April 2011
Volume 52, Issue 14
ARVO Annual Meeting Abstract  |   April 2011
Retinal Progenitor Cells Acquire Tri-potentiality Following In vitro Expansion
Author Affiliations & Notes
  • Magdalena W. Czekaj
    Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany
  • Jochen Haas
    Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany
  • Jane Farrar
    AG Ocular Genetics Unit, Smurfit Institute of Genetics, Trinity College Dublin, Dublin 2, Ireland
  • Udo Bartsch
    Department of Ophthalmology, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany
  • Marius Ader
    Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany
  • Footnotes
    Commercial Relationships  Magdalena W. Czekaj, None; Jochen Haas, None; Jane Farrar, None; Udo Bartsch, None; Marius Ader, None
  • Footnotes
    Support  CRTD
Investigative Ophthalmology & Visual Science April 2011, Vol.52, 2228. doi:
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      Magdalena W. Czekaj, Jochen Haas, Jane Farrar, Udo Bartsch, Marius Ader; Retinal Progenitor Cells Acquire Tri-potentiality Following In vitro Expansion. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2228.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: : One of the treatment strategies proposed for retinal degenerations is to replace lost neurons with cells expanded in vitro, primed to specific neuronal fates that after transplantation would integrate into the retina and restore its visual function. Here we investigated the differentiation potential of in vitro expanded retinal stem cells (RSCs).

Methods: : RSC cultures were generated by isolating retinal cells from E14.5 and PN0 actin:GFP or actin:dsRed mice and expanded in medium containing N2, EGF and FGF2. For in vitro oligodendrocyte differentiation primary as well as expanded RSCs from early and late passages were subjected to a stepwise replacement of EGF and FGF2 by forskolin, PDGFα, ascorbic acid and triiodothyonine and characterized by immunocytochemistry and RT-PCR. We also performed intravitreal transplantation experiments of expanded and primed RSCs into the eyes of adult wild-type mice. 4 weeks after grafting dissected retinae were either subjected to transmission electron microscopy or flat-mounted/sectioned retinae were analyzed by fluorescence microscopy following immunohistochemistry.

Results: : Retinal cells upon in vitro expansion maintain the potential to generate glial and neuronal cell types (GFAP- and β-III-tubulin-positive, respectively). However, RT-PCR analysis shows that they loose expression of some genes that are characteristic for retinal progenitors. In contrast to primary cells, RSCs are able to respond to oligodendrocyte priming treatment with expression of oligodendrocyte markers, as shown by immunocytochemistry and RT-PCR analyses, and following transplantation develop MBP-positive and ultra-structurally normal myelin sheaths around RGC axons.

Conclusions: : RSC cultures can be propagated over several passages and maintain the ability to differentiate along the glial and neuronal lineages. However, they do not differentiate into true retinal neurons and loose expression of genes characteristic for retinal progenitors. With increasing passages the expanded RSCs acquire the capacity to express proteins implicated in myelin formation and after transplantation differentiate into oligodendrocytes, a cell type that is not generated by primary retinal progenitors in vivo. Thus, we conclude that culture conditions lead to a loss of regional identity and conversion of RSCs into tri-potential neural stem cells.

Keywords: retinal culture • transplantation 

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