September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Investigating the molecular mechanisms of Late-Onset Retinal Degeneration using patient-specific induced pluripotent stem cells
Author Affiliations & Notes
  • Zoya Qureshy
    Unit on Ocular Stem Cell and Translational Research, National Eye Institute, National Institutes of Health, Bethesda , Maryland, United States
  • Kiyoharu Miyagishima
    Section on Epithelial & Retinal Physiology & Disease, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States
  • Katharina Clore-Gronenborn
    Unit on Ocular Stem Cell and Translational Research, National Eye Institute, National Institutes of Health, Bethesda , Maryland, United States
  • Congxiao Zhang
    Section on Epithelial & Retinal Physiology & Disease, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States
  • Ruchi Sharma
    Unit on Ocular Stem Cell and Translational Research, National Eye Institute, National Institutes of Health, Bethesda , Maryland, United States
  • Vaishakh Rajan
    Unit on Ocular Stem Cell and Translational Research, National Eye Institute, National Institutes of Health, Bethesda , Maryland, United States
  • Vladimir Khristov
    Section on Epithelial & Retinal Physiology & Disease, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States
  • Catherine Cukras
    Clinical Trials Branch, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States
  • Sheldon S Miller
    Section on Epithelial & Retinal Physiology & Disease, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States
  • Kapil Bharti
    Unit on Ocular Stem Cell and Translational Research, National Eye Institute, National Institutes of Health, Bethesda , Maryland, United States
  • Footnotes
    Commercial Relationships   Zoya Qureshy, None; Kiyoharu Miyagishima, None; Katharina Clore-Gronenborn, None; Congxiao Zhang, None; Ruchi Sharma, None; Vaishakh Rajan, None; Vladimir Khristov, None; Catherine Cukras, None; Sheldon Miller, None; Kapil Bharti, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science September 2016, Vol.57, No Pagination Specified. doi:
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      Zoya Qureshy, Kiyoharu Miyagishima, Katharina Clore-Gronenborn, Congxiao Zhang, Ruchi Sharma, Vaishakh Rajan, Vladimir Khristov, Catherine Cukras, Sheldon S Miller, Kapil Bharti; Investigating the molecular mechanisms of Late-Onset Retinal Degeneration using patient-specific induced pluripotent stem cells. Invest. Ophthalmol. Vis. Sci. 201657(12):.

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

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Abstract

Purpose : Late-Onset Retinal Degeneration (L-ORD) is a rare autosomal dominant disorder that shares clinical similarities to more prevalent retinal degenerations, such as age-related macular degeneration. L-ORD is caused by a single S163R amino acid substitution in the globular domain of the CTRP5 protein. The CTRP5 gene is contained within the 3’-untranslated region of another gene, which encodes the membrane-frizzled related protein (MFRP). Investigating this genetically simple disease will provide us with valuable insight into the shared mechanisms of other retinal degenerative diseases and help us identify novel targets for therapy.

Methods : We generated induced pluripotent stem cells (iPSCs) from fibroblasts of two L-ORD-affected and two unaffected siblings. iPSCs were characterized by the expression of pluripotent markers: SSEA4, SOX2, OCT4, and NANOG. Karyotype analysis revealed no aberrations. Sequencing confirmed that the patients’ cells retained the S163R point mutation characteristic of L-ORD. iPSCs were differentiated into RPE using a directed differentiation protocol. CTRP5 and MFRP expression levels were assessed by qPCR, Western blot, and immunostaining. Intracellular calcium levels were measured with ratiometric calcium dye, Fura-2.

Results : RPE differentiated from iPSCs of L-ORD patients and their healthy sibling express similar amounts of CTRP5 protein. mRNA expression of MFRP is also not significantly different between patients and healthy cells. However, patient’s cells show characteristic deposits that are reminiscent of deposits seen in the eyes of L-ORD patients. Calcium imaging results demonstrate lower baseline intracellular calcium levels in patient cells as compared to their healthy siblings.

Conclusions : L-ORD patient RPE cells show cellular endophenotypes that are reminiscent of phenotype seen in the eyes of L-ORD patients. Reduced baseline calcium in L-ORD patient cells suggests defective calcium homeostasis that might contribute to accumulation of intra and sub-cellular deposits.

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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