Skip to main content
PMC home page
Brain Pathology logoLink to Brain Pathology
. 2006 Apr 5;14(3):237–248. doi: 10.1111/j.1750-3639.2004.tb00060.x

Direct Stimulation of Adult Neural Stem Cells In Vitro and Neurogenesis In Vivo by Vascular Endothelial Growth Factor

Anne Schänzer 1,*, Frank‐Peter Wachs 2,*, Daniel Wilhelm 1, Till Acker 1, Christiana Cooper‐Kuhn 2, Heike Beck 1, Jürgen Winkler 2, Ludwig Aigner 2, Karl H Plate 1,*, H Georg Kuhn 2,*,✉
PMCID: PMC8096047  PMID: 15446578

Abstract

Hypoxia as well as global and focal ischemia are strong activators of neurogenesis in the adult mammalian central nervous system. Here we show that the hypoxia‐inducible vascular endothelial growth factor (VEGF) and its receptor VEGFR‐2/Flk‐1 are expressed in clonally‐derived adult rat neural stem cells in vitro. VEGF stimulated the expansion of neural stem cells whereas blockade of VEGFR‐2/Flk‐1‐kinase activity reduced neural stem cell expansion. VEGF was also infused into the lateral ventricle to study changes in neurogenesis in the ventricle wall, olfactory bulb and hippocampus. Using a low dose (2.4 ng/d) to avoid endothelial proliferation and changes in vascular permeability, VEGF stimulated adult neurogenesis in vivo. After VEGF infusion, we observed reduced apoptosis but unaltered proliferation suggesting a survival promoting effect of VEGF in neural progenitor cells. Strong expression of VEGFR‐2/Flk‐1 was detected in the ventricle wall adjacent to the choroid plexus, a site of significant VEGF production, which suggests a paracrine function of endogenous VEGF on neural stem cells in vivo. We propose that VEGF acts as a trophic factor for neural stem cells in vitro and for sustained neurogenesis in the adult nervous system. These findings may have implications for the pathogenesis and therapy of neurodegenerative diseases.

Full Text

The Full Text of this article is available as a PDF (971.0 KB).

REFERENCES

  • 1. Adamis AP, Shima DT, Yeo KT, Yeo TK, Brown LF, Berse B, D'Amore PA, Folkman J (1993) Synthesis and secretion of vascular permeability factor/vascular endothelial growth factor by human retinal pigment epithelial cells. Biochem Biophys Res Commun 193:631–638. [DOI] [PubMed] [Google Scholar]
  • 2. Alvarez‐Buylla A, Seri B, Doetsch F (2002) Identification of neural stem cells in the adult vertebrate brain. Brain Res Bull 57:751–758. [DOI] [PubMed] [Google Scholar]
  • 3. Bagnard D, Vaillant C, Khuth ST, Dufay N, Lohrum M, Puschel AW, Belin MF, Bolz J, Thomasset N (2001) Semaphorin 3A‐vascular endothelial growth factor‐165 balance mediates migration and apoptosis of neural progenitor cells by the recruitment of shared receptor. J Neurosci 21:3332–3341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4. Beck H, Acker T, Wiessner C, Allegrini PR, Plate KH (2000) Expression of angiopoietin‐1, angiopoietin‐2, and tie receptors after middle cerebral artery occlusion in the rat. Am J Pathol 157:1473–1483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5. Bernier PJ, Vinet J, Cossette M, Parent A (2000) Characterization of the subventricular zone of the adult human brain: evidence for the involvement of Bcl‐2. Neurosci Res 37:67–78. [DOI] [PubMed] [Google Scholar]
