Abstract
During development and in various diseases of the CNS, new blood vessel formation starts with endothelial tip cell selection and vascular sprout migration, followed by the establishment of functional, perfused blood vessels. Here we describe a method that allows the assessment of these distinct angiogenic steps together with antibody-based protein detection in the postnatal mouse brain. Intravascular and perivascular markers such as Evans blue (EB), isolectin B4 (IB4) or laminin (LN) are used alongside simultaneous immunofluorescence on the same sections. By using confocal laser-scanning microscopy and stereological methods for analysis, detailed quantification of the 3D postnatal brain vasculature for perfused and nonperfused vessels (e.g., vascular volume fraction, vessel length and number, number of branch points and perfusion status of the newly formed vessels) and characterization of sprouting activity (e.g., endothelial tip cell density, filopodia number) can be obtained. The entire protocol, from mouse perfusion to vessel analysis, takes ∼10 d.
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References
Carmeliet, P. & Jain, R.K. Molecular mechanisms and clinical applications of angiogenesis. Nature 473, 298–307 (2011).
Potente, M., Gerhardt, H. & Carmeliet, P. Basic and therapeutic aspects of angiogenesis. Cell 146, 873–887 (2011).
Jain, R.K. & Carmeliet, P. SnapShot: tumor angiogenesis. Cell 149, 1408–1408 (2012).
Burri, P.H., Hlushchuk, R. & Djonov, V. Intussusceptive angiogenesis: its emergence, its characteristics, and its significance. Dev. Dyn. 231, 474–488 (2004).
Makanya, A.N., Hlushchuk, R. & Djonov, V.G. Intussusceptive angiogenesis and its role in vascular morphogenesis, patterning, and remodeling. Angiogenesis 12, 113–123 (2009).
Herbert, S.P. & Stainier, D.Y. Molecular control of endothelial cell behaviour during blood vessel morphogenesis. Nat. Rev. Mol. Cell Biol. 12, 551–564 (2011).
Mancuso, M.R., Kuhnert, F. & Kuo, C.J. Developmental angiogenesis of the central nervous system. Lymphat. Res. Biol. 6, 173–180 (2008).
Quaegebeur, A., Lange, C. & Carmeliet, P. The neurovascular link in health and disease: molecular mechanisms and therapeutic implications. Neuron 71, 406–424 (2011).
Fantin, A., Vieira, J.M., Plein, A., Maden, C.H. & Ruhrberg, C. The embryonic mouse hindbrain as a qualitative and quantitative model for studying the molecular and cellular mechanisms of angiogenesis. Nat. Protoc. 8, 418–429 (2013).
Daneman, R., Zhou, L., Kebede, A.A. & Barres, B.A. Pericytes are required for blood-brain barrier integrity during embryogenesis. Nature 468, 562–566 (2010).
Zeller, K., Vogel, J. & Kuschinsky, W. Postnatal distribution of Glut1 glucose transporter and relative capillary density in blood-brain barrier structures and circumventricular organs during development. Brain Res. Dev. Brain Res. 91, 200–208 (1996).
Walchli, T. et al. Nogo-A is a negative regulator of CNS angiogenesis. Proc. Natl. Acad. Sci. USA 110, E1943–E1952 (2013).
Geudens, I. & Gerhardt, H. Coordinating cell behaviour during blood vessel formation. Development 138, 4569–4583 (2011).
Eilken, H.M. & Adams, R.H. Dynamics of endothelial cell behavior in sprouting angiogenesis. Curr. Opin. Cell Biol. 22, 617–625 (2010).
Strilic, B. et al. The molecular basis of vascular lumen formation in the developing mouse aorta. Dev. Cell 17, 505–515 (2009).
Lammert, E. & Axnick, J. Vascular lumen formation. Cold Spring Harb. Perspect. Med. 2, a006619 (2012).
Tung, J.J., Tattersall, I.W. & Kitajewski, J. Tips, stalks, tubes: notch-mediated cell fate determination and mechanisms of tubulogenesis during angiogenesis. Cold Spring Harb. Perspect. Med. 2, a006601 (2012).
