Advertisement

Redirection of Neurite Outgrowth by Coupling Chondroitin Sulfate Proteoglycans to Polymer Membranes

  • 464 Accesses

  • 7 Citations

Abstract

Upon nerve injury, the body creates an environment consisting of permissive and non-permissive cues that instruct the function of cells involved in nerve repair. Among other roles, the developing extracellular matrix (ECM) acts as an underlying substrate to guide the union of neurites extending from the proximal stump for bridging the nerve gap. Chondroitin sulfate proteoglycans (CSPGs) are present in the nerve ECM and inhibit axon growth, potentially providing molecular cues to prevent aberrant growth and direct regeneration. In this study, we examined the potential of CSPGs to guide dorsal root ganglia (DRG) neurite outgrowth when freely available in the media or presented from a polymeric membrane. Soluble CSPGs added to the media of DRG explant cultures inhibited neurite outgrowth without spatial bias, caused retraction of axons, and decreased neurite extension in a dose-dependent manner. Poly-l-lactic acid membranes were chemically treated to enhance adsorption of CSPGs to the surface. CSPGs bound to 1,6-hexanediamine-treated membranes directed the orientation of neurite outgrowth, as neurites avoided bound CSPGs and a higher number and percentage grew on treated membranes lacking CSPGs. DRG explants cultured on CSPG-coated membranes without 1,6-hexanediamine-treatment had a smaller number of neurites and decreased neurite outgrowth, suggesting CSPGs were not retained on the membrane and were released into the culture medium. Taken together, these data demonstrate the potential of CSPG presentation to guide axonal growth. This approach offers a strategy to improve upon existing nerve guidance conduits by incorporating axon guidance molecules to direct nerve regeneration.

This is a preview of subscription content, log in to check access.

Access options

Buy single article

Instant unlimited access to the full article PDF.

US$ 39.95

Price includes VAT for USA

Subscribe to journal

Immediate online access to all issues from 2019. Subscription will auto renew annually.

US$ 199

This is the net price. Taxes to be calculated in checkout.

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

References

  1. 1.

    Aframian, D. J., R. S. Redman, S. Yamano, J. Nikolovski, E. Cukierman, K. M. Yamada, M. F. Kriete, W. D. Swaim, D. J. Mooney, and B. J. Baum. Tissue compatibility of two biodegradable tubular scaffolds implanted adjacent to skin or buccal mucosa in mice. Tissue Eng. 8(4):649–659, 2002.

  2. 2.

    Ara, J., P. Bannerman, A. Hahn, S. Ramirez, and D. Pleasure. Modulation of sciatic nerve expression of class 3 semaphorins by nerve injury. Neurochem. Res. 29(6):1153–1159, 2004.

  3. 3.

    Bertelli, J. A., M. Taleb, J. C. Mira, and M. F. Ghizoni. Variation in nerve autograft length increases fibre misdirection and decreases pruning effectiveness: an experimental study in the rat median nerve. Neurol. Res. 27(6):657–665, 2005.

  4. 4.

    Braunewell, K. H., R. Martini, R. LeBaron, H. Kresse, A. Faissner, B. Schmitz, and M. Schachner. Up-regulation of a chondroitin sulphate epitope during regeneration of mouse sciatic nerve: evidence that the immunoreactive molecules are related to the chondroitin sulphate proteoglycans decorin and versican. Eur. J. Neurosci. 7(4):792–804, 1995.

  5. 5.

    Ceballos, D., X. Navarro, N. Dubey, G. Wendelschafer-Crabb, W. R. Kennedy, and R. T. Tranquillo. Magnetically aligned collagen gel filling a collagen nerve guide improves peripheral nerve regeneration. Exp. Neurol. 158(2):290–300, 1999.

  6. 6.

    Chernousov, M. A., and D. J. Carey. Schwann cell extracellular matrix molecules and their receptors. Histol. Histopathol. 15(2):593–601, 2000.

  7. 7.

