June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
α-Synuclein modulates extracellular matrix proteins in the trabecular meshwork
Author Affiliations & Notes
  • Neena Singh
    Case Western Reserve University, Cleveland, Ohio, United States
  • Alexander Kritikos
    Case Western Reserve University, Cleveland, Ohio, United States
  • Dallas McDonald
    Case Western Reserve University, Cleveland, Ohio, United States
  • Sachin Pillai
    Case Western Reserve University, Cleveland, Ohio, United States
  • Shu Chen
    Case Western Reserve University, Cleveland, Ohio, United States
  • Min H Kang
    Case Western Reserve University, Cleveland, Ohio, United States
  • Douglas Rhee
    Case Western Reserve University, Cleveland, Ohio, United States
  • Ewald Lindner
    Case Western Reserve University, Cleveland, Ohio, United States
  • Ajay Ashok
    Case Western Reserve University, Cleveland, Ohio, United States
  • Footnotes
    Commercial Relationships   Neena Singh None; Alexander Kritikos None; Dallas McDonald None; Sachin Pillai None; Shu Chen None; Min Kang None; Douglas Rhee None; Ewald Lindner None; Ajay Ashok None
  • Footnotes
    Support  NIH Grant NS092145 and NS125228
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 3309 – A0409. doi:
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      Neena Singh, Alexander Kritikos, Dallas McDonald, Sachin Pillai, Shu Chen, Min H Kang, Douglas Rhee, Ewald Lindner, Ajay Ashok; α-Synuclein modulates extracellular matrix proteins in the trabecular meshwork. Invest. Ophthalmol. Vis. Sci. 2022;63(7):3309 – A0409.

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

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Abstract

Purpose : α-Synuclein (α-Syn) is implicated in Parkinson’s disease (PD), a neuro-motor disorder with prominent ocular symptoms. Here, we explored whether α-Syn plays a functional role in the anterior segment of the eye, and whether its dysfunction due to aggregation contributes to ocular pathology.

Methods : Primary human trabecular meshwork (TM) cells, cadaveric human and bovine TM tissue and aqueous humor (AH), and human AH collected during cataract surgery were used for this study. Silencing of α-Syn and overexpression of TGFβ2 in TM cells and tissue was achieved by conventional techniques. Expression of fibronectin (FN), α-smooth muscle actin (α-SMA), and ROCK-1 was determined by Western blotting (WB). Binding, uptake, and response of TM cells to extracellular α-Syn was determined by immunostaining and WB.

Results : α-Syn was expressed in the ciliary body and TM, and monomeric and oligomeric α-Syn was present in the AH. Silencing of α-Syn in TM cells downregulated FN, α-SMA, and ROCK-1 expression in the absence and presence of active TGFβ2. Extracellular monomeric and oligomeric α-Syn was internalized by β1-integrin, and upregulated FN, α-SMA, and ROCK-1 in TM cells and TM tissue. Active TGFβ2 was also increased.

Conclusions : Intracellular α-Syn altered the expression of extracellular matrix (ECM) proteins in TM cells through ROCK-1 independently and downstream of TGFβ2, the principal trigger for dysregulation of ECM in primary open angle glaucoma (POAG). Extracellular α-Syn was endocytosed by TM cells through β1-integrin, and likewise, upregulated ECM proteins. These observations have significant implications for POAG, and warrant exploration of the underlying biochemical pathways.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

 

Silencing of α-Syn downregulates ECM proteins in primary human TM cells: (A) Silencing of α-Syn in TM cells downregulates FN, α-SMA, and ROCK-1. (B) Densitometry confirms these results. (C & D) Silencing of α-Syn downregulates FN, α-SMA, and ROCK-1 in the absence and presence of over-expressed TGFβ2.

Silencing of α-Syn downregulates ECM proteins in primary human TM cells: (A) Silencing of α-Syn in TM cells downregulates FN, α-SMA, and ROCK-1. (B) Densitometry confirms these results. (C & D) Silencing of α-Syn downregulates FN, α-SMA, and ROCK-1 in the absence and presence of over-expressed TGFβ2.

 

Uptake of recombinant α-Syn increases FN, α-SMA, ROCK-1, and active TGFβ2 in primary human TM cells: (A) Exposure of TM cells to preformed fibrils (PFF) of α-Syn increases the expression of FN, α-SMA, and ROCK-1 in a dose-dependent manner relative to controls. Active TGFβ2 is increased as well. (B) Densitometric analysis confirmes these results.

Uptake of recombinant α-Syn increases FN, α-SMA, ROCK-1, and active TGFβ2 in primary human TM cells: (A) Exposure of TM cells to preformed fibrils (PFF) of α-Syn increases the expression of FN, α-SMA, and ROCK-1 in a dose-dependent manner relative to controls. Active TGFβ2 is increased as well. (B) Densitometric analysis confirmes these results.

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