Cell and Tissue Banking

, Volume 15, Issue 2, pp 227–239 | Cite as

In toto differentiation of human amniotic membrane towards the Schwann cell lineage

  • Asmita Banerjee
  • Sylvia Nürnberger
  • Simone Hennerbichler
  • Sabrina Riedl
  • Christina M. A. P. Schuh
  • Ara Hacobian
  • Andreas Teuschl
  • Johann Eibl
  • Heinz Redl
  • Susanne Wolbank
Original Paper


Human amniotic membrane (hAM) is a tissue containing cells with proven stem cell properties. In its decellularized form it has been successfully applied as nerve conduit biomaterial to improve peripheral nerve regeneration in injury models. We hypothesize that viable hAM without prior cell isolation can be differentiated towards the Schwann cell lineage to generate a possible alternative to commonly applied tissue engineering materials for nerve regeneration. For in vitro Schwann cell differentiation, biopsies of hAM of 8 mm diameter were incubated with a sequential order of neuronal induction and growth factors for 21 days and characterized for cellular viability and the typical glial markers glial fibrillary acidic protein (GFAP), S100β, p75 and neurotrophic tyrosine kinase receptor (NTRK) using immunohistology. The secretion of the neurotrophic factors brain-derived neurotrophic factor (BDNF) and glial cell-derived neurotrophic factor (GDNF) was quantified by ELISA. The hAM maintained high viability, especially under differentiation conditions (90.2 % ± 41.6 day 14; 80.0 % ± 44.5 day 21 compared to day 0). Both, BDNF and GDNF secretion was up-regulated upon differentiation. The fresh membrane stained positive for GFAP and p75 and NTRK, which was strongly increased after culture in differentiation conditions. Especially the epithelial layer within the membrane exhibited a change in morphology upon differentiation forming a multi-layered epithelium with intense accumulations of the marker proteins. However, S100β was expressed at equal levels and equal distribution in fresh and cultured hAM conditions. Viable hAM may be a promising alternative to present formulations used for peripheral nerve regeneration.


Schwann cells Human amniotic membrane In toto differentiation Amniotic epithelial cells Peripheral nerve regeneration Adipose-derived stem cells 



We would like to thank Daniela Dopler, Alice Zimmermann and Sidrah Chaudry for their technical assistance and Eva Schwingenschlögl for support with graphic design.

Conflict of interest

We would like to disclose that the co-authors Johann Eibl and Heinz Redl own the patent rights for “Process for differentiating stem cells of the amniotic membrane”.