  • 6. Biebl M, Cooper CM, Winkler J, Kuhn HG (2000) Analysis of neurogenesis and programmed cell death reveals a self‐ renewing capacity in the adult rat brain. Neurosci Lett 291:17–20. [DOI] [PubMed] [Google Scholar]
  • 7. Blaschke AJ, Weiner JA, Chun J (1998) Programmed cell death is a universal feature of embryonic and postnatal neuroproliferative regions throughout the central nervous system. J Comp Neurol 396:39–50. [DOI] [PubMed] [Google Scholar]
  • 8. Breier G, Albrecht U, Sterrer S, Risau W (1992) Expression of vascular endothelial growth factor during embryonic angiogenesis and endothelial cell differentiation. Development 114:521–532. [DOI] [PubMed] [Google Scholar]
  • 9. Carlen M, Cassidy RM, Brismar H, Smith GA, Enquist LW, Frisen J (2002) Functional integration of adult‐born neurons. Curr Biol 12:606–608. [DOI] [PubMed] [Google Scholar]
  • 10. Carmeliet P, Ferreira V, Breier G, Pollefeyt S, Kieckens L, Gertsenstein M, Fahrig M, Vandenhoeck A, Harpal K, Eberhardt C, Declercq C, Pawling J, Moons L, Collen D, Risau W, Nagy A (1996) Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele. Nature 380:435–439. [DOI] [PubMed] [Google Scholar]
  • 11. Carmeliet P (2003) Blood vessels and nerves: common signals, pathways and diseases. Nat Rev Genet 4:710–720. [DOI] [PubMed] [Google Scholar]
  • 12. Chodobski A, Szmydynger‐Chodobska J (2001) Choroid plexus: target for polypeptides and site of their synthesis. Microsc Res Tech 52:65–82. [DOI] [PubMed] [Google Scholar]
  • 13. Cooper‐Kuhn CM, Vroemen M, Brown J, Ye H, Thompson MA, Winkler J, Kuhn HG (2002) Impaired adult neurogenesis in mice lacking the transcription factor E2F1. Mol Cell Neurosci 21:312–323. [DOI] [PubMed] [Google Scholar]
  • 14. Dayer AG, Ford AA, Cleaver KM, Yassaee M, Cameron HA (2003) Short‐term and long‐term survival of new neurons in the rat dentate gyrus. J Comp Neurol 460:563–572. [DOI] [PubMed] [Google Scholar]
  • 15. Fabel K, Tam B, Kaufer D, Baiker A, Simmons N, Kuo CJ, Palmer TD (2003) VEGF is necessary for exercise‐induced adult hippocampal neurogenesis. Eur J Neurosci 18:2803–2812. [DOI] [PubMed] [Google Scholar]
  • 16. Ferrara N, Henzel WJ (1989) Pituitary follicular cells secrete a novel heparin‐binding growth factor specific for vascular endothelial cells. Biochem Biophys Res Commun 161:851–858. [DOI] [PubMed] [Google Scholar]
  • 17. Ferrara N, Carver‐Moore K, Chen H, Dowd M, Lu L, O'Shea KS, Powell‐Braxton L, Hillan KJ, Moore MW (1996) Heterozygous embryonic lethality induced by targeted inactivation of the VEGF gene. Nature 380:439–442. [DOI] [PubMed] [Google Scholar]
  • 18. Fong GH, Rossant J, Gertsenstein M, Breitman ML (1995) Role of the Flt‐1 receptor tyrosine kinase in regulating the assembly of vascular endothelium. Nature 376:66–70. [DOI] [PubMed] [Google Scholar]
  • 19. Haigh JJ, Morelli PI, Gerhardt H, Haigh K, Tsien J, Damert A, Miquerol L, Muhlner U, Klein R, Ferrara N, Wagner EF, Betsholtz C, Nagy A (2003) Cortical and retinal defects caused by dosage‐dependent reductions in VEGF‐A paracrine signaling. Dev Biol 262:225–241. [DOI] [PubMed] [Google Scholar]
  • 20. Harrigan MR, Ennis SR, Sullivan SE, Keep RF (2003) Effects of intraventricular infusion of vascular endothelial growth factor on cerebral blood flow, edema, and infarct volume. Acta Neurochir (Wien) 145:49–53. [DOI] [PubMed] [Google Scholar]