Wacker, A. & Gerhardt, H. Endothelial development taking shape. Curr. Opin. Cell Biol. 23, 676–685 (2011).
Mazzone, M. et al. Heterozygous deficiency of PHD2 restores tumor oxygenation and inhibits metastasis via endothelial normalization. Cell 136, 839–851 (2009).
Pries, A.R., Secomb, T.W. & Gaehtgens, P. Structural adaptation and stability of microvascular networks: theory and simulations. Am. J. Physiol. 275, H349–H360 (1998).
Zhang, L., Cooper-Kuhn, C.M., Nannmark, U., Blomgren, K. & Kuhn, H.G. Stimulatory effects of thyroid hormone on brain angiogenesis in vivo and in vitro. J. Cereb. Blood Flow Metab. 30, 323–335 (2010).
Frahm, K.A., Nash, C.P. & Tobet, S.A. Endocan immunoreactivity in the mouse brain: method for identifying nonfunctional blood vessels. J. Immunol. Methods 398–399, 27–32 (2013).
Lobov, I.B. et al. Delta-like ligand 4 (Dll4) is induced by VEGF as a negative regulator of angiogenic sprouting. Proc. Natl. Acad. Sci. USA 104, 3219–3224 (2007).
Blanco, R. & Gerhardt, H. VEGF and Notch in tip and stalk cell selection. Cold Spring Harb. Perspect. Med. 3, a006569 (2013).
Phng, L.K. & Gerhardt, H. Angiogenesis: a team effort coordinated by Notch. Dev. Cell 16, 196–208 (2009).
Thurston, G., Noguera-Troise, I. & Yancopoulos, G.D. The delta paradox: DLL4 blockade leads to more tumour vessels but less tumour growth. Nat. Rev. Cancer 7, 327–331 (2007).
Yan, M. et al. Chronic DLL4 blockade induces vascular neoplasms. Nature 463, E6–E7 (2010).
Noguera-Troise, I. et al. Blockade of Dll4 inhibits tumour growth by promoting non-productive angiogenesis. Nature 444, 1032–1037 (2006).
Ridgway, J. et al. Inhibition of Dll4 signalling inhibits tumour growth by deregulating angiogenesis. Nature 444, 1083–1087 (2006).
Noguera-Troise, I. et al. Blockade of Dll4 inhibits tumour growth by promoting non-productive angiogenesis. Novartis Found. Symp. 283, 106–120 discussion 121–125, 238–241 (2007).
Gobel, U., Theilen, H. & Kuschinsky, W. Congruence of total and perfused capillary network in rat brains. Circ. Res. 66, 271–281 (1990).
Carmeliet, P. Angiogenesis in life, disease and medicine. Nature 438, 932–936 (2005).
Storkebaum, E., Quaegebeur, A., Vikkula, M. & Carmeliet, P. Cerebrovascular disorders: molecular insights and therapeutic opportunities. Nat. Neurosci. 14, 1390–1397 (2011).
Pitulescu, M.E., Schmidt, I., Benedito, R. & Adams, R.H. Inducible gene targeting in the neonatal vasculature and analysis of retinal angiogenesis in mice. Nat. Protoc. 5, 1518–1534 (2010).
Connor, K.M. et al. Quantification of oxygen-induced retinopathy in the mouse: a model of vessel loss, vessel regrowth and pathological angiogenesis. Nat. Protoc. 4, 1565–1573 (2009).
Rawson, R.A. The binding of T-1824 and structurally related diazo dyes by the plasma proteins. Am. J. Physiol. 138, 708–717 (1943).
Weintraub, H. et al. Storage of glycoprotein in NCTR-BALB/c mouse. Lectin histochemistry, and biochemical studies. Virchows Arch. B Cell Pathol. Incl. Mol. Pathol. 62, 347–352 (1992).
DeGasperi, R. et al. Glycoprotein storage in Gaucher disease: lectin histochemistry and biochemical studies. Lab. Invest. 63, 385–392 (1990).
Laitinen, L. Griffonia simplicifolia lectins bind specifically to endothelial cells and some epithelial cells in mouse tissues. Histochem. J. 19, 225–234 (1987).
