May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
PEDF Supports the Growth of Retinal Progenitors and Embryonic Stem Cells
Author Affiliations & Notes
  • C. J. Barnstable
    Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, Pennsylvania
  • X. Liao
    Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, Pennsylvania
  • H. Li
    Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, Pennsylvania
  • J. Tombran-Tink
    Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, Pennsylvania
  • Footnotes
    Commercial Relationships  C.J. Barnstable, None; X. Liao, None; H. Li, None; J. Tombran-Tink, None.
  • Footnotes
    Support  David Woods Kemper Memorial Foundation and grants from the NIH
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 3570. doi:https://doi.org/
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    • Get Citation

      C. J. Barnstable, X. Liao, H. Li, J. Tombran-Tink; PEDF Supports the Growth of Retinal Progenitors and Embryonic Stem Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3570. doi: https://doi.org/.

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

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Abstract

Purpose: : Control of the proliferation and differentiation of stem and progenitor cells is essential for their effective use in any form of regenerative medicine. Pigment epithelium derived factor (PEDF) is a potent neurotrophic factor that is expressed early in embryogenesis and can maintain the self-renewal properties of stem cells in the subventricular zone of mature animals. In this study we have examined the effects of PEDF and other factors on retinal progenitors and mouse embryonic stem cells.

Methods: : Retinal progenitors were isolated by plating dissociated cells in either DMEM or Ultraculture medium in non-adherent plastic dishes. Mouse embryonic stem cells were routinely cultured in DMEM supplemented with FBS and LIF (1000U/ml). For immunocytochemistry cells were fixed in freshly prepared cold Methanol:Acetic acid solution for 10 min. After blocking for 30 minutes with 25% horse serum in PBS, cells were incubated for 1 hour at room temperature in 25% horse serum in PBS with anti-Nanog, Oct4 and Pax6 antibodies, and for 30 minutes with Cy3-conjugated second antibody. For immunoblot analysis cell lysates were prepared and Western blot was performed. Immunoblots were developed using HRP-conjugated goat anti-mouse or -rabbit IgG, followed by chemiluminescent detection.

Results: : 100 ng/ml PEDF substantially increased the number of neurospheres formed from retinal cell preparations. VEGF and FGF did not increase the number of neurospheres but did increase the size. Immunocytochemical analysis of the neurospheres showed a core of Nanog, Oct4, CD44 and CD133 positive cells surrounded by a shell of differentiated progeny. Passaging of retinal progenitors was also more efficient in the presence of PEDF. PEDF was able to support the continued growth of mouse ES cells in DMEM and was as effective as LIF, the standard supplement. Both immunocytochemical and Western blot analysis showed that PEDF maintained expression of Nanog, Oct4 and Pax6. PEDF rapidly induced phosphorylation of Akt, Erk1/2, Stat3 and p38 in both retinal progenitors and ES cells. Peak phosphorylation occurred 15 min after initiation of treatment.

Conclusions: : PEDF activates a number of key signal transduction pathways in stem cells. It is effective at maintaining the self-renewal properties of ES cells and also improves the survival and growth of retinal progenitor cells. Use of well-defined factors such as PEDF will be an important tool in the development of stem cell technology to treat ocular diseases.

Keywords: retinal development • growth factors/growth factor receptors • regeneration 
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