September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Disease in a Dish Model of Primary Open Angle Glaucoma
Author Affiliations & Notes
  • Iqbal Ahmad
    Ophthalmology & Visual Sciences, Univ of Nebraska Medical Ctr, Omaha, Nebraska, United States
  • Rand R Allingham
    Duke Eye Center, Duke, North Carolina, United States
  • Michael houser
    Duke Molecular Physiology Institute, Duke, North Carolina, United States
  • Pooja Teotia
    Ophthalmology & Visual Sciences, Univ of Nebraska Medical Ctr, Omaha, Nebraska, United States
  • Footnotes
    Commercial Relationships   Iqbal Ahmad, None; Rand Allingham, None; Michael houser, None; Pooja Teotia, None
  • Footnotes
    Support  National Eye Institute, Lincy foundation, Pearson foundation and Research to Prevent Blindness
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 6057. doi:
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    • Get Citation

      Iqbal Ahmad, Rand R Allingham, Michael houser, Pooja Teotia; Disease in a Dish Model of Primary Open Angle Glaucoma. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6057.

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

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Abstract

Purpose : Glaucoma is a complex group of diseases with a unifying theme where retinal ganglion cells (RGCs) degenerate, leading to irreversible blindness. Since RGCs are born embryonically and glaucoma is a late onset disease our knowledge about the mechanism underlying RGC susceptibility, which may facilitate early diagnosis and better therapeutic management of the disease, remains rudimentary. We are proposing to understand RGC susceptibility using a disease in a dish model of primary open angle glaucoma associated with the SIX6 risk allele (Asn141His). SIX6 is one of the primary eye field genes involved in retinal development and RGC integrity.

Methods : The model was created using the induced pluripotent stem (iPS) cell technology. Briefly, peripheral blood mononuclear cells (PBMCs) were harvested from patients’ and age matched controls’ blood samples. PBMCs were reprogrammed using the transduction of Sendai virus, expressing Yamanaka factors (Oct4, Klf4, Sox2, and c-Myc). Multiple SIX6 risk allele (Asn141His) and control iPS clones were generated for the characterization of pluripotency and derivation of RGCs by a chemical induction protocol, using small molecules and recombinant growth factors, that recapitulates the mechanism of normal RGC genesis.

Results : Both the SIX6 (Asn141His) patient-specific and control iPS colonies were morphologically similar and expressed pluripotency markers Nanog and Oct4. Further characterization of a control iPS clone, in addition to ongoing teratomas analyses, revealed that the iPS cells efficiently generated retinal progenitor cells (RPCs) either through the embryoid body- or monolayer-based culture protocols. The iPS cell-derived RPCs expressed retinal progenitor regulators, Rx, Pax6, and Chx10; their authenticity was confirmed by double immunocytochemical analysis of Rx and Pax6. The iPS cell-derived RPCs were temporally exposed to small molecules and growth factors over the period of 15 days to influence signaling pathways (Shh, FGF8, TGFb, and Notch pathways) involved in the normal genesis of RGCs. Cells emerged co-expressing RGC markers, and displaying RGC electrophysiological features.

Conclusions : Both SIX6 (Asn141His) patient and control PBMCs are re-programmed to pluripotency using the non-integrating Sendai virus transduction of Yamanaka factors approach. The reprogrammed cells are capable of differentiating into RGCs by recapitulating developmental mechanism.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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