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Purification and in vitro characterization of adult canine olfactory ensheathing cells

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Abstract

Olfactory ensheathing cells (OECs) are known to promote neural repair under experimental conditions. The experimental focus has so far been almost entirely on rodent OECs (rOECs), and hence whether human OECs (humOECs) display similar properties is unclear. Studies on larger mammals as an “intermediate” model may be helpful for translating the experimental evidence gathered so far into novel therapeutic strategies. In the present study, we purified adult canine OECs (caOECs) from the olfactory bulb and analyzed their in vitro properties with respect to antigen expression, proliferation, and differentiation. Secondary caOECs shared the expression of marker molecules and the reactivity toward growth factors, with rOECs and humOECs. CaOECs were positively immunostained for the low affinity neurotrophin receptor p75, GFAP, and O4 and proliferated in response to fibroblast growth factor-2 and heregulin-1β. No decline in proliferation was noted at higher passages (>8). The effects of forskolin, which neither increased proliferation nor stimulated the expression of O4, were clearly different from those on rOECs. Moreover, caOECs displayed their typical spindle-shaped morphology only upon growth factor/forskolin addition, whereas mitotically quiescent caOECs had a flattened morphology. Thus, caOECs can readily be purified from adult canine olfactory bulb and expanded by using established OEC mitogens. The behavior of caOECs toward forskolin suggests that caOECs and humOECs share a number of properties amd implies the presence of common intracellular signalling pathways. CaOECs therefore represent a suitable model system relevant for humOECs in neural repair studies.

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References

  • Alexander CL, Fitzgerald UF, Barnett SC (2002) Identification of growth factors that promote long-term proliferation of olfactory ensheathing cells and modulate their antigenic phenotype. Glia 37:349–364

    Article  PubMed  Google Scholar 

  • Barber PC (1982) Neurogenesis and regeneration in the primary olfactory pathway of mammals. Bibl Anat 23:12–25

    PubMed  Google Scholar 

  • Barnett SC, Chang L (2004) Olfactory ensheathing cells and CNS repair: going solo or in need of a friend? Trends Neurosci 27:54–60

    Article  PubMed  CAS  Google Scholar 

  • Barnett SC, Riddell JS (2004) Olfactory ensheathing cells (OECs) and the treatment of CNS injury: advantages and possible caveats. J Anat 204:57–67

    Article  PubMed  Google Scholar 

  • Barnett SC, Alexander CL, Iwashita Y, Gilson JM, Crowther J, Clark L, Dunn LT, Papanastassiou V, Kennedy PG, Franklin RJ (2000) Identification of a human olfactory ensheathing cell that can effect transplant-mediated remyelination of demyelinated CNS axons. Brain 123:1581–1588

    Article  PubMed  Google Scholar 

  • Bartholomei JC, Greer CA (2000) Olfactory ensheathing cells: bridging the gap in spinal cord repair. Neurosurgery 47:1057–1069

    Article  Google Scholar 

  • Bianco JI, Perry C, Harkin DG, Mackay-Sim A, Féron F (2004) Neurotrophin-3 promotes proliferation of olfactory ensheathing cells from human nose. Glia 45:111–123

    Article  PubMed  Google Scholar 

  • Borenfreud E, Babinet H (1996) Neutral red (NR) assay. In: Doyle A, Griffiths JB, Newell DG (eds) Cell and tissue culture: laboratory procedures. Wiley, Chichester, pp 4B:7.1–4B:7.7

    Google Scholar 

  • Boyd JG, Doucette R, Kawaja MD (2005) Defining the role of olfactory ensheathing cells in facilitating axon remyelination following damage to the spinal cord. FASEB J 7:694–703

    Article  Google Scholar 

  • Chu L, Norota I, Yomogida S, Ishii K, Endoh M (2004) Differential inotropic effects of endothelin-1, angiotensin II, and phenylephrine induced by crosstalk with cAMP-mediated signaling process in dog ventricular myocardium. J Pharmacol Sci 96:199–207

    Article  PubMed  CAS  Google Scholar 

  • Chuah MI, West AK (2002) Cellular and molecular biology of ensheathing cells. Microsc Res Tech 58:216–227