Laitinen, L., Virtanen, I. & Saxen, L. Changes in the glycosylation pattern during embryonic development of mouse kidney as revealed with lectin conjugates. J. Histochem. Cytochem. 35, 55–65 (1987).
Alroy, J., Goyal, V. & Warren, C.D. Lectin histochemistry of gangliosidosis. I. Neural tissue in four mammalian species. Acta Neuropathol. 76, 109–114 (1988).
Gerhardt, H. et al. Neuropilin-1 is required for endothelial tip cell guidance in the developing central nervous system. Dev. Dyn. 231, 503–509 (2004).
Gerhardt, H. et al. VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia. J. Cell Biol. 161, 1163–1177 (2003).
Sorokin, S.P. & Hoyt, R.F. Jr. Macrophage development: I. Rationale for using Griffonia simplicifolia isolectin B4 as a marker for the line. Anat. Rec. 232, 520–526 (1992).
Sorokin, S.P., Hoyt, R.F. Jr., Blunt, D.G. & McNelly, N.A. Macrophage development: II. Early ontogeny of macrophage populations in brain, liver, and lungs of rat embryos as revealed by a lectin marker. Anat. Rec. 232, 527–550 (1992).
Theilen, H., Schrock, H. & Kuschinsky, W. Capillary perfusion during incomplete forebrain ischemia and reperfusion in rat brain. Am. J. Physiol. 265, H642–H648 (1993).
Gundersen, H.J. Stereology of arbitrary particles. A review of unbiased number and size estimators and the presentation of some new ones, in memory of William R. Thompson. J. Microsc. 143, 3–45 (1986).
West, M.J. Basic Stereology for Biologists and Neuroscientists (Cold Spring Harbor Laboratory Press, 2012).
Gundersen, H.J. & Jensen, E.B. The efficiency of systematic sampling in stereology and its prediction. J. Microsc. 147, 229–263 (1987).
Boyce, R.W., Dorph-Petersen, K.A., Lyck, L. & Gundersen, H.J. Design-based stereology: introduction to basic concepts and practical approaches for estimation of cell number. Toxicol. Pathol. 38, 1011–1025 (2010).
Mouton, P.R., Gokhale, A.M., Ward, N.L. & West, M.J. Stereological length estimation using spherical probes. J. Microsc. 206, 54–64 (2002).
Gundersen, H.J. et al. Some new, simple and efficient stereological methods and their use in pathological research and diagnosis. APMIS 96, 379–394 (1988).
Gundersen, H.J. et al. The new stereological tools: disector, fractionator, nucleator and point sampled intercepts and their use in pathological research and diagnosis. APMIS 96, 857–881 (1988).
Schmitz, C. & Hof, P.R. Design-based stereology in neuroscience. Neuroscience 130, 813–831 (2005).
Howard, C.V. & Reed, M.G. Unbiased Stereology: Three-dimensional Measurements in Microscopy (QTP Publications, 2010).
Lokkegaard, A., Nyengaard, J.R. & West, M.J. Stereological estimates of number and length of capillaries in subdivisions of the human hippocampal region. Hippocampus 11, 726–740 (2001).
Sawamiphak, S., Ritter, M. & Acker-Palmer, A. Preparation of retinal explant cultures to study ex vivo tip endothelial cell responses. Nat. Protoc. 5, 1659–1665 (2010).
Gore, A.V., Monzo, K., Cha, Y.R., Pan, W. & Weinstein, B.M. Vascular development in the zebrafish. Cold Spring Harb. Perspect. Med. 2, a006684 (2012).
Ellertsdottir, E. et al. Developmental role of zebrafish protease-activated receptor 1 (PAR1) in the cardiovascular system. PLoS ONE 7, e42131 (2012).
Carmeliet, P., De Smet, F., Loges, S. & Mazzone, M. Branching morphogenesis and antiangiogenesis candidates: tip cells lead the way. Nat. Rev. Clin. Oncol. 6, 315–326 (2009).