    Choi, D., and G. Raisman. After facial nerve damage, regenerating axons become aberrant throughout the length of the nerve and not only at the site of the lesion: an experimental study. Br. J. Neurosurg. 18(1):45–48, 2004.

  8. 8.

    Clement, A. M., K. Sugahara, and A. Faissner. Chondroitin sulfate E promotes neurite outgrowth of rat embryonic day 18 hippocampal neurons. Neurosci. Lett. 269(3):125–128, 1999.

  9. 9.

    Davies, J. E., X. Tang, J. W. Denning, S. J. Archibald, and S. J. Davies. Decorin suppresses neurocan, brevican, phosphacan and NG2 expression and promotes axon growth across adult rat spinal cord injuries. Eur. J. Neurosci. 19(5):1226–1242, 2004.

  10. 10.

    de Ruiter, G. C., M. J. Malessy, A. O. Alaid, R. J. Spinner, J. K. Engelstad, E. J. Sorenson, K. R. Kaufman, P. J. Dyck, and A. J. Windebank. Misdirection of regenerating motor axons after nerve injury and repair in the rat sciatic nerve model. Exp. Neurol. 211(2):339–350, 2008.

  11. 11.

    Dubey, N., P. C. Letourneau, and R. T. Tranquillo. Neuronal contact guidance in magnetically aligned fibrin gels: effect of variation in gel mechano-structural properties. Biomaterials 22(10):1065–1075, 2001.

  12. 12.

    Evans, G. R., K. Brandt, S. Katz, P. Chauvin, L. Otto, M. Bogle, B. Wang, R. K. Meszlenyi, L. Lu, A. G. Mikos, et al. Bioactive poly(l-lactic acid) conduits seeded with Schwann cells for peripheral nerve regeneration. Biomaterials 23(3):841–848, 2002.

  13. 13.

    Evans, G. R., K. Brandt, M. S. Widmer, L. Lu, R. K. Meszlenyi, P. K. Gupta, A. G. Mikos, J. Hodges, J. Williams, A. Gurlek, et al. In vivo evaluation of poly(l-lactic acid) porous conduits for peripheral nerve regeneration. Biomaterials 20(12):1109–1115, 1999.

  14. 14.

    Fernaud-Espinosa, I., M. Nieto-Sampedro, and P. Bovolenta. Differential effects of glycosaminoglycans on neurite outgrowth from hippocampal and thalamic neurones. J. Cell Sci. 107(Pt 6):1437–1448, 1994.

  15. 15.

    Funakoshi, H., J. Frisen, G. Barbany, T. Timmusk, O. Zachrisson, V. M. Verge, and H. Persson. Differential expression of mRNAs for neurotrophins and their receptors after axotomy of the sciatic nerve. J. Cell Biol. 123(2):455–465, 1993.

  16. 16.

    Giger, R. J., J. F. Cloutier, A. Sahay, R. K. Prinjha, D. V. Levengood, S. E. Moore, S. Pickering, D. Simmons, S. Rastan, F. S. Walsh, et al. Neuropilin-2 is required in vivo for selective axon guidance responses to secreted semaphorins. Neuron 25(1):29–41, 2000.

  17. 17.

    Hammarberg, H., F. Piehl, S. Cullheim, J. Fjell, T. Hokfelt, and K. Fried. GDNF mRNA in Schwann cells and DRG satellite cells after chronic sciatic nerve injury. NeuroReport 7(4):857–860, 1996.

  18. 18.

    Hawkins, R. L., and N. W. Seeds. Protease inhibitors influence the direction of neurite outgrowth. Brain Res. Dev. Brain Res. 45(2):203–209, 1989.

  19. 19.

    He, Z., and M. Tessier-Lavigne. Neuropilin is a receptor for the axonal chemorepellent Semaphorin III. Cell 90(4):739–751, 1997.

  20. 20.