    Article  PubMed  CAS  Google Scholar 

  • Chuah MI, Cossins J, Woodhall E, Tennent R, Nash G, West AK (2000) Glial growth factor 2 induces proliferation and structural changes in ensheathing cells. Brain Res 28:265–274

    Article  Google Scholar 

  • Doucette R, Devon R (1995) Elevated intracellular levels of cAMP induce olfactory ensheathing cells to express GAL-C and GFAP but not MBP. Glia 13:130–140

    Article  PubMed  CAS  Google Scholar 

  • Féron F, Perry C, Cochrane J, Licina P, Nowitzke A, Urquhart S, Geraghty T, Mackay-Sim A (2005) Autologous olfactory ensheathing cell transplantation in human spinal cord injury. Brain 128:2951–2960

    Article  PubMed  Google Scholar 

  • Field P, Li Y, Raisman G (2003) Ensheathment of the olfactory nerves in the adult rat. J Neurocytol 32:317–324

    Article  PubMed  Google Scholar 

  • Franceschini IA, Barnett SC (1996) Low-affinity NGF-receptor and E-N-CAM expression define two types of olfactory nerve ensheathing cells that share a common lineage. Dev Biol 173:327–343

    Article  PubMed  CAS  Google Scholar 

  • Franklin RJ, Barnett SC (2000) Olfactory ensheathing cells and CNS regeneration: the sweet smell of success? Neuron 28:15–18

    Article  PubMed  CAS  Google Scholar 

  • Franklin RJM, Gilson JM, Franceschini IA, Barnett SC (1996) Schwann cell-like myelination folowing transplantation of an olfactory bulb-ensheathing cell line into areas of demyelination in the adult CNS. Glia 17:217–224

    Article  PubMed  CAS  Google Scholar 

  • Giulian D, Baker TJ (1986) Characterization of ameboid microglia isolated from developing mammalian brain. J Neurosci 6:2163–2178

    PubMed  CAS  Google Scholar 

  • Hahn CG, Han LY, Rawson NE, Mirza N, Borgmann-Winter K, Lenox RH, Arnold SE (2005) In vivo and in vitro neurogenesis in human olfactory epithelium. J Comp Neurol 483:154–163

    Article  PubMed  Google Scholar 

  • Herrera LP, Casas CE, Bates ML, Guest JD (2005) Ultrastructural study of the primary olfactory pathway in Macaca fascicularis. J Comp Neurol 488:427–441

    Article  PubMed  Google Scholar 

  • Imaizumi T, Lankford KL, Waxman SG, Greer CA, Kocsis JD (1998) Transplanted olfactory ensheathing cells remyelinate and enhance axonal conduction in the demyelinated dorsal columns of the rat spinal cord. J Neurosci 18:6176–6185

    PubMed  CAS  Google Scholar 

  • Imaizumi T, Lankford KL, Burton WV, Fodor WL, Kocsis JD (2000) Xenotransplantation of transgenic pig olfactory ensheathing cells promotes axonal regeneration in rat spinal cord. Nat Biotechnol 18:925–927

    Article  Google Scholar 

  • Jeffery ND, Lakatos A, Franklin RJ (2005) Autologous olfactory glial cell transplantation is reliable and safe in naturally occurring canine spinal cord injury. J Neurotrauma 22:1282–1293

    Article  PubMed  Google Scholar 

  • Jessen KR, Mirsky R (1991) Schwann cell precursor and their development. Glia 4:185–194

    Article  PubMed  CAS  Google Scholar 

  • Jessen KR, Mirsky R (2002) Signals that determine Schwann cell identity. J Anat 200:367–376

    Article  PubMed  CAS  Google Scholar 

  • Jessen KR, Mirsky R, Morgan L (1991) Role of cyclic AMP and proliferation controls in Schwann cell differentiation. Ann N Y Acad Sci 633:78–89

    PubMed  CAS  Google Scholar 

  • Kirk AD (2003) Crossing the bridge: large animal models in translational transplantation research. Immunol Rev 196:176–196

    Article  PubMed  CAS  Google Scholar 

  • Levi AD, Guénard V, Aebischer P, Bunge RP (1994) The functional characteristics of Schwann cells cultured from human peripheral nerve after transplantation into a gap within the rat sciatic nerve. J Neurosci 14:1309–1319

