Skip to main content
PMC home page
Journal of Anatomy logoLink to Journal of Anatomy
. 2000 Nov;197(Pt 4):591–605. doi: 10.1046/j.1469-7580.2000.19740591.x

VEGF enhances intraneural angiogenesis and improves nerve regeneration after axotomy

MARK I HOBSON 1, COLIN J GREEN 1, GIORGIO TERENGHI 1,✉
PMCID: PMC1468175  PMID: 11197533

Abstract

Whilst there is an increased understanding of the cell biology of nerve regeneration, it remains unclear whether there is a direct interrelationship between vascularisation and efficacy of nerve regeneration within a nerve conduit. To establish this is important as in clinical surgery peripheral nerve conduit grafting has been widely investigated as a possible alternative to the use of nerve autografts. The aim of this study was to assess whether vascular endothelial growth factor (VEGF), a highly specific endothelial cell mitogen, can enhance vascularisation and, indirectly, axonal regeneration within a silicone nerve regeneration chamber. Chambers containing VEGF (500–700 ng/ml) in a laminin-based gel (Matrigel) were inserted into 1 cm rat sciatic nerve defects and nerve regeneration examined in relation to angiogenesis between 5 and 180 d. Longitudinal sections were stained with antibodies against endothelial cells (RECA-1), axons (neurofilament) and Schwann cells (S-100) to follow the progression of vascular and neural elements. Computerised image analysis demonstrated that the addition of VEGF significantly increased blood vessel penetration within the chamber from d 5, and by d 10 this correlated with an increase of axonal regeneration and Schwann cell migration. The pattern of increased nerve regeneration due to VEGF administration was maintained up to 180 d, when myelinated axon counts were increased by 78% compared with plain Matrigel control. Furthermore the dose-response of blood vessel regeneration to VEGF was clearly reflected in the increase of axonal regrowth and Schwann cell proliferation, indicating the close relationship between regenerating nerves and blood vessels within the chamber. Target organ reinnervation was enhanced by VEGF at 180 d as measured through the recovery of gastrocnemius muscle weights and footpad axonal terminal density, the latter showing a significant increase over controls (P < 0.05). The results demonstrate an overall relationship between increased vascularisation and enhanced nerve regeneration within an acellular conduit, and highlight the interdependence of the 2 processes.

