Neurochemical Research

, Volume 39, Issue 11, pp 2047–2057 | Cite as

The Beneficial Effect of Chitooligosaccharides on Cell Behavior and Function of Primary Schwann Cells is Accompanied by Up-Regulation of Adhesion Proteins and Neurotrophins

Original Paper


Chitosan-based tissue engineered nerve grafts are successfully used for bridging peripheral nerve gaps. The biodegradation products of chitosan are water-dissolvable chitooligosaccharides (COSs), which have been shown to support peripheral nerve regeneration. In this study, we aimed to examine in vitro interactions between COSs and Schwann cells (SCs), the principal glial cells in the peripheral nervous system. Treatment of primary SCs with COSs enhanced cell survival and promoted cell proliferation in a dose-dependent manner (0.25–1.0 mg/ml), as determined by real-time cell analyzer-based assay, cell growth assay, cell cycle analysis, and EdU incorporation. Western blot analysis and immunocytochemistry with antibodies against MBP and MAG (two myelin-specific markers) showed that COSs enhanced axonal myelination in a co-culture system consisting of SCs and dorsal root ganglia (DRGs). Furthermore, we observed that COSs enhanced the protein expression of N-cadherin and β-catenin in primary SCs, and also increased the release of BDNF and NGF in co-culture of SCs with DRGs. And we also noted that knockdown of N-cadherin in primary SCs reduced COSs-induced increase in cell proliferation. Our findings suggested that beneficial effects of COSs on cell behavior and functions of primary SCs might be accompanied by up-regulation of adhesion proteins and neurotrophins, thus providing a new insight into the supportive role of COSs during peripheral nerve regeneration.


Chitooligosaccharides Schwann cell Dorsal root ganglion Proliferation Myelination Cell adhesion proteins 



This study was supported by the National Natural Science Foundation of China (Grant No. 81101159), General Natural Science Research Project of Colleges and Universities of Jiangsu Province (Grant No. 12KJB310010), and a Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD). We thank Professor Jie Liu for assistance in manuscript preparation.