  • 21. Hayashi T, Abe K, Itoyama Y (1998) Reduction of ischemic damage by application of vascular endothelial growth factor in rat brain after transient ischemia. J Cereb Blood Flow Metab 18:887–895. [DOI] [PubMed] [Google Scholar]
  • 22. Hennequin LF, Thomas AP, Johnstone C, Stokes ES, PI inverted question Marke PA, Lohmann JJ, Ogilvie DJ, Dukes M, Wedge SR, Curwen JO, Kendrew J, Lambert‐van der Brempt C (1999) Design and structure‐activity relationship of a new class of potent VEGF receptor tyrosine kinase inhibitors. J Med Chem 42:5369–5389. [DOI] [PubMed] [Google Scholar]
  • 23. Jellinger KA (2003) General aspects of neuro‐degeneration. J Neural Transm Suppl:101–144. [DOI] [PubMed] [Google Scholar]
  • 24. Jin K, Zhu Y, Sun Y, Mao XO, Xie L, Greenberg DA (2002) Vascular endothelial growth factor (VEGF) stimulates neurogenesis in vitro and in vivo. Proc Natl Acad Sci U S A 99:11946–11950. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25. Jin KL, Mao XO, Greenberg DA (2000) Vascular endothelial growth factor: direct neuroprotective effect in in vitro ischemia. Proc Natl Acad Sci U S A 97:10242–10247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26. Johansson CB, Momma S, Clarke DL, Risling M, Lendahl U, Frisen J (1999) Identification of a neural stem cell in the adult mammalian central nervous system. Cell 96:25–34. [DOI] [PubMed] [Google Scholar]
  • 27. Keck PJ, Hauser SD, Krivi G, Sanzo K, Warren T, Feder J, Connolly DT (1989) Vascular permeability factor, an endothelial cell mitogen related to PDGF. Science 246:1309–1312. [DOI] [PubMed] [Google Scholar]
  • 28. Kremer C, Breier G, Risau W, Plate KH (1997) Up‐regulation of flk‐1/vascular endothelial growth factor receptor 2 by its ligand in a cerebral slice culture system. Cancer Res 57:3852–3859. [PubMed] [Google Scholar]
  • 29. Kroll J, Waltenberger J (1997) The vascular endothelial growth factor receptor KDR activates multiple signal transduction pathways in porcine aortic endothelial cells. J Biol Chem 272:32521–32527. [DOI] [PubMed] [Google Scholar]
  • 30. Krum JM, Rosenstein JM (1998) VEGF mRNA and its receptor flt‐1 are expressed in reactive astrocytes following neural grafting and tumor cell implantation in the adult CNS. Exp Neurol 154:57–65. [DOI] [PubMed] [Google Scholar]
  • 31. Krum JM, Mani N, Rosenstein JM (2002) Angiogenic and astroglial responses to vascular endothelial growth factor administration in adult rat brain. Neuroscience 110:589–604. [DOI] [PubMed] [Google Scholar]
  • 32. Kuhn HG, Dickinson‐Anson H, Gage FH (1996) Neurogenesis in the dentate gyrus of the adult rat: age‐related decrease of neuronal progenitor proliferation. J Neurosci 16:2027–2033. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33. Kuhn HG, Winkler J, Kempermann G, Thal LJ, Gage FH (1997) Epidermal growth factor and fibroblast growth factor‐2 have different effects on neural progenitors in the adult rat brain. J Neurosci 17:5820–5829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 34. Kuhn HG, Palmer TD, Fuchs E (2001) Adult neurogenesis: a compensatory mechanism for neuronal damage. Eur Arch Psychiatry Clin Neurosci 251:152–158. [DOI] [PubMed] [Google Scholar]