Keywords: Peripheral nerve, vasculature

Full Text

The Full Text of this article is available as a PDF (563.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Angiolillo A. L., Sgadari C., Taub D. D., Liao F., Farber J. M., Maheshwari S., Kleinman H. K., Reaman G. H., Tosato G. Human interferon-inducible protein 10 is a potent inhibitor of angiogenesis in vivo. J Exp Med. 1995 Jul 1;182(1):155–162. doi: 10.1084/jem.182.1.155. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bell M. A., Weddell A. G. A descriptive study of the blood vessels of the sciatic nerve in the rat, man and other mammals. Brain. 1984 Sep;107(Pt 3):871–898. doi: 10.1093/brain/107.3.871. [DOI] [PubMed] [Google Scholar]
  3. Best T. J., Mackinnon S. E. Peripheral nerve revascularization: a current literature review. J Reconstr Microsurg. 1994 May;10(3):193–204. doi: 10.1055/s-2007-1006587. [DOI] [PubMed] [Google Scholar]
  4. Bissell D. M., Arenson D. M., Maher J. J., Roll F. J. Support of cultured hepatocytes by a laminin-rich gel. Evidence for a functionally significant subendothelial matrix in normal rat liver. J Clin Invest. 1987 Mar;79(3):801–812. doi: 10.1172/JCI112887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Clauss M., Gerlach M., Gerlach H., Brett J., Wang F., Familletti P. C., Pan Y. C., Olander J. V., Connolly D. T., Stern D. Vascular permeability factor: a tumor-derived polypeptide that induces endothelial cell and monocyte procoagulant activity, and promotes monocyte migration. J Exp Med. 1990 Dec 1;172(6):1535–1545. doi: 10.1084/jem.172.6.1535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Connolly D. T., Heuvelman D. M., Nelson R., Olander J. V., Eppley B. L., Delfino J. J., Siegel N. R., Leimgruber R. M., Feder J. Tumor vascular permeability factor stimulates endothelial cell growth and angiogenesis. J Clin Invest. 1989 Nov;84(5):1470–1478. doi: 10.1172/JCI114322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Devor M., Keller C. H., Ellisman M. H. Spontaneous discharge of afferents in a neuroma reflects original receptor tuning. Brain Res. 1990 May 28;517(1-2):245–250. doi: 10.1016/0006-8993(90)91033-d. [DOI] [PubMed] [Google Scholar]
  8. Fawcett J. W., Keynes R. J. Peripheral nerve regeneration. Annu Rev Neurosci. 1990;13:43–60. doi: 10.1146/annurev.ne.13.030190.000355. [DOI] [PubMed] [Google Scholar]
  9. Fields R. D., Le Beau J. M., Longo F. M., Ellisman M. H. Nerve regeneration through artificial tubular implants. Prog Neurobiol. 1989;33(2):87–134. doi: 10.1016/0301-0082(89)90036-1. [DOI] [PubMed] [Google Scholar]
  10. Gitay-Goren H., Soker S., Vlodavsky I., Neufeld G. The binding of vascular endothelial growth factor to its receptors is dependent on cell surface-associated heparin-like molecules. J Biol Chem. 1992 Mar 25;267(9):6093–6098. [PubMed] [Google Scholar]
  11. Grant D. S., Kleinman H. K., Goldberg I. D., Bhargava M. M., Nickoloff B. J., Kinsella J. L., Polverini P., Rosen E. M. Scatter factor induces blood vessel formation in vivo. Proc Natl Acad Sci U S A. 1993 Mar 1;90(5):1937–1941. doi: 10.1073/pnas.90.5.1937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hobson M. I., Brown R., Green C. J., Terenghi G. Inter-relationships between angiogenesis and nerve regeneration: a histochemical study. Br J Plast Surg. 1997 Feb;50(2):125–131. doi: 10.1016/s0007-1226(97)91325-4. [DOI] [PubMed] [Google Scholar]
  13. Hu D. E., Fan T. P. Suppression of VEGF-induced angiogenesis by the protein tyrosine kinase inhibitor, lavendustin A. Br J Pharmacol. 1995 Jan;114(2):262–268. doi: 10.1111/j.1476-5381.1995.tb13221.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jabaley M. E., Burns J. E., Orcutt B. S., Bryant M. Comparison of histologic and functional recovery after peripheral nerve repair. J Hand Surg Am. 1976 Sep;1(2):119–130. doi: 10.1016/s0363-5023(76)80005-6. [DOI] [PubMed] [Google Scholar]
  15. Kanaya F., Firrell J. C., Breidenbach W. C. Sciatic function index, nerve conduction tests, muscle contraction, and axon morphometry as indicators of regeneration. Plast Reconstr Surg. 1996 Dec;98(7):1264-71, discussion 1272-4. doi: 10.1097/00006534-199612000-00023. [DOI] [PubMed] [Google Scholar]