Hellstrom, M. et al. Dll4 signalling through Notch1 regulates formation of tip cells during angiogenesis. Nature 445, 776–780 (2007).
Tammela, T. et al. Blocking VEGFR-3 suppresses angiogenic sprouting and vascular network formation. Nature 454, 656–660 (2008).
Strasser, G.A., Kaminker, J.S. & Tessier-Lavigne, M. Microarray analysis of retinal endothelial tip cells identifies CXCR4 as a mediator of tip cell morphology and branching. Blood 115, 5102–5110 (2010).
del Toro, R. et al. Identification and functional analysis of endothelial tip cell-enriched genes. Blood 116, 4025–4033 (2010).
Siemerink, M.J. et al. CD34 marks angiogenic tip cells in human vascular endothelial cell cultures. Angiogenesis 15, 151–163 (2012).
Jakobsson, L. et al. Endothelial cells dynamically compete for the tip cell position during angiogenic sprouting. Nat. Cell Biol. 12, 943–953 (2010).
Vogel, J., Gehrig, M., Kuschinsky, W. & Marti, H.H. Massive inborn angiogenesis in the brain scarcely raises cerebral blood flow. J. Cereb. Blood Flow Metab. 24, 849–859 (2004).
Heinzer, S. et al. Novel three-dimensional analysis tool for vascular trees indicates complete micro-networks, not single capillaries, as the angiogenic endpoint in mice overexpressing human VEGF(165) in the brain. Neuroimage 39, 1549–1558 (2008).
Kienast, Y. et al. Real-time imaging reveals the single steps of brain metastasis formation. Nat. Med. 16, 116–122 (2010).
Whiteus, C., Freitas, C. & Grutzendler, J. Perturbed neural activity disrupts cerebral angiogenesis during a postnatal critical period. Nature 505, 407–411 (2014).
Harb, R., Whiteus, C., Freitas, C. & Grutzendler, J. In vivo imaging of cerebral microvascular plasticity from birth to death. J. Cereb. Blood Flow Metab. 33, 146–156 (2013).
Chappell, J.C., Wiley, D.M. & Bautch, V.L. Regulation of blood vessel sprouting. Semin. Cell Dev. Biol. 22, 1005–1011 (2011).
Chappell, J.C., Wiley, D.M. & Bautch, V.L. How blood vessel networks are made and measured. Cells Tissues Organs 195, 94–107 (2012).
West, M.J., Slomianka, L. & Gundersen, H.J. Unbiased stereological estimation of the total number of neurons in the subdivisions of the rat hippocampus using the optical fractionator. Anat. Rec. 231, 482–497 (1991).
West, M.J. Introduction to stereology. Cold Spring Harb. Protoc. 2012 doi:10.1101/pdb.top070623 (2012).
West, M.J. The precision of estimates in stereological analyses. Cold Spring Harb. Protoc. 2012, 937–949 (2012).
West, M.J. Estimating object number in biological structures. Cold Spring Harb. Protoc. 2012, 1049–1066 (2012).
West, M.J. Estimating volume in biological structures. Cold Spring Harb. Protoc. 2012, 1129–1139 (2012).
West, M.J. Systematic versus random sampling in stereological studies. Cold Spring Harb. Protoc. 2012 doi:10.1101/pdb.top071837 (2012).
Reed, M.G., Howard, C.V. & GS, D.E.Y. One-stop stereology: the estimation of 3D parameters using isotropic rulers. J. Microsc. 239, 54–65 (2010).
Howard, C.V. & Reed, M.G. Unbiased Stereology (BIOS Scientific Publishers, 1998).
Sawamiphak, S. et al. Ephrin-B2 regulates VEGFR2 function in developmental and tumour angiogenesis. Nature 465, 487–491 (2010).
Gampel, A. et al. VEGF regulates the mobilization of VEGFR2/KDR from an intracellular endothelial storage compartment. Blood 108, 2624–2631 (2006).
Jopling, H.M., Howell, G.J., Gamper, N. & Ponnambalam, S. The VEGFR2 receptor tyrosine kinase undergoes constitutive endosome-to-plasma membrane recycling. Biochem. Biophys. Res. Commun. 410, 170–176 (2011).
