraembryonic origin of circulating endothelial cells. Plos One, 6: e25889.

- Tammela T., Zarkada G., Nurmi H., Jacobsson L., Heinolainen K., Tvogorov D., Mutomäki A., Franco C., Aranda E., Yla-Herttuala S., Fruttiger M., Mäkinen T., Eichmann A., Pollard J., Gerhardt H., Alitalo K. (2011), VEGFR-3 reinforces Notch signaling through FoxC2 to control angiogenesis. Nat Cell Biol, 11: 1202-13.

- Calvo C.F., Fontaine R.H., Soueid J., Tammela T., Makinen T., Alfaro-Cervello C., Bonnaud F., Miguez A., Benhaim L., Xu Y., Barallobre M.J., Moutkine I., Lyytikkä J., Tatlisumak T., Pytowski B., Zalc B., Richardson W., Kessaris N., Garcia-Verdugo J.M., Alitalo K., Eichmann A. & Thomas J.L. (2011), Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis. Genes & Dev., 25: 831-844.

- Koch A.W.*, Mathivet T.*, Larrivée B., Tong R.K., Kowalski J., Pibouin-Fragner L., Bouvrée K., Stawicki S., Nicholes K., Rathore N., Scales S.J., Luis E., del Toro R., Freitas C., Bréant C., Michaud A., Corvol P., Thomas J.L., Wu J., Peale F., Watts R.J., Tessier-Lavigne M., Bagri A. & Eichmann A.* (2011), Robo4 maintains vessel integrity and inhibits angiogenesis by interacting with UNC5B. Dev. Cell, 20: 33-46. * equal contribution

- Del Toro R., Prahst C., Mathivet T., Siegfried G., Kaminker J., Larrivée B., Bréant C., Duarte A., Takakura N., Fukamizu A., Penninger J. & Eichmann A. (2010), Identification and functional analysis of novel endothelial tip cell-enriched genes. Blood, 116: 4025-33.

- Lebrin F., Srun S., Raymond K., Martin S., van den Brink S., Freitas C., Bréant C., Mathivet T., Larrivée B., Thomas J.L., Arthur H.A., Westermann C.J.J., Disch F., Mager J.J., Snijder R.J., Eichmann A.* & Mummery C. (2010), Thalidomide enhances mural cell recruitment and reduces epistaxis in patients with hereditary hemorrhagic telangieectasia. Nat. Med.; 16: 420-8. * equal contribution

- Xu Y., Yuan L., Mak J., Pardanaud L., Caunt M., Kasman I., Larrivée B., Suchting S., del Toro R., Medvinsky A., Yang J., Kolodkin A., Thomas J.L., Koch A., Alitalo K., Eichmann A.* & Bagri A.* (2010), Neuropilin-2 mediates VEGF-C induced lymphatic sprouting together with VEGFR3. J. Cell Biol.; 188:115-30. * equal contribution


Group leader:
Brunet Isabelle, CR1 INSERM

Senior researcher:
Malkinson Guy, MDC CDF

Technical staff:

Martin Sabrina, IE2 INSERM

Postdoctoral fellows & PhD Students:
Azar Safa, Postdoctoral fellow
Simonnet Emilie, PhD student
Taib Sonia, PhD student