  1. 1.
    Gruart A, Streppel M, Guntinas-Lichius O, Angelov DN, Neiss WF, Delgado-Garcia JM (2003) Motoneuron adaptability to new motor tasks following two types of facial–facial anastomosis in cats. Brain 126:115–133PubMedCrossRefGoogle Scholar
  2. 2.
    Mokarram N, Merchant A, Mukhatyar V, Patel G, Bellamkonda RV (2012) Effect of modulating macrophage phenotype on peripheral nerve repair. Biomaterials 33:8793–8801PubMedCrossRefPubMedCentralGoogle Scholar
  3. 3.
    Wang Y, Tang X, Yu B, Gu Y, Yuan Y, Yao D, Ding F, Gu X (2012) Gene network revealed involvements of Birc2, Birc3 and Tnfrsf1a in anti-apoptosis of injured peripheral nerves. PLoS ONE 7:e43436PubMedCrossRefPubMedCentralGoogle Scholar
  4. 4.
    Jessen KR, Mirsky R (2005) The origin and development of glial cells in peripheral nerves. Nat Rev Neurosci 6:671–682PubMedCrossRefGoogle Scholar
  5. 5.
    Jiang H, Qu W, Li Y, Zhong W, Zhang W (2013) Platelet-derived growth factors-BB and fibroblast growth factors-base induced proliferation of Schwann cells in a 3D environment. Neurochem Res 38:346–355PubMedCrossRefGoogle Scholar
  6. 6.
    Gu X, Ding F, Yang Y, Liu J (2011) Construction of tissue engineered nerve grafts and their application in peripheral nerve regeneration. Prog Neurobiol 93:204–230PubMedCrossRefGoogle Scholar
  7. 7.
    Pereira JA, Lebrun-Julien F, Suter U (2012) Molecular mechanisms regulating myelination in the peripheral nervous system. Trends Neurosci 35:123–134PubMedCrossRefGoogle Scholar
  8. 8.
    Freier T, Koh HS, Kazazian K, Shoichet MS (2005) Controlling cell adhesion and degradation of chitosan films by N-acetylation. Biomaterials 26:5872–5878PubMedCrossRefGoogle Scholar
  9. 9.
    Hu N, Wu H, Xue C, Gong Y, Wu J, Xiao Z, Yang Y, Ding F, Gu X (2013) Long-term outcome of the repair of 50 mm long median nerve defects in rhesus monkeys with marrow mesenchymal stem cells-containing, chitosan-based tissue engineered nerve grafts. Biomaterials 34:100–111PubMedCrossRefGoogle Scholar
  10. 10.
    He Q, Zhang T, Yang Y, Ding F (2009) In vitro biocompatibility of chitosan-based materials to primary culture of hippocampal neurons. J Mater Sci Mater Med 20:1457–1466PubMedCrossRefGoogle Scholar
  11. 11.
    Jiao H, Yao J, Yang Y, Chen X, Lin W, Li Y, Gu X, Wang X (2009) Chitosan/polyglycolic acid nerve grafts for axon regeneration from prolonged axotomized neurons to chronically denervated segments. Biomaterials 30:5004–5018PubMedCrossRefGoogle Scholar
  12. 12.
    Yang Y, Liu M, Gu Y, Lin S, Ding F, Gu X (2009) Effect of chitooligosaccharide on neuronal differentiation of PC-12 cells. Cell Biol Int 33:352–356PubMedCrossRefGoogle Scholar
  13. 13.
    Ding F, Wu J, Yang Y, Hu W, Zhu Q, Tang X, Liu J, Gu X (2010) Use of tissue-engineered nerve grafts consisting of a chitosan/poly(lactic-co-glycolic acid)-based scaffold included with bone marrow mesenchymal cells for bridging 50-mm dog sciatic nerve gaps. Tissue Eng Part A 16:3779–3790PubMedCrossRefGoogle Scholar
  14. 14.
    Joodi G, Ansari N, Khodagholi F (2011) Chitooligosaccharide-mediated neuroprotection is associated with modulation of Hsps expression and reduction of MAPK phosphorylation. Int J Biol Macromol 48:726–735PubMedCrossRefGoogle Scholar
  15. 15.
    Huang R, Mendis E, Rajapakse N, Kim SK (2006) Strong electronic charge as an important factor for anticancer activity of chitooligosaccharides (COS). Life Sci 78:2399–2408PubMedCrossRefGoogle Scholar
  16. 16.
    Ryu B, Himaya SW, Napitupulu RJ, Eom TK, Kim SK (2012) Sulfated chitooligosaccharide II (SCOS II) suppress collagen degradation in TNF-induced chondrosarcoma cells via NF-kappaB pathway. Carbohydr Res 350:55–61PubMedCrossRefGoogle Scholar
  17. 17.