  1. Adinolfi M, Akle CA, McColl I, Fensom AH, Tansley L, Connolly P, Hsi BL, Faulk WP, Travers P, Bodmer WF (1982) Expression of HLA antigens, beta 2-microglobulin and enzymes by human amniotic epithelial cells. Nature 295:325–327PubMedCrossRefGoogle Scholar
  2. Akle CA, Adinolfi M, Welsh KI, Leibowitz S, McColl I (1981) Immunogenicity of human amniotic epithelial cells after transplantation into volunteers. Lancet 2:1003–1005PubMedCrossRefGoogle Scholar
  3. Bailo M, Soncini M, Vertua E, Signoroni PB, Sanzone S, Lombardi G, Arienti D, Calamani F, Zatti D, Paul P, Albertini A, Zorzi F, Cavagnini A, Candotti F, Wengler GS, Parolini O (2004) Engraftment potential of human amnion and chorion cells derived from term placenta. Transplantation 78:1439–1448PubMedCrossRefGoogle Scholar
  4. Caddick J, Kingham PJ, Gardiner NJ, Wiberg M, Terenghi G (2006) Phenotypic and functional characteristics of mesenchymal stem cells differentiated along a Schwann cell lineage. Glia 54:840–849PubMedCrossRefGoogle Scholar
  5. Cargnoni A, Gibelli L, Tosini A, Signoroni PB, Nassuato C, Arienti D, Lombardi G, Albertini A, Wengler GS, Parolini O (2009) Transplantation of allogeneic and xenogeneic placenta-derived cells reduces bleomycin-induced lung fibrosis. Cell Transpl 18:405–422CrossRefGoogle Scholar
  6. Chernousov MA, Yu WM, Chen ZL, Carey DJ, Strickland S (2008) Regulation of Schwann cell function by the extracellular matrix. Glia 56:1498–1507PubMedCrossRefGoogle Scholar
  7. Davis GE, Blaker SN, Engvall E, Varon S, Manthorpe M, Gage FH (1987) Human amnion membrane serves as a substratum for growing axons in vitro and in vivo. Science 236:1106–1109PubMedCrossRefGoogle Scholar
  8. di Summa PG, Kalbermatten DF, Raffoul W, Terenghi G, Kingham PJ (2013) Extracellular matrix molecules enhance the neurotrophic effect of Schwann cell-like differentiated adipose-derived stem cells and increase cell survival under stress conditions. Tissue Eng Part A 19:368–379PubMedCentralPubMedCrossRefGoogle Scholar
  9. Hennerbichler S, Reichl B, Pleiner D, Gabriel C, Eibl J, Redl H (2007) The influence of various storage conditions on cell viability in amniotic membrane. Cell Tissue Bank 8:1–8PubMedCrossRefGoogle Scholar
  10. Hubert T, Grimal S, Carroll P, Fichard-Carroll A (2009) Collagens in the developing and diseased nervous system. Cell Mol Life Sci 66:1223–1238PubMedCrossRefGoogle Scholar
  11. Insausti CL, Blanquer M, Bleda P, Iniesta P, Majado MJ, Castellanos G, Moraleda JM (2010) The amniotic membrane as a source of stem cells. Histol Histopathol 25:91–98PubMedGoogle Scholar
  12. Jiang L, Zhu JK, Liu XL, Xiang P, Hu J, Yu WH (2008) Differentiation of rat adipose tissue-derived stem cells into Schwann-like cells in vitro. Neuroreport 19:1015–1019PubMedCrossRefGoogle Scholar
  13. Kakishita K, Nakao N, Sakuragawa N, Itakura T (2003) Implantation of human amniotic epithelial cells prevents the degeneration of nigral dopamine neurons in rats with 6-hydroxydopamine lesions. Brain Res 980:48–56PubMedCrossRefGoogle Scholar
  14. Keilhoff G, Goihl A, Stang F, Wolf G, Fansa H (2006a) Peripheral nerve tissue engineering: autologous Schwann cells versus transdifferentiated mesenchymal stem cells. Tissue Eng 12:1451–1465PubMedCrossRefGoogle Scholar
  15. Keilhoff G, Stang F, Goihl A, Wolf G, Fansa H (2006b) Transdifferentiated mesenchymal stem cells as alternative therapy in supporting nerve regeneration and myelination. Cell Mol Neurobiol 26:1235–1252PubMedCrossRefGoogle Scholar
  16. Kingham PJ, Kalbermatten DF, Mahay D, Armstrong SJ, Wiberg M, Terenghi G (2007) Adipose-derived stem cells differentiate into a Schwann cell phenotype and promote neurite outgrowth in vitro. Exp Neurol 207:267–274PubMedCrossRefGoogle Scholar
  17. Kronsteiner B, Peterbauer-Scherb A, Grillari-Voglauer R, Redl H, Gabriel C, van Griensven M, Wolbank S (2011a) Human mesenchymal stem cells and renal tubular epithelial cells differentially influence monocyte-derived dendritic cell differentiation and maturation. Cell Immunol 267:30–38PubMedCrossRefGoogle Scholar
  18. Kronsteiner B, Wolbank S, Peterbauer A, Hackl C, Redl H, van Griensven M, Gabriel C (2011b) Human mesenchymal stem cells from adipose tissue and amnion influence T-cells depending on stimulation method and presence of other immune cells. Stem Cells Dev 20:2115–2126PubMedCrossRefGoogle Scholar
