April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Adherent Differentiation of Human Embryonic Stem Cells Promotes Generation of Retinal Pigment Epithelial Cells
Author Affiliations & Notes
  • H. Vaajasaari
    Regea - Inst. for Regenerative Medicine, University of Tampere, Tampere, Finland
  • T. Ilmarinen
    Regea - Inst. for Regenerative Medicine, University of Tampere, Tampere, Finland
  • H. Hongisto
    Regea - Inst. for Regenerative Medicine, University of Tampere, Tampere, Finland
  • R. Suuronen
    Regea - Inst. for Regenerative Medicine, University of Tampere, Tampere, Finland
    Tampere University Hospital, Department of Eye, Ear and Oral Diseases, Finland
  • H. Uusitalo
    Medical School, SILK, University of Tampere, Tampere, Finland
  • H. Skottman
    Regea - Inst. for Regenerative Medicine, University of Tampere, Tampere, Finland
  • Footnotes
    Commercial Relationships  H. Vaajasaari, None; T. Ilmarinen, None; H. Hongisto, None; R. Suuronen, None; H. Uusitalo, None; H. Skottman, None.
  • Footnotes
    Support  Academy of Finland, the Competitive Research Funding of the Tampere University Hospital, TGSBB,Evald and Hilda Nissi Foundation, Sokeain ystävät Foundation and two private donations
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 2658. doi:https://doi.org/
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      H. Vaajasaari, T. Ilmarinen, H. Hongisto, R. Suuronen, H. Uusitalo, H. Skottman; Adherent Differentiation of Human Embryonic Stem Cells Promotes Generation of Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2658. doi: https://doi.org/.

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

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Abstract

Purpose: : Human embryonic stem cell (hESC) derived retinal pigment epithelial (RPE) cells can provide a cell model for various retinal diseases or drug discovery and essentially, a regenerative cure for vision-threatening diseases. The utilization of hESC derived RPE cells for these purposes requires an efficient differentiation method. In this study, the efficiency of the adherent differentiation was compared to suspension (EB) differentiation in the generation of the RPE cells from hESCs.

Methods: : Human ESCs (Regea 08/023) were derived and maintained without serum and animal feeder cells. The differentiation was performed by plating the hESCs on protein coating or by culturing them as cell aggregates in suspension using a serum-free medium without any growth factors. The differentiation efficiency of the methods was compared by using qRT-PCR. The maturation of the cells was confirmed by RT-PCR and immunocytochemistry.

Results: : First pigmented cells were observed by day 14 with both differentiation methods. qRT-PCR data demonstrated that the adherent differentiation of the hESCs enhanced the expression of the essential genes for RPE development (PAX6, RAX, SIX3, MITF) compared to EB differentiation. Furthermore, the expression of neural retinal marker, CHX10, was down-regulated in adherent differentiation compared to EB differentiation. The RPE cell maturation was confirmed at gene level (MITF, RPE65, BEST, PEDF, PMEL, tyrosinase) and at protein level (MITF, CRALB, Bestrophin, ZO1).

Conclusions: : The adherent differentiation method was more efficient for the generation of RPE cells than the EB method. The constituents responsible for the differences between the two methods are currently under investigation.

Keywords: retinal pigment epithelium • retinal development • age-related macular degeneration 
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