  • 35. Larrivee B, Lane DR, Pollet I, Olive PL, Humphries RK, Karsan A (2003) VEGFR‐2 induces survival of hematopoietic progenitor cells. J Biol Chem 278:22006–22013. [DOI] [PubMed] [Google Scholar]
  • 36. Lennmyr F, Ata KA, Funa K, Olsson Y, Terent A (1998) Expression of vascular endothelial growth factor (VEGF) and its receptors (Flt‐1 and Flk‐1) following permanent and transient occlusion of the middle cerebral artery in the rat. J Neuropathol Exp Neurol 57:874–882. [DOI] [PubMed] [Google Scholar]
  • 37. Leung DW, Cachianes G, Kuang WJ, Goeddel DV, Ferrara N (1989) Vascular endothelial growth factor is a secreted angiogenic mitogen. Science 246:1306–1309. [DOI] [PubMed] [Google Scholar]
  • 38. Louissaint A, Jr. , Rao S, Leventhal C, Goldman SA (2002) Coordinated interaction of neurogenesis and angiogenesis in the adult songbird brain. Neuron 34:945–960. [DOI] [PubMed] [Google Scholar]
  • 39. Marti HH, Risau W (1998) Systemic hypoxia changes the organ‐specific distribution of vascular endothelial growth factor and its receptors. Proc Natl Acad Sci U S A 95:15809–15814. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 40. Matthews W, Jordan CT, Gavin M, Jenkins NA, Copeland NG, Lemischka IR (1991) A receptor tyrosine kinase cDNA isolated from a population of enriched primitive hematopoietic cells and exhibiting close genetic linkage to c‐ kit. Proc Natl Acad Sci U S A 88:9026–9030. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 41. Maurer MH, Tripps WK, Feldmann RE, Jr. , Kuschinsky W (2003) Expression of vascular endothelial growth factor and its receptors in rat neural stem cells. Neurosci Lett 344:165–168. [DOI] [PubMed] [Google Scholar]
  • 42. Mercier F, Kitasako JT, Hatton GI (2002) Anatomy of the brain neurogenic zones revisited: fractones and the fibroblast/macrophage network. J Comp Neurol 451:170–188. [DOI] [PubMed] [Google Scholar]
  • 43. Naruse I, Keino H (1995) Apoptosis in the developing CNS. Prog Neurobiol 47:135–155. [DOI] [PubMed] [Google Scholar]
  • 44. Ogunshola OO, Stewart WB, Mihalcik V, Solli T, Madri JA, Ment LR (2000) Neuronal VEGF expression correlates with angiogenesis in postnatal developing rat brain. Brain Res Dev Brain Res 119:139–153. [DOI] [PubMed] [Google Scholar]
  • 45. Ogunshola OO, Antic A, Donoghue MJ, Fan SY, Kim H, Stewart WB, Madri JA, Ment LR (2002) Paracrine and autocrine functions of neuronal VEGF in the CNS. J Biol Chem 277:11410–11415. [DOI] [PubMed] [Google Scholar]
  • 46. Oosthuyse B, Moons L, Storkebaum E, Beck H, Nuyens D, Brusselmans K, van Dorpe J, Hellings P, Gorselink M, Heymans S, Theilmeier G, Dewerchin M, et al (2001) Deletion of the hypoxia‐response element in the vascular endothelial growth factor promoter causes motor neuron degeneration. Nat Genet 28:131–138. [DOI] [PubMed] [Google Scholar]
  • 47. Palmer TD, Ray J, Gage FH (1995) FGF‐2‐responsive neuronal progenitors reside in proliferative and quiescent regions of the adult rodent bra in. Mol Cell Neurosci 6:474–486. [DOI] [PubMed] [Google Scholar]
  • 48. Palmer TD, Willhoite AR, Gage FH (2000) Vascular niche for adult hippocampal neurogenesis. J Comp Neurol 425:479–494. [DOI] [PubMed] [Google Scholar]
