July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Identification of new survival pathways activated by neurotrophic factors using whole transcriptome analysis of flow-sorted photoreceptors.
Author Affiliations & Notes
  • Jerome E Roger
    NEUROPSI, CERTO / CNRS, Orsay, France
  • Elodie-Kim Grellier
    NEUROPSI, CERTO / CNRS, Orsay, France
  • Linn Gieser
    N-NRL, NEI / NIH, Bethesda, Maryland, United States
  • Anand Swaroop
    N-NRL, NEI / NIH, Bethesda, Maryland, United States
  • Muriel Perron
    NEUROPSI, CERTO / CNRS, Orsay, France
  • Footnotes
    Commercial Relationships   Jerome Roger, None; Elodie-Kim Grellier, None; Linn Gieser, None; Anand Swaroop, None; Muriel Perron, None
  • Footnotes
    Support  Retina France.; NEI Intramural Research Program EY000450 and EY000546
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4886. doi:https://doi.org/
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      Jerome E Roger, Elodie-Kim Grellier, Linn Gieser, Anand Swaroop, Muriel Perron; Identification of new survival pathways activated by neurotrophic factors using whole transcriptome analysis of flow-sorted photoreceptors.. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4886. doi: https://doi.org/.

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

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Abstract

Purpose : Retinal degenerative diseases are clinically and genetically heterogeneous, with a vast majority leading to loss of photoreceptors. Neuroprotective strategies are promising because they have the advantage of being mutation independent. We hypothesize that specific molecules and signaling pathways, yet to be identified, shall be commonly used by neurotrophic factors to preserve photoreceptors. Our goal is to identify downstream targets regulated by different neurotrophic factors specifically in photoreceptors. We first focused on CNTF target genes, a well-known neurotrophic factor.

Methods : CNTF intraocular injections were performed in Nrl-GFP wild-type (WT) and Nrl-GFP, Pde6brd10/rd10 (rd10) mice. Intravitreal injection efficiency was controlled by immunohistochemistry, qPCR and immunoblot analysis. RNAs were purified from flow-sorted photoreceptors (FSPR) collected at 1, 3 and 5 days after CNTF injection, followed by whole-transcriptome sequencing. After mapping with STAR, differentially expressed genes (DEGs) were identified using EdgeR between CNTF and PBS-injected retina in WT and rd10 groups. Pathway analysis was performed using Panther database. Additional factors were also injected (Edn2, Pedf, Gdnf and Bdnf) for candidate gene validation by qPCR.

Results : As predicted, the expression of CNTF target genes (e.g. Gfap) was increased, reflecting reactive gliosis in Muller cells following CNTF injection. Maximum upregulation of Gfap expression was observed after 3 days, followed by a decrease at 5 days. Whole transcriptome analysis of FSPR revealed 252 DEGs (Fold Change of at least 1.5, FDR lower than 0.05). Among known CNTF-photoreceptor target genes, Fgf2 and Edn2 were upregulated. Hierarchical clustering allowed the identification of clusters of upregulated genes following CNTF injection with even higher induction in rd10. Pathway analysis from these 160 DEGs revealed enriched biological processes among others related to regulation of cell death (28 DEGs) and to response to toxic substance (10 DEGs). Importantly, some of these candidate genes were also induced by other neurotrophic factors (Edn2, Pedf, Gdnf and Bdnf) injected in WT and rd10 mice.

Conclusions : Our study revealed promising novel candidate factor that could potentially prevent photoreceptor cell death. We will next validate their neuroprotective effects in rd10 mice.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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