March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Directing Virus-free Human Induced Pluripotent Stem Cells To Differentiate Into Retinal Cells
Author Affiliations & Notes
  • Xiufeng Zhong
    Wilmer Eye Inst, Johns Hopkins Univ Sch, Baltimore, Maryland
  • Christopher Hampton
    Wilmer Eye Inst, Johns Hopkins Univ Sch, Baltimore, Maryland
  • Tea Soon Park
    Institute for Cell Engineering, Johns Hopkins Univ Sch, Baltimore, Maryland
  • David M. Gamm
    Stem Cell Research Program at Waisman Center and Ophthalmology and Visual Sciences, University of Wisconsin-Madison, Madison, Wisconsin
  • Elias Zambidis
    Institute for Cell Engineering, Johns Hopkins Univ Sch, Baltimore, Maryland
  • Valeria Canto-Soler
    Wilmer Eye Inst, Johns Hopkins Univ Sch, Baltimore, Maryland
  • Footnotes
    Commercial Relationships  Xiufeng Zhong, None; Christopher Hampton, None; Tea Soon Park, None; David M. Gamm, None; Elias Zambidis, None; Valeria Canto-Soler, None
  • Footnotes
    Support  TEDCO and NIH EY004859, FFB WG-TRAP Award, Lincy Foundation
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 5910. doi:
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      Xiufeng Zhong, Christopher Hampton, Tea Soon Park, David M. Gamm, Elias Zambidis, Valeria Canto-Soler; Directing Virus-free Human Induced Pluripotent Stem Cells To Differentiate Into Retinal Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):5910.

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

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Abstract

Purpose: : Human induced pluripotent stem cells (iPS) hold great promise for cell-based regenerative interventions aimed at repairing the retina, but their use in cell therapy is still far from feasible. One of the major limitations for the use of iPS cells in therapeutic applications is the integration of viral vectors in the host genome, which might lead to unpredictable genetic dysfunction and increased risk of tumorigenicity. Therefore, a key step for future clinical applications is the establishment of efficient protocols to induce virus-free human iPS cells to differentiate into the cell-type of interest. The purpose of this study is to test the potential of virus-free hiPS cells to differentiate into retinal cells.

Methods: : A cell line, 6.2-CB-hiPS, established by viral-free technology was cultured and maintained on MatriGel with mTeSRTM1 medium. The IMR90-C4 cell line, established by viral technology (WiCell) and known to efficiently differentiate into retinal cells, was used as control. The procedure for inducing differentiation of hiPS cells toward a retinal fate was based on a previously described protocol with some modifications. Differentiating hiPS cells were followed by morphological observation under inverted microscope every other day and characterized by immunocytochemistry and RT-PCR with molecular markers of pluripotency and/or specific for different stages of retinal cell differentiation.

Results: : 6.2 -CB-hiPS cells were morphologically similar to IMR90-C4 hiPS counterpart, formed sharp-edged, flat, tightly-packed colonies, and expressed molecular markers specific for undifferentiated hiPS cells, such as SSEA-4, OCT4 and TRA-1-60. Under specific differentiation conditions, hiPS cells lost expression of the pluripotency genes and gradually acquired expression of molecular markers characteristic of different stages of retinal cell differentiation, as well as of specific retinal cell types, such as retinal pigment epithelium (RPE) and photoreceptors. Interestingly, the timeframe for acquisition of retinal cell fates is very close to that of IMR90-C4 hiPS cells and human retina embryogenesis.

Conclusions: : Our results indicate that virus-free hiPS cells can be efficiently directed to differentiate into retinal cell types. The efficiency and timeframe for acquisition of retinal cells is at least the same as that of IMR90-C4 hiPS cells. This study provides promising hope for the potential development of safe clinical applications of hiPS cells in retinal degenerative diseases in the future.

Keywords: retinal development • differentiation • gene/expression 
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