September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Redox Status of Wallerian Degeneration Slow (WldS) and Wild-Type (WT) Axons Following Single Cell Laser Axotomy of Cultured Retinal Ganglion Cells (RGCs)
Author Affiliations & Notes
  • Mohammadali Almasieh
    Centre de recherche de l’Hôpital Maisonneuve-Rosemont (CRHMR) & Department of Ophthalmology, Université de Montréal, Montreal, Quebec, Canada
    Ophthalmology, McGill University, Montreal, Quebec, Canada
  • Maria-Magdalena Catrinescu
    Centre de recherche de l’Hôpital Maisonneuve-Rosemont (CRHMR) & Department of Ophthalmology, Université de Montréal, Montreal, Quebec, Canada
    Ophthalmology, McGill University, Montreal, Quebec, Canada
  • Loïc Binan
    Centre de recherche de l’Hôpital Maisonneuve-Rosemont (CRHMR) & Department of Ophthalmology, Université de Montréal, Montreal, Quebec, Canada
  • Santiago Costantino
    Centre de recherche de l’Hôpital Maisonneuve-Rosemont (CRHMR) & Department of Ophthalmology, Université de Montréal, Montreal, Quebec, Canada
  • Leonard A Levin
    Centre de recherche de l’Hôpital Maisonneuve-Rosemont (CRHMR) & Department of Ophthalmology, Université de Montréal, Montreal, Quebec, Canada
    Ophthalmology, McGill University, Montreal, Quebec, Canada
  • Footnotes
    Commercial Relationships   Mohammadali Almasieh, None; Maria-Magdalena Catrinescu, None; Loïc Binan, None; Santiago Costantino, None; Leonard Levin, None
  • Footnotes
    Support  CIHR MOP 112357
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 2538. doi:
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      Mohammadali Almasieh, Maria-Magdalena Catrinescu, Loïc Binan, Santiago Costantino, Leonard A Levin; Redox Status of Wallerian Degeneration Slow (WldS) and Wild-Type (WT) Axons Following Single Cell Laser Axotomy of Cultured Retinal Ganglion Cells (RGCs)
      . Invest. Ophthalmol. Vis. Sci. 2016;57(12):2538.

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      © 2017 Association for Research in Vision and Ophthalmology.

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Abstract

Purpose : Glaucoma is characterized by axonal degeneration and loss of RGCs. Understanding the mechanisms that lead to axonal degeneration is essential to delay RGC loss and preserve vision in glaucoma. The delayed axonal degeneration in WldS mutant animals can provide insight to such mechanisms. Here, we tested the hypothesis that initiation and progression of axonal degeneration involves the critical step of oxidative modification of sulfhydryls contained within the axon, and that its progression down the axon is significantly different in WldS rats compared to wild-type.

Methods : Retina suspensions from postnatal wild-type and WldS rat pups were immunopanned using anti-Thy-1 to obtain purified primary RGC cultures. RGCs cultured in laminin-coated dishes were allowed to extend neurites until a dominant process with a typical axon hillock was identified. A two-photon laser was used to induce single-cell axotomy. Axonal membrane degeneration was monitored by time-lapse fluorescent imaging.

Results : Following axonal damage, there was a significant delay in detection of the polarity-sensitive thiol-reactive fluorogenic marker annexin B12-IANBD signal in WldS axons compared to wild-type. The velocity of the spread of the fluorescent signal along the segments proximal and distal to the axotomy was significantly slower for WldS compared to wild-type. When the optimized cell-permeable disulfide reducing agent bis(3-propionic acid methyl ester) phenylphosphine borane complex was added to the medium, it significantly lowered the velocity of the spread of fluorescence in axotomized wild-type RGC axons.

Conclusions : The spread of axonal degeneration after axotomy can be detected by fluorescenct imaging of annexin B12-IANBD. The significant delay in the initiation and velocity of this spread in WldS axons, which is partly replicated by application of a cell-permeable disulfide-reducing agent, suggests a role for a redox signal in axonal degeneration after axotomy. This might provide a basis for a therapeutic strategy to delay axonal degeneration in glaucoma.

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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