  16. Kleinman H. K., McGarvey M. L., Liotta L. A., Robey P. G., Tryggvason K., Martin G. R. Isolation and characterization of type IV procollagen, laminin, and heparan sulfate proteoglycan from the EHS sarcoma. Biochemistry. 1982 Nov 23;21(24):6188–6193. doi: 10.1021/bi00267a025. [DOI] [PubMed] [Google Scholar]
  17. Le Beau J. M., Ellisman M. H., Powell H. C. Ultrastructural and morphometric analysis of long-term peripheral nerve regeneration through silicone tubes. J Neurocytol. 1988 Apr;17(2):161–172. doi: 10.1007/BF01674203. [DOI] [PubMed] [Google Scholar]
  18. Levy A. P., Tamargo R., Brem H., Nathans D. An endothelial cell growth factor from the mouse neuroblastoma cell line NB41. Growth Factors. 1989;2(1):9–19. doi: 10.3109/08977198909069077. [DOI] [PubMed] [Google Scholar]
  19. Lundborg G. Structure and function of the intraneural microvessels as related to trauma, edema formation, and nerve function. J Bone Joint Surg Am. 1975 Oct;57(7):938–948. [PubMed] [Google Scholar]
  20. Madison R. D., da Silva C., Dikkes P., Sidman R. L., Chiu T. H. Peripheral nerve regeneration with entubulation repair: comparison of biodegradeable nerve guides versus polyethylene tubes and the effects of a laminin-containing gel. Exp Neurol. 1987 Feb;95(2):378–390. doi: 10.1016/0014-4886(87)90146-4. [DOI] [PubMed] [Google Scholar]
  21. Madison R., da Silva C. F., Dikkes P., Chiu T. H., Sidman R. L. Increased rate of peripheral nerve regeneration using bioresorbable nerve guides and a laminin-containing gel. Exp Neurol. 1985 Jun;88(3):767–772. doi: 10.1016/0014-4886(85)90087-1. [DOI] [PubMed] [Google Scholar]
  22. Mellick R. S., Cavanagh J. B. Changes in blood vessel permeability during degeneration and regeneration in peripheral nerves. Brain. 1968 Mar;91(1):141–160. doi: 10.1093/brain/91.1.141. [DOI] [PubMed] [Google Scholar]
  23. Millauer B., Wizigmann-Voos S., Schnürch H., Martinez R., Møller N. P., Risau W., Ullrich A. High affinity VEGF binding and developmental expression suggest Flk-1 as a major regulator of vasculogenesis and angiogenesis. Cell. 1993 Mar 26;72(6):835–846. doi: 10.1016/0092-8674(93)90573-9. [DOI] [PubMed] [Google Scholar]
  24. Müller H., Williams L. R., Varon S. Nerve regeneration chamber: evaluation of exogenous agents applied by multiple injections. Brain Res. 1987 Jun 16;413(2):320–326. doi: 10.1016/0006-8993(87)91023-7. [DOI] [PubMed] [Google Scholar]
  25. Nicosia R. F., Nicosia S. V., Smith M. Vascular endothelial growth factor, platelet-derived growth factor, and insulin-like growth factor-1 promote rat aortic angiogenesis in vitro. Am J Pathol. 1994 Nov;145(5):1023–1029. [PMC free article] [PubMed] [Google Scholar]
  26. Nicosia R. F., Ottinetti A. Modulation of microvascular growth and morphogenesis by reconstituted basement membrane gel in three-dimensional cultures of rat aorta: a comparative study of angiogenesis in matrigel, collagen, fibrin, and plasma clot. In Vitro Cell Dev Biol. 1990 Feb;26(2):119–128. doi: 10.1007/BF02624102. [DOI] [PubMed] [Google Scholar]
  27. Norrby K. Vascular endothelial growth factor and de novo mammalian angiogenesis. Microvasc Res. 1996 Mar;51(2):153–163. doi: 10.1006/mvre.1996.0017. [DOI] [PubMed] [Google Scholar]
  28. Nukada H. Post-traumatic endoneurial neovascularization and nerve regeneration: a morphometric study. Brain Res. 1988 May 24;449(1-2):89–96. doi: 10.1016/0006-8993(88)91027-x. [DOI] [PubMed] [Google Scholar]
  29. Passaniti A., Taylor R. M., Pili R., Guo Y., Long P. V., Haney J. A., Pauly R. R., Grant D. S., Martin G. R. A simple, quantitative method for assessing angiogenesis and antiangiogenic agents using reconstituted basement membrane, heparin, and fibroblast growth factor. Lab Invest. 1992 Oct;67(4):519–528. [PubMed] [Google Scholar]
  30. Podhajsky R. J., Myers R. R. A diffusion-reaction model of nerve regeneration. J Neurosci Methods. 1995 Aug;60(1-2):79–88. doi: 10.1016/0165-0270(94)00222-3. [DOI] [PubMed] [Google Scholar]
  31. Podhajsky R. J., Myers R. R. The vascular response to nerve crush: relationship to Wallerian degeneration and regeneration. Brain Res. 1993 Sep 24;623(1):117–123. doi: 10.1016/0006-8993(93)90018-i. [DOI] [PubMed] [Google Scholar]