Nakayama, M. et al. Spatial regulation of VEGF receptor endocytosis in angiogenesis. Nat. Cell Biol. 15, 249–260 (2013).
Gaengel, K. & Betsholtz, C. Endocytosis regulates VEGF signalling during angiogenesis. Nat. Cell Biol. 15, 233–235 (2013).
Wang, Y. et al. Ephrin-B2 controls VEGF-induced angiogenesis and lymphangiogenesis. Nature 465, 483–486 (2010).
Kent, T.A. et al. Cerebral blood volume in a rat model of ischemia by MR imaging at 4.7 T. AJNR Am. J. Neuroradiol. 10, 335–338 (1989).
Lin, W., Paczynski, R.P., Kuppusamy, K., Hsu, C.Y. & Haacke, E.M. Quantitative measurements of regional cerebral blood volume using MRI in rats: effects of arterial carbon dioxide tension and mannitol. Magn. Reson. Med. 38, 420–428 (1997).
Ruhrberg, C. & Bautch, V.L. Neurovascular development and links to disease. Cell. Mol. Life Sci. 70, 1675–1684 (2013).
Eichmann, A. & Thomas, J.L. Molecular parallels between neural and vascular development. Cold Spring Harb. Perspect. Med. 3, a006551 (2013).
Acknowledgements
We thank L. Slomianka for help with stereological analysis and A. Wacker for critical reading of the manuscript. T.W. was supported by an MD-PhD fellowship of the Swiss National Science Foundation, by the Olga Mayenfisch Foundation, the Hartmann Muller Foundation, the EMDO Foundation and by the MD-PhD student allowance of the Swiss Society for Microvascular Research (SSMVR). J.V. was supported by the Swiss National Science Foundation (no. 310000 120321/1). All the animal experiments were conducted in J.V.'s laboratory and were approved by the Veterinary office of the Canton of Zurich. The histological studies were performed in M.E.S.'s laboratory. Microscopy image acquisition and analysis was performed in M.E.S.'s laboratory and at the Center for Microscopy and Image Analysis, University of Zurich.
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T.W. and J.V. designed the experiments and wrote the manuscript. T.W., O.W. and J.V. conducted the experiments. T.W. and J.M.M. analyzed the data. J.V. and M.E.S. supervised the experiments in their respective laboratories. D.B. helped with the endothelial tip cell-marker experiments and gave critical inputs to the manuscript. H.G., S.P.H. and L.R. gave critical inputs to the manuscript. All authors read and approved the final version of the manuscript.
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Supplementary Figure 1 Optimization of the tissue fixation protocol to combine EB perfusion and IB4 endothelial tip cell staining in the postnatal mouse brain.
a-l Labeling of EB-perfused P8 mice with biotinylated IB4 in combination with immunfluorescence (e.g. LN to label the basement membrane). Different fixation methods of EB-perfused P8 mice to optimize the combination of EB perfusion with IB4 endothelial tip cell labeling and antibody stainings were tested. Combinations of immersion-fixation with 4% FA and different amounts of GA were used. Subsequently, 40 µm coronal brain sections were labeled with biotinylated IB4 to visualize blood vessel endothelial cells (red) and with an antibody against LN (green) to test if antigenicity is still present despite GA treatment (a-l,). EB (perfused blood vessels, cyan). Whereas postfixation in 4% FA only (no GA) resulted in a considerable loss of the EB signal (a-d), the combination of 4% FA + 0.025% GA (e-h) or 4% FA + 0.05% GA (i-l) were optimal for combination of the different label procedures (IB4, LN) with EB-perfused vessels.
Scale bars: 50 µm (a-l).
Supplementary Figure 2 Classical endothelial tip cell markers in direct comparison to IB4 labeling in the P8 mouse brain cortex.