  • 49. Patt S, Danner S, Theallier‐Janko A, Breier G, Hottenrott G, Plate KH, Cervos‐Navarro J (1998) Upregulation of vascular endothelial growth factor in severe chronic brain hypoxia of the rat. Neurosci Lett 252:199–202. [DOI] [PubMed] [Google Scholar]
  • 50. Pelosi E, Valtieri M, Coppola S, Botta R, Gabbianelli M, Lulli V, Marziali G, Masella B, Muller R, Sgadari C, Testa U, Bonanno G, Peschle C (2002) Identification of the hemangioblast in postnatal life. Blood 100:3203–3208. [DOI] [PubMed] [Google Scholar]
  • 51. Plate KH, Breier G, Weich HA, Risau W (1992) Vascular endothelial growth factor is a potential tumour angiogenesis factor in human gliomas in vivo. Nature 359:845–848. [DOI] [PubMed] [Google Scholar]
  • 52. Plate KH, Beck H, Danner S, Allegrini PR, Wiessner C (1999) Cell type specific upregulation of vascular endothelial growth factor in an MCA‐occlusion model of cerebral infarct. J Neuropathol Exp Neurol 58:654–666. [DOI] [PubMed] [Google Scholar]
  • 53. Pong K (2003) Oxidative stress in neurode‐generative diseases: therapeutic implications for superoxide dismutase mimetics. Expert Opin Biol Ther 3:127–139. [DOI] [PubMed] [Google Scholar]
  • 54. Raff MC, Barres BA, Burne JF, Coles HS, Ishizaki Y, Jacobson MD (1993) Programmed cell death and the control of cell survival: lessons from the nervous system. Science 262:695–700. [DOI] [PubMed] [Google Scholar]
  • 55. Rafii S, Avecilla S, Shmelkov S, Shido K, Tejada R, Moore MA, Heissig B, Hattori K (2003) Angiogenic factors reconstitute hematopoiesis by recruiting stem cells from bone marrow microenvironment. Ann N Y Acad Sci 996:49–60. [DOI] [PubMed] [Google Scholar]
  • 56. Reynolds BA, Weiss S (1992) Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 255:1707–1710. [DOI] [PubMed] [Google Scholar]
  • 57. Rivard A, Berthou‐Soulie L, Principe N, Kearney M, Curry C, Branellec D, Semenza GL, Isner JM (2000) Age‐dependent defect in vascular endothelial growth factor expression is associated with reduced hypoxia‐inducible factor 1 activity. J Biol Chem 275:29643–29647. [DOI] [PubMed] [Google Scholar]
  • 58. Schratzberger P, Schratzberger G, Silver M, Curry C, Kearney M, Magner M, Alroy J, Adelman LS, Weinberg DH, Ropper AH, Isner JM (2000) Favorable effect of VEGF gene transfer on ischemic peripheral neuropathy. Nat Med 6:405–413. [DOI] [PubMed] [Google Scholar]
  • 59. Senger DR, Perruzzi CA, Feder J, Dvorak HF (1986) A highly conserved vascular permeability factor secreted by a variety of human and rodent tumor cell lines. Cancer Res 46:5629–5632. [PubMed] [Google Scholar]
  • 60. Shalaby F, Rossant J, Yamaguchi TP, Gertsenstein M, Wu XF, Breitman ML, Schuh AC (1995) Failure of blood‐island formation and vasculogenesis in Flk‐1‐deficient mice. Nature 376:62–66. [DOI] [PubMed] [Google Scholar]
  • 61. Shingo T, Sorokan ST, Shimazaki T, Weiss S (2001) Erythropoietin regulates the in vitro and in vivo production of neuronal progenitors by mammalian forebrain neural stem cells. J Neurosci 21:9733–9743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 62. Shweiki D, Itin A, Soffer D, Keshet E (1992) Vascular endothelial growth factor induced by hypoxia may mediate hypoxia‐initiated angiogenesis. Nature 359:843–845. [DOI] [PubMed] [Google Scholar]