    PubMed  CAS  Google Scholar 

  • Li Y, Field PM, Raisman G (1998) Regeneration of adult rat corticospinal axons induced by transplanted olfactory ensheathing cells. J Neurosci 18:10514–10524

    PubMed  CAS  Google Scholar 

  • Li Y, Sauve Y, Li D, Lund RD, Raisman G (2003a) Transplanted olfactory ensheathing cells promote regeneration of cut adult rat optic nerve axons. J Neurosci 23:7783–7788

    PubMed  CAS  Google Scholar 

  • Li Y, Decherchi P, Raisman G (2003b) Transplantation of olfactory ensheathing cells into spinal cord lesions restores breathing and climbing. J Neurosci 23:727–731

    PubMed  CAS  Google Scholar 

  • Luo SF, Chiu CT, Tsao HL, Fan LW, Tsai CT, Pan SL, Yang CM (1997) Effect of forskolin on bradykinin-induced calcium mobilization in cultured canine tracheal smooth muscle cells. Cell Signal 9:159–167

    Article  PubMed  CAS  Google Scholar 

  • Mackay-Sim A (2005) Olfactory ensheathing cells and spinal cord repair. Keio J Med 1:8–14

    Article  Google Scholar 

  • Morgan L, Jessen KR, Mirsky R (1991) The effects of cAMP on differentiation of cultured Schwann cells: progression from an early phenotype (04+) to a myelin phenotype (P0+, GFAP-, N-CAM-, NGF-receptor-) depends on growth inhibition. J Cell Biol 112:457–467

    Article  PubMed  CAS  Google Scholar 

  • Pollock GS, Franceschini IA, Graham G, Marchionni MA, Barnett SC (1999) Neuregulin is a mitogen and survival factor for olfactory bulb ensheathing cells and an isoform is produced by astrocytes. Eur J Neurosci 11:769–780

    Article  PubMed  CAS  Google Scholar 

  • Radtke C, Akiyama Y, Brokaw J, Lankford KL, Wewetzer K, Fodor WL, Kocsis JD (2004) Remyelination of the nonhuman primate spinal cord by transplantation of H-transferase transgenic adult pig olfactory ensheathing cells. FASEB J 18:335–337

    PubMed  CAS  Google Scholar 

  • Raisman G (1985) Specialized neuroglial arrangement may explain the capacity of vomeronasal axons to reinnervate central neurons. Neuroscience 14:237–254

    Article  PubMed  CAS  Google Scholar 

  • Raisman G (2001) Olfactory ensheathing cells—another miracle cure for spinal cord injury? Nat Rev Neurosci 2:369–375

    Article  PubMed  CAS  Google Scholar 

  • Ramón-Cueto A, Avila J (1998) Olfactory ensheathing glia: properties and function. Brain Res Bull 46:175–187

    Article  PubMed  Google Scholar 

  • Ramón-Cueto A, Plant GW, Avila J, Bunge MB (1998) Long-distance axonal regeneration in the transected rat spinal cordis promoted by olfactory ensheathing glia transplants. J Neurosci 18:3803–3815

    PubMed  Google Scholar 

  • Reier PJ (2004) Cellular transplantation strategies for spinal cord injury and translational neurobiology. NeuroRx 4:424–451

    Article  Google Scholar 

  • Ross AH, Grob P, Bothwell M, Elder DE, Ernst CS, Marano N, Ghrist BF, Slemp CC, Herlyn M, Atkinson B (1984) Characterization of nerve growth factor receptor in neural crest tumors using monoclonal antibodies. Proc Natl Acad USA 81:6681–6685

    Article  CAS  Google Scholar 

  • Santos-Benito FF, Ramón-Cueto A (2003) Olfactory ensheathing glia transplantation: a therapy to promote repair in the mammalian central nervous system. Anat Rec 271B:77–85

    Article  Google Scholar 

  • Sasaki M, Lankford KL, Zemedkun M, Kocsis JD (2004) Identified olfactory ensheathing cells transplanted into the transected dorsal funiculus bridge the lesion and form myelin. J Neurosci 24:8485–8493

    Article  PubMed  CAS  Google Scholar 

  • Smith PM, Lakatos A, Barnett SC, Jeffery ND, Franklin RJ (2002) Cryopreserved cells isolated from the adult canine olfactory bulb are capable of extensive remyelination following transplantation into the adult rat CNS. Exp Neurol 176:402–406

