June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Extracellular vesicles purified from porcine vitreous positively regulate retinal ganglion cell axon survival and growth
Author Affiliations & Notes
  • Yolandi van der Merwe
    Department of Bioengineering, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Anne Faust
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Bianca Leonard
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Mark Curtis
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    Center for Neuroscience, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Apoorva Kandakatla
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Kevin C Chan
    Department of Bioengineering, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Michael Steketee
    Department of Ophthalmology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
    Center for Neuroscience, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
  • Footnotes
    Commercial Relationships   Yolandi van der Merwe, None; Anne Faust, None; Bianca Leonard, None; Mark Curtis, None; Apoorva Kandakatla, None; Kevin Chan, None; Michael Steketee, None
  • Footnotes
    Support  Department of Defense office of the Congressionally Directed Medical Research Programs and the Clinical and Rehabilitative Medicine Research Program (MBS, W81XWH-15-1-0026), Start-up funds, Department of Ophthalmology, University of Pittsburgh (MBS), National Institutes of Health CORE Grant P30 EY008098, Eye and Ear Foundation of Pittsburgh, PA, Unrestricted Grant from Research to Prevent Blindness, New York, NY
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 1754. doi:
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      Yolandi van der Merwe, Anne Faust, Bianca Leonard, Mark Curtis, Apoorva Kandakatla, Kevin C Chan, Michael Steketee; Extracellular vesicles purified from porcine vitreous positively regulate retinal ganglion cell axon survival and growth. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1754.

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

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Abstract

Purpose : Retinal injury can lead to retinal ganglion cell (RGC) axon degeneration and ultimately RGC death due to the default healing response in the CNS. Extracellular vesicles (EV) are emerging as naturally-derived therapeutics with the potential to modulate the default healing response by positively regulating the phenotypes of multiple cell types simultaneously. EVs are released from all cells and carry unique bioactive cargoes that reflect the state of the cell from which they are released, which during injury or disease tends to potentiate inflammatory signaling and secondary trauma. We hypothesize EVs derived from healthy porcine vitreous can increase RGC survival and neurite growth in vitro and neuroprotect injured RGCs in vivo

Methods : Vitreous vesicles (VV) were isolated from collagenase treated porcine vitreous by differential centrifugation, visualized by transmission electron microscopy (TEM), analyzed for exosome markers by flow cytometry, and miRNA content by sequencing. In vitro, primary RGCs from P3 Sprague-Dawley rats were cultured with VVs. After 3 days, viability and neurite growth were analyzed. In vivo, intraocular pressure was elevated to 130mmHg for 60 minutes via anterior chamber perfusion with or without VV injection 24 hours after injury. After 7 days, RGC survival was analyzed by TUNEL

Results : TEM showed relatively uniform VVs, ranging in size from 10-30 nm (Fig. 1). Flow cytometry showed VVs were positive for exosome markers, including CD63 but not CD81. In vitro, fluorescent VVs were internalized within minutes into RGC cell bodies, neurites, and growth cones. VVs increased RGC viability and neurite growth (Fig. 1). In vivo, intravitreally injected VVs were internalized by glial and RGCs and RGC survival is being analyzed. VVs contain various miRNAs known regulate cellular differentiation and apoptotic signaling

Conclusions : Porcine VVs are uniform in size, have exosome markers, and carry unique miRNA cargoes. In vitro, VVs increase primary RGC survival and growth. In vivo, VVs integrate into multiple layers of the retina. We are currently investigating the in vivo effects of VVs on RGC survival in injury models and the roles that VV miRNAs play in regulating RGC survival and growth. These initial studies indicate VVs hold promise as biologically derived neuroprotective therapeutics for retinal injury

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

 

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