    Heumann, R., D. Lindholm, C. Bandtlow, M. Meyer, M. J. Radeke, T. P. Misko, E. Shooter, and H. Thoenen. Differential regulation of mRNA encoding nerve growth factor and its receptor in rat sciatic nerve during development, degeneration, and regeneration: role of macrophages. Proc. Natl. Acad. Sci. USA 84(23):8735–8739, 1987.

  21. 21.

    Hoke, A., C. Cheng, and D. W. Zochodne. Expression of glial cell line-derived neurotrophic factor family of growth factors in peripheral nerve injury in rats. NeuroReport 11(8):1651–1654, 2000.

  22. 22.

    Jones, L. L., D. Sajed, and M. H. Tuszynski. Axonal regeneration through regions of chondroitin sulfate proteoglycan deposition after spinal cord injury: a balance of permissiveness and inhibition. J. Neurosci. 23(28):9276–9288, 2003.

  23. 23.

    Kofron, C. M., V. J. Fong, and D. Hoffman-Kim. Neurite outgrowth at the interface of 2D and 3D growth environments. J. Neural Eng. 6(1):016002, 2009.

  24. 24.

    Lindholm, T., M. K. Skold, A. Suneson, T. Carlstedt, S. Cullheim, and M. Risling. Semaphorin and neuropilin expression in motoneurons after intraspinal motoneuron axotomy. NeuroReport 15(4):649–654, 2004.

  25. 25.

    Lubinska, L. Patterns of Wallerian degeneration of myelinated fibres in short and long peripheral stumps and in isolated segments of rat phrenic nerve. Interpretation of the role of axoplasmic flow of the trophic factor. Brain Res. 233(2):227–240, 1982.

  26. 26.

    Mahoney, M. J., R. R. Chen, J. Tan, and W. M. Saltzman. The influence of microchannels on neurite growth and architecture. Biomaterials 26(7):771–778, 2005.

  27. 27.

    McGeachie, A. B., K. Koishi, T. Imamura, and I. S. McLennan. Fibroblast growth factor-5 is expressed in Schwann cells and is not essential for motoneurone survival. Neuroscience 104(3):891–899, 2001.

  28. 28.

    McKeon, R. J., R. C. Schreiber, J. S. Rudge, and J. Silver. Reduction of neurite outgrowth in a model of glial scarring following CNS injury is correlated with the expression of inhibitory molecules on reactive astrocytes. J. Neurosci. 11(11):3398–3411, 1991.

  29. 29.

    Meijering, E., M. Jacob, J. C. Sarria, P. Steiner, H. Hirling, and M. Unser. Design and validation of a tool for neurite tracing and analysis in fluorescence microscopy images. Cytometry A 58(2):167–176, 2004.

  30. 30.

    Meyer, M., I. Matsuoka, C. Wetmore, L. Olson, and H. Thoenen. Enhanced synthesis of brain-derived neurotrophic factor in the lesioned peripheral nerve: different mechanisms are responsible for the regulation of BDNF and NGF mRNA. J. Cell Biol. 119(1):45–54, 1992.

  31. 31.

    Mizuno, H., H. Warita, M. Aoki, and Y. Itoyama. Accumulation of chondroitin sulfate proteoglycans in the microenvironment of spinal motor neurons in amyotrophic lateral sclerosis transgenic rats. J. Neurosci. Res. 86(11):2512–2523, 2008.

  32. 32.

    Morrison, S., L. S. Mitchell, M. S. Ecob-Prince, I. R. Griffiths, C. E. Thomson, J. A. Barrie, and D. Kirkham. P0 gene expression in cultured Schwann cells. J. Neurocytol. 20(9):769–780, 1991.

  33. 33.

    National Center for Health Statistics based on Classification of Diseases tR. Clinical modification for the following categories: ICD-9 CM code: 04.3, 04.5, 04.6, 04.7.

  34. 34.

    Naveilhan, P., W. M. ElShamy, and P. Ernfors. Differential regulation of mRNAs for GDNF and its receptors Ret and GDNFR alpha after sciatic nerve lesion in the mouse. Eur. J. Neurosci. 9(7):1450–1460, 1997.