  • 63. Stopa EG, Berzin TM, Kim S, Song P, Kuo‐LeBlanc V, Rodriguez‐Wolf M, Baird A, Johanson CE (2001) Human choroid plexus growth factors: What are the implications for CSF dynamics in Alzheimer's disease Exp Neurol 167:40–47. [DOI] [PubMed] [Google Scholar]
  • 64. Strawn LM, McMahon G, App H, Schreck R, Kuchler WR, Longhi MP, Hui TH, Tang C, Levitzki A, Gazit A, Chen I, Keri G, Orfi L, Risau W, Flamme I, Ullrich A, Hirth KP, Shawver LK (1996) Flk‐1 as a target for tumor growth inhibition. Cancer Res 56:3540–3545. [PubMed] [Google Scholar]
  • 65. Studer L, Csete M, Lee SH, Kabbani N, Walikonis J, Wold B, McKay R (2000) Enhanced proliferation, survival, and dopaminergic differentiation of CNS precursors in lowered oxygen. J Neurosci 20:7377–7383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 66. Sun Y, Jin K, Xie L, Childs J, Mao XO, Logvinova A, Greenberg DA (2003) VEGF‐induced neuroprotection, neurogenesis, and angiogenesis after focal cerebral ischemia. J Clin Invest 111:1843–1851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 67. Uchida K, Uchida S, Nitta K, Yumura W, Marumo F, Nihei H (1994) Glomerular endothelial cells in culture express and secrete vascular endothelial growth factor. Am J Physiol 266:F81–88. [DOI] [PubMed] [Google Scholar]
  • 68. van Praag H, Schinder AF, Christie BR, Toni N, Palmer TD, Gage FH (2002) Functional neurogenesis in the adult hippocampus. Nature 415:1030–1034. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 69. Wachs FP, Couillard‐Despres S, Engelhardt M, Wilhelm D, Plötz S, Vroemen M, Klucken J, Karl C, Tebbing J, Svendsen C, Weidner N, Kuhn HG, Winkler J, Aigner L (2003) High efficacy of clonal growth and expansion of adult neural stem cells. Lab Invest 7:949–962. [DOI] [PubMed] [Google Scholar]
  • 70. Williams RW, Rakic P (1988) Three‐dimensional counting: an accurate and direct method to estimate numbers of cells in sectioned material. J Comp Neurol 278:344–352. [DOI] [PubMed] [Google Scholar]
  • 71. Winner B, Cooper‐Kuhn CM, Aigner R, Winkler J, Kuhn HG (2002) Long‐term survival and cell death of newly generated neurons in the adult rat olfactory bulb. Eur J Neurosci 16:1681–1689. [DOI] [PubMed] [Google Scholar]
  • 72. Yang K, Cepko CL (1996) Flk‐1, a receptor for vascular endothelial growth factor (VEGF), is expressed by retinal progenitor cells. J Neurosci 16:6089–6099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 73. Yu X, Shacka JJ, Eells JB, Suarez‐Quian C, Przygodzki RM, Beleslin‐Cokic B, Lin CS, Nikodem VM, Hempstead B, Flanders KC, Costantini F, Noguchi CT (2002) Erythropoietin receptor signalling is required for normal brain development. Development 129:505–516. [DOI] [PubMed] [Google Scholar]
  • 74. Zhu Y, Jin K, Mao XO, Greenberg DA (2003) Vascular endothelial growth factor promotes proliferation of cortical neuron precursors by regulating E2F expression. Faseb J 17:186–193. [DOI] [PubMed] [Google Scholar]
  • 75. Ziegler BL, Valtieri M, Porada GA, De Maria R, Muller R, Masella B, Gabbianelli M, Casella I, Pelosi E, Bock T, Zanjani ED, Peschle C (1999) KDR receptor: a key marker defining hematopoietic stem cells. Science 285:1553–1558. [DOI] [PubMed] [Google Scholar]

Articles from Brain Pathology are provided here courtesy of Wiley

RESOURCES