  32. Podhajsky R. J., Myers R. R. The vascular response to nerve transection: neovascularization in the silicone nerve regeneration chamber. Brain Res. 1994 Oct 31;662(1-2):88–94. doi: 10.1016/0006-8993(94)90799-4. [DOI] [PubMed] [Google Scholar]
  33. Rosen E. M., Goldberg I. D. Scatter factor and angiogenesis. Adv Cancer Res. 1995;67:257–279. doi: 10.1016/s0065-230x(08)60715-0. [DOI] [PubMed] [Google Scholar]
  34. Seetharam L., Gotoh N., Maru Y., Neufeld G., Yamaguchi S., Shibuya M. A unique signal transduction from FLT tyrosine kinase, a receptor for vascular endothelial growth factor VEGF. Oncogene. 1995 Jan 5;10(1):135–147. [PubMed] [Google Scholar]
  35. Shen H., Clauss M., Ryan J., Schmidt A. M., Tijburg P., Borden L., Connolly D., Stern D., Kao J. Characterization of vascular permeability factor/vascular endothelial growth factor receptors on mononuclear phagocytes. Blood. 1993 May 15;81(10):2767–2773. [PubMed] [Google Scholar]
  36. Smith J. W. Factors influencing nerve repair. I. Blood supply of peripheral nerves. Arch Surg. 1966 Aug;93(2):335–341. doi: 10.1001/archsurg.1966.01330020127022. [DOI] [PubMed] [Google Scholar]
  37. Smith J. W. Factors influencing nerve repair. II. Collateral circulation of peripheral nerves. Arch Surg. 1966 Sep;93(3):433–437. doi: 10.1001/archsurg.1966.01330030063014. [DOI] [PubMed] [Google Scholar]
  38. Sondell M., Lundborg G., Kanje M. Vascular endothelial growth factor has neurotrophic activity and stimulates axonal outgrowth, enhancing cell survival and Schwann cell proliferation in the peripheral nervous system. J Neurosci. 1999 Jul 15;19(14):5731–5740. doi: 10.1523/JNEUROSCI.19-14-05731.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Sparrow J. R., Kiernan J. A. Endoneurial vascular permeability in degenerating and regenerating peripheral nerves. Acta Neuropathol. 1981;53(3):181–188. doi: 10.1007/BF00688020. [DOI] [PubMed] [Google Scholar]
  40. Thomas K. A. Vascular endothelial growth factor, a potent and selective angiogenic agent. J Biol Chem. 1996 Jan 12;271(2):603–606. doi: 10.1074/jbc.271.2.603. [DOI] [PubMed] [Google Scholar]
  41. Van Beek A. L., Jacobs S. C., Zook E. G. Examination of peripheral nerves with the scanning electron microscope. Plast Reconstr Surg. 1979 Apr;63(4):509–519. doi: 10.1097/00006534-197904000-00012. [DOI] [PubMed] [Google Scholar]
  42. Weddell G. Axonal regeneration in cutaneous nerve plexuses. J Anat. 1942 Oct;77(Pt 1):49–62.3. doi: 10.1093/oxfordjournals.bmb.a070231. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Weerasuriya A. Patterns of change in endoneurial capillary permeability and vascular space during Wallerian degeneration. Brain Res. 1988 Mar 29;445(1):181–187. doi: 10.1016/0006-8993(88)91091-8. [DOI] [PubMed] [Google Scholar]
  44. Weerasuriya A. Patterns of change in endoneurial capillary permeability and vascular space during nerve regeneration. Brain Res. 1990 Feb 26;510(1):135–139. doi: 10.1016/0006-8993(90)90739-x. [DOI] [PubMed] [Google Scholar]
  45. Williams L. R., Longo F. M., Powell H. C., Lundborg G., Varon S. Spatial-temporal progress of peripheral nerve regeneration within a silicone chamber: parameters for a bioassay. J Comp Neurol. 1983 Aug 20;218(4):460–470. doi: 10.1002/cne.902180409. [DOI] [PubMed] [Google Scholar]
  46. Williams L. R., Powell H. C., Lundborg G., Varon S. Competence of nerve tissue as distal insert promoting nerve regeneration in a silicone chamber. Brain Res. 1984 Feb 20;293(2):201–211. doi: 10.1016/0006-8993(84)91227-7. [DOI] [PubMed] [Google Scholar]
  47. Williams L. R., Varon S. Modification of fibrin matrix formation in situ enhances nerve regeneration in silicone chambers. J Comp Neurol. 1985 Jan 8;231(2):209–220. doi: 10.1002/cne.902310208. [DOI] [PubMed] [Google Scholar]
  48. de Vries C., Escobedo J. A., Ueno H., Houck K., Ferrara N., Williams L. T. The fms-like tyrosine kinase, a receptor for vascular endothelial growth factor. Science. 1992 Feb 21;255(5047):989–991. doi: 10.1126/science.1312256. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Anatomy are provided here courtesy of Anatomical Society of Great Britain and Ireland

RESOURCES