  35. 35.

    Omura, T., M. Sano, K. Omura, T. Hasegawa, M. Doi, T. Sawada, and A. Nagano. Different expressions of BDNF, NT3, and NT4 in muscle and nerve after various types of peripheral nerve injuries. J. Peripher. Nerv. Syst. 10(3):293–300, 2005.

  36. 36.

    Pierucci, A., E. A. de Duek, and A. L. de Oliveira. Peripheral nerve regeneration through biodegradable conduits prepared using solvent evaporation. Tissue Eng. Part A 14(5):595–606, 2008.

  37. 37.

    Sango, K., A. Oohira, K. Ajiki, A. Tokashiki, M. Horie, and H. Kawano. Phosphacan and neurocan are repulsive substrata for adhesion and neurite extension of adult rat dorsal root ganglion neurons in vitro. Exp. Neurol. 182(1):1–11, 2003.

  38. 38.

    Scarlato, M., J. Ara, P. Bannerman, S. Scherer, and D. Pleasure. Induction of neuropilins-1 and -2 and their ligands, Sema3A, Sema3F, and VEGF, during Wallerian degeneration in the peripheral nervous system. Exp. Neurol. 183(2):489–498, 2003.

  39. 39.

    Scarlato, M., T. Xu, P. Bannerman, J. Beesley, U. R. Reddy, A. Rostami, S. S. Scherer, and D. Pleasure. Axon-Schwann cell interactions regulate the expression of fibroblast growth factor-5 (FGF-5). J. Neurosci. Res. 66(1):16–22, 2001.

  40. 40.

    Schlaepfer, W. W. Structural alterations of peripheral nerve induced by the calcium ionophore A23187. Brain Res. 136(1):1–9, 1977.

  41. 41.

    Snow, D. M., E. M. Brown, and P. C. Letourneau. Growth cone behavior in the presence of soluble chondroitin sulfate proteoglycan (CSPG), compared to behavior on CSPG bound to laminin or fibronectin. Int. J. Dev. Neurosci. 14(3):331–349, 1996.

  42. 42.

    Snow, D. M., and P. C. Letourneau. Neurite outgrowth on a step gradient of chondroitin sulfate proteoglycan (CS-PG). J. Neurobiol. 23(3):322–336, 1992.

  43. 43.

    Stewart, H. J., P. A. Eccleston, K. R. Jessen, and R. Mirsky. Interaction between cAMP elevation, identified growth factors, and serum components in regulating Schwann cell growth. J. Neurosci. Res. 30(2):346–352, 1991.

  44. 44.

    Tansey, K. E., J. L. Seifert, B. Botterman, M. R. Delgado, and M. I. Romero. Peripheral nerve repair through multi-luminal biosynthetic implants. Ann. Biomed. Eng. 39(6):1815–1828, 2011.

  45. 45.

    Tomita, K., T. Kubo, K. Matsuda, R. Hattori, T. Fujiwara, K. Yano, and K. Hosokawa. Effect of conduit repair on aberrant motor axon growth within the nerve graft in rats. Microsurgery 27(5):500–509, 2007.

  46. 46.

    Trupp, M., M. Ryden, H. Jornvall, H. Funakoshi, T. Timmusk, E. Arenas, and C. F. Ibanez. Peripheral expression and biological activities of GDNF, a new neurotrophic factor for avian and mammalian peripheral neurons. J. Cell Biol. 130(1):137–148, 1995.

  47. 47.

    Vial, J. D. The early changes in the axoplasm during Wallerian degeneration. J. Biophys. Biochem. Cytol. 4(5):551–555, 1958.

  48. 48.

    Wood, M. D., M. R. MacEwan, A. R. French, A. M. Moore, D. A. Hunter, S. E. Mackinnon, D. W. Moran, G. H. Borschel, and S. E. Sakiyama-Elbert. Fibrin matrices with affinity-based delivery systems and neurotrophic factors promote functional nerve regeneration. Biotechnol. Bioeng. 106(6):970–979, 2010.