    Bahar B, O’Doherty JV, Maher S, McMorrow J, Sweeney T (2012) Chitooligosaccharide elicits acute inflammatory cytokine response through AP-1 pathway in human intestinal epithelial-like (Caco-2) cells. Mol Immunol 51:283–291PubMedCrossRefGoogle Scholar
  18. 18.
    Lillo L, Alarcon J, Cabello G, Cespedes C, Caro C (2008) Antibacterial activity of chitooligosaccharides. Z Naturforsch C 63:644–648PubMedGoogle Scholar
  19. 19.
    Fernandes JC, Tavaria FK, Fonseca SC, Ramos OS, Pintado ME, Malcata FX (2010) In vitro screening for anti-microbial activity of chitosans and chitooligosaccharides, aiming at potential uses in functional textiles. J Microbiol Biotechnol 20:311–318PubMedCrossRefGoogle Scholar
  20. 20.
    Zhou S, Yang Y, Gu X, Ding F (2008) Chitooligosaccharides protect cultured hippocampal neurons against glutamate-induced neurotoxicity. Neurosci Lett 444:270–274PubMedCrossRefGoogle Scholar
  21. 21.
    Xu Y, Zhang Q, Yu S, Yang Y, Ding F (2011) The protective effects of chitooligosaccharides against glucose deprivation-induced cell apoptosis in cultured cortical neurons through activation of PI3K/Akt and MEK/ERK1/2 pathways. Brain Res 1375:49–58PubMedCrossRefGoogle Scholar
  22. 22.
    Jiang M, Zhuge X, Yang Y, Gu X, Ding F (2009) The promotion of peripheral nerve regeneration by chitooligosaccharides in the rat nerve crush injury model. Neurosci Lett 454:239–243PubMedCrossRefGoogle Scholar
  23. 23.
    He B, Liu SQ, Chen Q, Li HH, Ding WJ, Deng M (2011) Carboxymethylated chitosan stimulates proliferation of Schwann cells in vitro via the activation of the ERK and Akt signaling pathways. Eur J Pharmacol 667:195–201PubMedCrossRefGoogle Scholar
  24. 24.
    Tao HY, He B, Liu SQ, Wei AL, Tao FH, Tao HL, Deng WX, Li HH, Chen Q (2013) Effect of carboxymethylated chitosan on the biosynthesis of NGF and activation of the Wnt/beta-catenin signaling pathway in the proliferation of Schwann cells. Eur J Pharmacol 702:85–92PubMedCrossRefGoogle Scholar
  25. 25.
    Yang Y, Shu R, Shao J, Xu G, Gu X (2006) Radical scavenging activity of chitooligosaccharide with different molecular weights. Eur Food Res Technol 222:36–40CrossRefGoogle Scholar
  26. 26.
    Weinstein DE, Wu R (2001) Isolation and purification of primary Schwann cells. Curr Protoc Neurosci Chapter 3:Unit 3 17Google Scholar
  27. 27.
    Eshed Y, Feinberg K, Poliak S, Sabanay H, Sarig-Nadir O, Spiegel I, Bermingham JR Jr, Peles E (2005) Gliomedin mediates Schwann cell-axon interaction and the molecular assembly of the nodes of Ranvier. Neuron 47:215–229PubMedCrossRefGoogle Scholar
  28. 28.
    Garcia SN, Gutierrez L, McNulty A (2013) Real-time cellular analysis as a novel approach for in vitro cytotoxicity testing of medical device extracts. J Biomed Mater Res, Part A 101:2097–2106CrossRefGoogle Scholar
  29. 29.
    Teng Z, Kuang X, Wang J, Zhang X (2013) Real-time cell analysis–a new method for dynamic, quantitative measurement of infectious viruses and antiserum neutralizing activity. J Virol Methods 193:364–370PubMedCrossRefGoogle Scholar
  30. 30.
    Baldin V, Lukas J, Marcote MJ, Pagano M, Draetta G (1993) Cyclin D1 is a nuclear protein required for cell cycle progression in G1. Genes Dev 7:812–821PubMedCrossRefGoogle Scholar
  31. 31.
    Abdelaal MY, Sobahi TR, Al-Shareef HF (2013) Modification of chitosan derivatives of environmental and biological interest: a green chemistry approach. Int J Biol Macromol 55:231–239PubMedCrossRefGoogle Scholar
  32. 32.
    Jangouk P, Dehmel T, Meyer Zu, Horste G, Ludwig A, Lehmann HC, Kieseier BC (2009) Involvement of ADAM10 in axonal outgrowth and myelination of the peripheral nerve. Glia 57:1765–1774PubMedCrossRefGoogle Scholar
  33. 33.
    Fancy SP, Chan JR, Baranzini SE, Franklin RJ, Rowitch DH (2011) Myelin regeneration: a recapitulation of development? Annu Rev Neurosci 34:21–43PubMedCrossRefGoogle Scholar
  34. 34.