  19. Liang HS, Liang P, Xu Y, Wu JN, Liang T, Xu XP, Liu EZ (2009) Denuded human amniotic membrane seeding bone marrow stromal cells as an effective composite matrix stimulates axonal outgrowth of rat neural cortical cells in vitro. Acta Neurochir (Wien) 151:1113–1120CrossRefGoogle Scholar
  20. Liang H, Li C, Gao A, Liang P, Shao Y, Lin T, Zhang X (2012) Spinal duraplasty with two novel substitutes restored locomotor function after acute laceration spinal cord injury in rats. J Biomed Mater Res B Appl Biomater 100:2131–2140PubMedCrossRefGoogle Scholar
  21. Lindenmair A, Wolbank S, Stadler G, Meinl A, Peterbauer-Scherb A, Eibl J, Polin H, Gabriel C, van Griensven M, Redl H (2010) Osteogenic differentiation of intact human amniotic membrane. Biomaterials 31:8659–8665PubMedCrossRefGoogle Scholar
  22. Mahay D, Terenghi G, Shawcross SG (2008) Schwann cell mediated trophic effects by differentiated mesenchymal stem cells. Exp Cell Res 314:2692–2701PubMedCrossRefGoogle Scholar
  23. Mahmoudi-Rad M, Abolhasani E, Moravvej H, Mahmoudi-Rad N, Mirdamadi Y (2013) Acellular amniotic membrane: an appropriate scaffold for fibroblast proliferation. Clin Exp Dermatol 38:646–651PubMedCrossRefGoogle Scholar
  24. Manuelpillai U, Lourensz D, Vaghjiani V, Tchongue J, Lacey D, Tee JY, Murthi P, Chan J, Hodge A, Sievert W (2012) Human amniotic epithelial cell transplantation induces markers of alternative macrophage activation and reduces established hepatic fibrosis. PLoS ONE 7:e38631PubMedCentralPubMedCrossRefGoogle Scholar
  25. Miki T, Strom SC (2006) Amnion-derived pluripotent/multipotent stem cells. St Cell Rev 2:133–142CrossRefGoogle Scholar
  26. Mligiliche N, Endo K, Okamoto K, Fujimoto E, Ide C (2002) Extracellular matrix of human amnion manufactured into tubes as conduits for peripheral nerve regeneration. J Biomed Mater Res 63:591–600PubMedCrossRefGoogle Scholar
  27. Ndubaku U, de Bellard ME (2008) Glial cells: old cells with new twists. Acta Histochem 110:182–195PubMedCentralPubMedCrossRefGoogle Scholar
  28. Park HW, Lim MJ, Jung H, Lee SP, Paik KS, Chang MS (2010) Human mesenchymal stem cell-derived Schwann cell-like cells exhibit neurotrophic effects, via distinct growth factor production, in a model of spinal cord injury. Glia 58:1118–1132PubMedCrossRefGoogle Scholar
  29. Parolini O, Alviano F, Bergwerf I, Boraschi D, De BC, De WP, Dominici M, Evangelista M, Falk W, Hennerbichler S, Hess DC, Lanzoni G, Liu B, Marongiu F, McGuckin C, Mohr S, Nolli ML, Ofir R, Ponsaerts P, Romagnoli L, Solomon A, Soncini M, Strom S, Surbek D, Venkatachalam S, Wolbank S, Zeisberger S, Zeitlin A, Zisch A, Borlongan CV (2010) Toward cell therapy using placenta-derived cells: disease mechanisms, cell biology, preclinical studies, and regulatory aspects at the round table. St Cells Dev 19:143–154CrossRefGoogle Scholar
  30. Peterbauer-Scherb A, Danzer M, Gabriel C, van Griensven M, Redl H, Wolbank S (2012) In vitro adipogenesis of adipose-derived stem cells in 3D fibrin matrix of low component concentration. J Tissue Eng Regen Med 6:434–442PubMedCrossRefGoogle Scholar
  31. Pratama G, Vaghjiani V, Tee JY, Liu YH, Chan J, Tan C, Murthi P, Gargett C, Manuelpillai U (2011) Changes in culture expanded human amniotic epithelial cells: implications for potential therapeutic applications. PLoS ONE 6:e26136PubMedCentralPubMedCrossRefGoogle Scholar
  32. Riau AK, Beuerman RW, Lim LS, Mehta JS (2010) Preservation, sterilization and de-epithelialization of human amniotic membrane for use in ocular surface reconstruction. Biomaterials 31:216–225PubMedCrossRefGoogle Scholar
  33. Ricci E, Vanosi G, Lindenmair A, Hennerbichler S, Peterbauer-Scherb A, Wolbank S, Cargnoni A, Signoroni PB, Campagnol M, Gabriel C, Redl H, Parolini O (2012) Anti-fibrotic effects of fresh and cryopreserved human amniotic membrane in a rat liver fibrosis model. Cell Tissue BankGoogle Scholar
  34. Sakuragawa N, Kakinuma K, Kikuchi A, Okano H, Uchida S, Kamo I, Kobayashi M, Yokoyama Y (2004) Human amnion mesenchyme cells express phenotypes of neuroglial progenitor cells. J Neurosci Res 78:208–214PubMedCrossRefGoogle Scholar
  35. Stadler G, Hennerbichler S, Lindenmair A, Peterbauer A, Hofer K, van Griensven M, Gabriel C, Redl H, Wolbank S (2008) Phenotypic shift of human amniotic epithelial cells in culture is associated with reduced osteogenic differentiation in vitro. Cytotherapy 10:743–752PubMedCrossRefGoogle Scholar