  49. 49.

    Wood, M. D., A. M. Moore, D. A. Hunter, S. Tuffaha, G. H. Borschel, S. E. Mackinnon, and S. E. Sakiyama-Elbert. Affinity-based release of glial-derived neurotrophic factor from fibrin matrices enhances sciatic nerve regeneration. Acta Biomater. 5(4):959–968, 2009.

  50. 50.

    Yao, L., S. Wang, W. Cui, R. Sherlock, C. O’Connell, G. Damodaran, A. Gorman, A. Windebank, and A. Pandit. Effect of functionalized micropatterned PLGA on guided neurite growth. Acta Biomater. 5(2):580–588, 2009.

  51. 51.

    Yi, J. H., Y. Katagiri, B. Susarla, D. Figge, A. J. Symes, and H. M. Geller. Alterations in sulfated chondroitin glycosaminoglycans following controlled cortical impact injury in mice. J. Comp. Neurol. 520(15):3295–3313, 2012.

  52. 52.

    Yu, X., and R. V. Bellamkonda. Dorsal root ganglia neurite extension is inhibited by mechanical and chondroitin sulfate-rich interfaces. J. Neurosci. Res. 66(2):303–310, 2001.

  53. 53.

    Zhu, Y., C. Gao, Y. Liu, and J. Shen. Endothelial cell functions in vitro cultured on poly(l-lactic acid) membranes modified with different methods. J. Biomed. Mater. Res. A 69(3):436–443, 2004.

  54. 54.

    Zuo, J., Y. J. Hernandez, and D. Muir. Chondroitin sulfate proteoglycan with neurite-inhibiting activity is up-regulated following peripheral nerve injury. J. Neurobiol. 34(1):41–54, 1998.

Download references

Author information

Correspondence to J. Kent Leach or Peter Bannerman.

Additional information

Associate Editor Scott I. Simon oversaw the review of this article.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Fig. 1. Soluble CSPG inhibits neurite extension. Time-lapse phase contrast images of DRG explants cultured in basal media with a CSPG concentration of 25 μg/mL were taken at 1 h intervals for 48 h. DRG explants were cultured in normal growth media for 24 h prior to exchange with CSPG media. Images at 10× magnification. Supplementary material 1 (MOV 2105 kb)

Supplementary Fig. 2. DRG explants cultured in basal media exhibited normal neurite extension. Time-lapse phase contrast images of DRG explants cultured in basal media were taken at 1 h intervals for 48 h. DRG explants were cultured in normal growth media for 24 h prior to exchange with basal media. Images at 10× magnification. Supplementary material 2 (MOV 1555 kb)

Supplementary Fig. 1. Soluble CSPG inhibits neurite extension. Time-lapse phase contrast images of DRG explants cultured in basal media with a CSPG concentration of 25 μg/mL were taken at 1 h intervals for 48 h. DRG explants were cultured in normal growth media for 24 h prior to exchange with CSPG media. Images at 10× magnification. Supplementary material 1 (MOV 2105 kb)

Supplementary Fig. 2. DRG explants cultured in basal media exhibited normal neurite extension. Time-lapse phase contrast images of DRG explants cultured in basal media were taken at 1 h intervals for 48 h. DRG explants were cultured in normal growth media for 24 h prior to exchange with basal media. Images at 10× magnification. Supplementary material 2 (MOV 1555 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Man, A.J., Leach, J.K. & Bannerman, P. Redirection of Neurite Outgrowth by Coupling Chondroitin Sulfate Proteoglycans to Polymer Membranes. Ann Biomed Eng 42, 1271–1281 (2014). https://doi.org/10.1007/s10439-014-0991-y

Download citation

Keywords

  • Dorsal root ganglia
  • Neurite
  • Chondroitin sulfate proteoglycans
  • PLLA