    Article  PubMed  CAS  Google Scholar 

  • Stewart HJ, Eccleston PA, Jessen KR, Mirsky R (1991) Interaction between cAMP elevation, identified growth factors, and serum components in regulating Schwann cell growth. J Neurosci Res 30:346–352

    Article  PubMed  CAS  Google Scholar 

  • Van Keymeulen A, Roger PP, Dumont JE, Dremier S (2000) TSH and cAMP do not signal mitogenesis through Ras activation. Biochem Biophys Res Commun 273:154–158

    Article  PubMed  Google Scholar 

  • Vincent AJ, West AK, Chuah MI (2005) Morphological and functional plasticity of olfactory ensheathing cells. J Neurocytol 34:65–80

    Article  PubMed  Google Scholar 

  • Vroemen M, Weidner N (2003) Purification of Schwann cells by selection of p75 low affinity nerve growth factor receptor expressing cells from adult peripheral nerve. J Neurosci Methods 124:135–143

    Article  PubMed  CAS  Google Scholar 

  • Wewetzer K, Grothe C, Christ B, Seilheimer B (1997) Identification and characterization of differentiation-dependent Schwann cell surface antigens by novel monoclonal antibodies: introduction of a marker common to the non-myelin-forming phenotype. Glia 19:213–226

    Article  PubMed  CAS  Google Scholar 

  • Wewetzer K, Grothe C, Claus P (2001) In vitro expression and regulation of ciliary neurotrophic factor and its alpha receptor subunit in neonatal rat olfactory ensheathing cells. Neurosci Lett 306:165–168

    Article  PubMed  CAS  Google Scholar 

  • Wewetzer K, Verdú E, Angelov DN, Navarro X (2002) Olfactory ensheathing glia and Schwann cells: two of a kind? Cell Tissue Res 309:337–345

    Article  PubMed  Google Scholar 

  • Wewetzer K, Kern N, Ebel C, Radtke C, Brandes G (2005) Phagocytosis of O4+ axonal fragments in vitro by p75− neonatal olfactory ensheathing cells. Glia 49:557–587

    Article  Google Scholar 

  • Yan H, Bunge MB, Wood PM, Plant GW (2001) Mitogenic response of adult rat olfactory ensheathing glia to four growth factors. Glia 33:334–342

    Article  PubMed  CAS  Google Scholar 

  • Zhang M, Schleicher RL, Fink AS, Gunter-Smith P, Savard C, Nguyen T, Lee SP (2000) Growth and function of isolated canine pancreatic ductal cells. Pancreas 20:67–76

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

We thank Drs. R. Niedenthal (Hannover, Germany) S. Barnett (Glasgow, UK), and E. Shooter (Stanford, USA) for providing monoclonal anti-human-p75 antibodies, monoclonal anti-human-p75 antibodies and hybridoma cells secreting antibodies to O4, respectively. We are also grateful to Dr. M. Kietzmann (Dept. Pharmacol. Toxicol., University of Veterinary Medicine Hannover) for providing the ELISA reader and to N. van Dornick for editorial help.

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Authors and Affiliations

  1. Department of Pathology, University of Veterinary Medicine Hannover, Bünteweg 17, 30559, Hannover, Germany

    Christiane Krudewig & Konstantin Wewetzer

  2. Department of Non-clinical Drug Safety, Boehringer Ingelheim Pharma, 88397, Biberach/Riß, Germany

    Ulrich Deschl

  3. Center of Anatomy, Neuroanatomy, Hannover Medical School, 30625, Hannover, Germany

    Konstantin Wewetzer

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  1. Christiane Krudewig
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  2. Ulrich Deschl
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  3. Konstantin Wewetzer
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Correspondence to Konstantin Wewetzer.

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This study was supported by a grant from the Hochschulinterne Leistungsförderung (HiLFII, Hannover Medical School) to K. Wewetzer.

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Krudewig, C., Deschl, U. & Wewetzer, K. Purification and in vitro characterization of adult canine olfactory ensheathing cells. Cell Tissue Res 326, 687–696 (2006). https://doi.org/10.1007/s00441-006-0238-9

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  • DOI: https://doi.org/10.1007/s00441-006-0238-9

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