  36. Takashima S, Yasuo M, Sanzen N, Sekiguchi K, Okabe M, Yoshida T, Toda A, Nikaido T (2008) Characterization of laminin isoforms in human amnion. Tissue Cell 40:75–81PubMedCrossRefGoogle Scholar
  37. Terenghi G (1999) Peripheral nerve regeneration and neurotrophic factors. J Anat 194(Pt 1):1–14PubMedCentralPubMedCrossRefGoogle Scholar
  38. Tohill M, Mantovani C, Wiberg M, Terenghi G (2004) Rat bone marrow mesenchymal stem cells express glial markers and stimulate nerve regeneration. Neurosci Lett 362:200–203PubMedCrossRefGoogle Scholar
  39. Tomita K, Madura T, Sakai Y, Yano K, Terenghi G, Hosokawa K (2013) Glial differentiation of human adipose-derived stem cells: implications for cell-based transplantation therapy. Neuroscience 236:55–65PubMedCrossRefGoogle Scholar
  40. Uchida S, Inanaga Y, Kobayashi M, Hurukawa S, Araie M, Sakuragawa N (2000) Neurotrophic function of conditioned medium from human amniotic epithelial cells. J Neurosci Res 62:585–590PubMedCrossRefGoogle Scholar
  41. Uziyel Y, Hall S, Cohen J (2000) Influence of laminin-2 on Schwann cell-axon interactions. Glia 32:109–121PubMedCrossRefGoogle Scholar
  42. Wallquist W, Plantman S, Thams S, Thyboll J, Kortesmaa J, Lannergren J, Domogatskaya A, Ogren SO, Risling M, Hammarberg H, Tryggvason K, Cullheim S (2005) Impeded interaction between Schwann cells and axons in the absence of laminin alpha4. J Neurosci 25:3692–3700PubMedCrossRefGoogle Scholar
  43. Wang J, Ding F, Gu Y, Liu J, Gu X (2009) Bone marrow mesenchymal stem cells promote cell proliferation and neurotrophic function of Schwann cells in vitro and in vivo. Brain Res 1262:7–15PubMedCrossRefGoogle Scholar
  44. Whittle WL, Gibb W, Challis JR (2000) The characterization of human amnion epithelial and mesenchymal cells: the cellular expression, activity and glucocorticoid regulation of prostaglandin output. Placenta 21:394–401PubMedCrossRefGoogle Scholar
  45. Wilshaw SP, Kearney J, Fisher J, Ingham E (2008) Biocompatibility and potential of acellular human amniotic membrane to support the attachment and proliferation of allogeneic cells. Tissue Eng Part A 14:463–472PubMedCrossRefGoogle Scholar
  46. Wolbank S, Hildner F, Redl H, van Griensven M, Gabriel C, Hennerbichler S (2009) Impact of human amniotic membrane preparation on release of angiogenic factors. J Tissue Eng Regen Med 3:651–654PubMedCrossRefGoogle Scholar
  47. Xu Y, Liu L, Li Y, Zhou C, Xiong F, Liu Z, Gu R, Hou X, Zhang C (2008) Myelin-forming ability of Schwann cell-like cells induced from rat adipose-derived stem cells in vitro. Brain Res 1239:49–55PubMedCrossRefGoogle Scholar
  48. Xue H, Zhang XY, Liu JM, Song Y, Li YF, Chen D (2013) Development of a chemically extracted acellular muscle scaffold seeded with amniotic epithelial cells to promote spinal cord repair. J Biomed Mater Res A 101:145–156PubMedCrossRefGoogle Scholar
  49. Yan ZJ, Zhang P, Hu YQ, Zhang HT, Hong SQ, Zhou HL, Zhang MY, Xu RX (2013) Neural stem-like cells derived from human amnion tissue are effective in treating traumatic brain injury in rat. Neurochem Res 38:1022–1033PubMedCrossRefGoogle Scholar
  50. Yang S, Xue DD, Wu B, Sun HM, Li XS, Dong F, Li WS, Ji FQ, Zhou DS (2013) Pleiotrophin is involved in the amniotic epithelial cell-induced differentiation of human umbilical cord blood-derived mesenchymal stem cells into dopaminergic neuron-like cells. Neurosci Lett 539:86–91PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Asmita Banerjee
    • 1
    • 2
  • Sylvia Nürnberger
    • 1
    • 7
    • 8
  • Simone Hennerbichler
    • 2
    • 3
  • Sabrina Riedl
    • 1
    • 2
  • Christina M. A. P. Schuh
    • 1
    • 2
  • Ara Hacobian
    • 1
    • 2
  • Andreas Teuschl
    • 2
    • 5
    • 6
  • Johann Eibl
    • 4
  • Heinz Redl
    • 1
    • 2
  • Susanne Wolbank
    • 1
    • 2
  1. 1.Ludwig Boltzmann Institute for Experimental and Clinical TraumatologyAUVA Research CenterViennaAustria
  2. 2.Austrian Cluster for Tissue RegenerationViennaAustria
  3. 3.Red Cross Blood Transfusion Service for Upper AustriaLinzAustria
  4. 4.Bio-Products and Bio-Engineering AGViennaAustria
  5. 5.City of Vienna Competence Team Tissue Engineering BioreactorsUniversity of Applied Science Technikum WienViennaAustria
  6. 6.Department of Biochemical EngineeringUniversity of Applied Science Technikum WienViennaAustria
  7. 7.Department of TraumatologyMedical University of ViennaViennaAustria
  8. 8.Bernhard Gottlieb University Clinic of DentistryViennaAustria

Personalised recommendations