September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Photoreceptor structure and gene expression in the canine NPHP5-LCA ciliopathy model
Author Affiliations & Notes
  • Louise M Downs
    University of Pennsylvania, School of Vetinary Medicine, Philadelphia, Pennsylvania, United States
  • Erin M Scott
    University of Pennsylvania, School of Vetinary Medicine, Philadelphia, Pennsylvania, United States
  • Sem Genini
    University of Pennsylvania, School of Vetinary Medicine, Philadelphia, Pennsylvania, United States
  • William A Beltran
    University of Pennsylvania, School of Vetinary Medicine, Philadelphia, Pennsylvania, United States
  • Gustavo D Aguirre
    University of Pennsylvania, School of Vetinary Medicine, Philadelphia, Pennsylvania, United States
  • Footnotes
    Commercial Relationships   Louise Downs, None; Erin Scott, None; Sem Genini, None; William Beltran, None; Gustavo Aguirre, None
  • Footnotes
    Support  EY-06855, -17549, the Foundation Fighting Blindness, the Macula Vision Research Foundation, the Van Sloun Fund for Canine Genetic Research, and Hope for Vision.
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 3188. doi:
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    • Get Citation

      Louise M Downs, Erin M Scott, Sem Genini, William A Beltran, Gustavo D Aguirre; Photoreceptor structure and gene expression in the canine NPHP5-LCA ciliopathy model. Invest. Ophthalmol. Vis. Sci. 2016;57(12):3188.

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

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Abstract

Purpose : A cytosine insertion in exon 10 of NPHP5 (IQCB1) results in an early onset, aggressive photoreceptor (PR) degeneration, a model of LCA in man (NPHP5-LCA). This study characterized the kinetics of PR loss, the progression of structural degeneration and the expression profiles of genes specific to PR and of signalling pathways involved in PR degeneration.

Methods : Retinas were obtained from 7 affected and control dogs at 6, 14, 31 and 42 weeks, embedded in OCT and examined with H&E and immunohistochemistry. Gene expression of rod and/or cone genes, and genes involved in pro-death and pro-survival pathways, was compared at 5 weeks of age. The ddCt method, using GAPDH for normalization, and an unpaired t-test were applied to identify differentially expressed genes (p<0.05 and FC>2).

Results : At 6 weeks of age, ONL thickness and the number of cone somas in affected dogs was similar to controls, although many other hallmarks of degeneration were observed in the affected dogs: Cones had shortened and distorted IS, and most had lost their OS; many PR underwent significant apoptosis or proliferation; and rod opsin mislocalized to the ONL and beyond. Retinal degeneration continued through the later time points and by 42 weeks the ONL was only 1-2 nuclei thick. At 14 weeks apoptosis and proliferation had dropped to a fraction of the level observed at 6 weeks, and remained at that level through 31 and 42 weeks. The number of cone somas counted remained similar to controls until 31 weeks, but by 42 weeks a sharp reduction was observed in the affected dogs. In contrast, by 14 weeks there were virtually no cone OS at all; by 42 weeks, a very small number of cone OS were present temporally near the area centralis. Connecting cilium (CC) marker labelling was observed at 6 and 14 weeks in the affected retina, as was PNA labelling of the cone sheaths, despite the absence of cone OS. qRT-PCR revealed that rod- or rod-and-cone-specific genes were downregulated at 5 weeks, but cone-specific genes were not. A number of pro-death or pro-survival genes were also down-regulated at 5 weeks.

Conclusions : Degeneration of rod and cone PR is observed from 6 weeks of age, and progresses until almost complete by 42 weeks. The relative preservation of the temporal region, and the remaining presence of CC and cone sheaths suggest that this model may be particularly suited to corrective gene augmentation therapy.

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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