Immunofluorescent labeling of 40 µm P8 coronal mouse brain sections labeled with the classical endothelial tip cells markers VEGFR2 (a-d, green), VEGFR3 (e-h, green) and Dll4 (i-l, green) and with biotinylated IB4 to visualize blood vessel endothelial (tip) cells (red). Cell nuclei (DAPI, blue).
a-l IB4 as well as antibodies against VEGFR2 (a-d), VEGFR3 (e-h) or Dll4 (i-l) visualize CNS blood vessel structures and endothelial tip-, stalk-, and phalanx cells. Boxed areas (a,b,e,f,i,j) highlight endothelial tip cells that are enlarged on the right hand (c,d,g,h,k,l). White arrowheads (a,b,i,j) mark additional endothelial tip cells
c,d,g,h,k,l IB4+ endothelial tip cells with clearly identifiable, multiple filopodia forming a typical “hand-like” structure (c,g,k). Note that neither VEGFR2 (d), VEGFR3 (h) nor Dll4 (l) label endothelial tip cell filopodia as accurately as IB4. As filopodia are the key morphological criterion for identifying endothelial tip cells, the classical tip cell markers VEGFR2, VEGFR3 and Dll4 do not facilitate the identification of endothelial tip cells in the postnatal mouse brain. Moreover, none of these markers allows a clear delineation of endothelial stalk- or phalanx cells from endothelial tip cells (d,h,l).
Scale bars: 50 µm (a,b,e,f,i,j) ; 10 µm (c,d,g,h,k,l).
Supplementary Figure 3 Immunofluorescence of perivascular cells in the vicinity of endothelial tip cells in the P8 mouse brain cortex.
a-p All samples used for the immunofluorescence shown in this figure have been immersion fixed with 4% FA and 0.025% GA, which was the final fixation protocol of the present study. This ensures optimal retaining of EB inside the vessels but also good antigenicity for a variety of cellular markers (see also Supplementary Fig. S1). For example a-d shows GFAP+ astrocytes and GFAP+ radial glia (green), IB4+ blood vessel endothelial cells (red) including an endothelial tip cell and an established blood vessel in the P8 mouse brain cortex. Endothelial tip cell filopodia do not follow a template of GFAP+ astrocytes and radial glia (a). Boxed area with zoom on endothelial tip cell is enlarged in b-d. Cell nuclei (DAPI, blue). e-h PDGFRB+ pericytes (green) and IB4+ blood vessels (red) in the P8 mouse cortex. Boxed area is enlarged in f-h. Endothelial tip cell filopodia are PDGFRB-. Cell nuclei (DAPI, blue). i-l LN+ (green) common basement membrane of IB4+ blood vessels (red) and PDGFRB+ pericytes. Boxed area with zoom on endothelial tip cell is enlarged in j-l. Note the faint LN-staining of endothelial tip cell filopodia at the base of the endothelial tip cell body (l). Cell nuclei (DAPI, blue). Note that no pericytes are present at the tip cell (f,h) whereas LN is ensheathing the tip cell body (j,l). m-p Nf160+ axons (cyan) and IB4+ endothelial tip cell (red, filopodia) in the P8 mouse corpus callosum. Boxed area with zoom on endothelial tip cell is enlarged in n-p. Cell nuclei (DAPI, blue).
Scale bars: 50 µm (a,e,i,m); 10 µm (b-d, f-h, j-l, n-p).
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Supplementary Figures
Supplementary Figures 1–3 (PDF 4937 kb)
Intracardial Evans blue injection of a P8 mouse.
This video shows the different steps of intracardial Evans blue (EB) injection of a P8 mouse pub. All steps are precisely explained in the “PROCEDURE” part of this manuscript. (MOV 9204 kb)
Brain dissection of a P8 mouse.
This video shows the different steps of brain dissection of an Evans-blue (EB) injected P8 mouse pup. All steps are precisely explained in the “PROCEDURE” part of this manuscript. (MOV 12461 kb)
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Wälchli, T., Mateos, J., Weinman, O. et al. Quantitative assessment of angiogenesis, perfused blood vessels and endothelial tip cells in the postnatal mouse brain. Nat Protoc 10, 53–74 (2015). https://doi.org/10.1038/nprot.2015.002
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DOI: https://doi.org/10.1038/nprot.2015.002
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