    Liang C, Tao Y, Shen C, Tan Z, Xiong WC, Mei L (2012) Erbin is required for myelination in regenerated axons after injury. J Neurosci 32:15169–15180PubMedCrossRefPubMedCentralGoogle Scholar
  35. 35.
    Lemke G (1988) Unwrapping the genes of myelin. Neuron 1:535–543PubMedCrossRefGoogle Scholar
  36. 36.
    Kristiansen LV, Bannon MJ, Meador-Woodruff JH (2009) Expression of transcripts for myelin related genes in postmortem brain from cocaine abusers. Neurochem Res 34:46–54PubMedCrossRefPubMedCentralGoogle Scholar
  37. 37.
    Zhang Y, Yeh J, Richardson PM, Bo X (2008) Cell adhesion molecules of the immunoglobulin superfamily in axonal regeneration and neural repair. Restor Neurol Neurosci 26:81–96PubMedGoogle Scholar
  38. 38.
    Parrinello S, Napoli I, Ribeiro S, Wingfield Digby P, Fedorova M, Parkinson DB, Doddrell RD, Nakayama M, Adams RH, Lloyd AC (2010) EphB signaling directs peripheral nerve regeneration through Sox2-dependent Schwann cell sorting. Cell 143:145–155PubMedCrossRefGoogle Scholar
  39. 39.
    Cavallaro U, Dejana E (2011) Adhesion molecule signalling: not always a sticky business. Nat Rev Mol Cell Biol 12:189–197PubMedCrossRefGoogle Scholar
  40. 40.
    Sakai N, Insolera R, Sillitoe RV, Shi SH, Kaprielian Z (2012) Axon sorting within the spinal cord marginal zone via Robo-mediated inhibition of N-cadherin controls spinocerebellar tract formation. J Neurosci 32:15377–15387PubMedCrossRefPubMedCentralGoogle Scholar
  41. 41.
    Corell M, Wicher G, Limbach C, Kilimann MW, Colman DR, Fex Svenningsen A (2010) Spatiotemporal distribution and function of N-cadherin in postnatal Schwann cells: a matter of adhesion? J Neurosci Res 88:2338–2349PubMedGoogle Scholar
  42. 42.
    Sakane F, Miyamoto Y (2013) N-cadherin regulates the proliferation and differentiation of ventral midbrain dopaminergic progenitors. Dev Neurobiol 73(7):518–529PubMedCrossRefGoogle Scholar
  43. 43.
    Lelievre EC, Plestant C, Boscher C, Wolff E, Mege RM, Birbes H (2012) N-cadherin mediates neuronal cell survival through Bim down-regulation. PLoS ONE 7:e33206PubMedCrossRefPubMedCentralGoogle Scholar
  44. 44.
    Gess B, Halfter H, Kleffner I, Monje P, Athauda G, Wood PM, Young P, Wanner IB (2008) Inhibition of N-cadherin and beta-catenin function reduces axon-induced Schwann cell proliferation. J Neurosci Res 86:797–812PubMedCrossRefGoogle Scholar
  45. 45.
    Martianez T, Lamarca A, Casals N, Gella A (2013) N-cadherin expression is regulated by UTP in schwannoma cells. Purinergic Signal 9:259–270PubMedCrossRefPubMedCentralGoogle Scholar
  46. 46.
    Wanner IB, Wood PM (2002) N-cadherin mediates axon-aligned process growth and cell–cell interaction in rat Schwann cells. J Neurosci 22:4066–4079PubMedGoogle Scholar
  47. 47.
    Lewallen KA, Shen YA, De la Torre AR, Ng BK, Meijer D, Chan JR (2011) Assessing the role of the cadherin/catenin complex at the Schwann cell-axon interface and in the initiation of myelination. J Neurosci 31:3032–3043PubMedCrossRefPubMedCentralGoogle Scholar
  48. 48.
    Wanner IB, Guerra NK, Mahoney J, Kumar A, Wood PM, Mirsky R, Jessen KR (2006) Role of N-cadherin in Schwann cell precursors of growing nerves. Glia 54:439–459PubMedCrossRefGoogle Scholar
  49. 49.
    Chan JR, Cosgaya JM, Wu YJ, Shooter EM (2001) Neurotrophins are key mediators of the myelination program in the peripheral nervous system. Proc Natl Acad Sci USA 98:14661–14668PubMedCrossRefPubMedCentralGoogle Scholar
  50. 50.
    Notterpek L (2003) Neurotrophins in myelination: a new role for a puzzling receptor. Trends Neurosci 26:232–234PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Jiangsu Key Laboratory of Neuroregeneration, Co-innovation Center of NeuroregenerationNantong UniversityNantongPeople’s Republic of China

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