June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Extensive Axonal Outgrowth and Pathfinding from Retinal Ganglion Cells Derived From Human Pluripotent Stem Cells
Author Affiliations & Notes
  • Clarisse Fligor
    Biology, IUPUI, Indianapolis, Indiana, United States
  • Akshayalakshmi Sridhar
    Biology, IUPUI, Indianapolis, Indiana, United States
  • Valentin Sluch
    Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States
  • Donald J Zack
    Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States
  • Daniel Suter
    Biology, Purdue University, West Lafayette, Indiana, United States
  • Jason S Meyer
    Biology, IUPUI, Indianapolis, Indiana, United States
    Stark Neurosciences Research Institute, Indiana University, Indianapolis, Indiana, United States
  • Footnotes
    Commercial Relationships   Clarisse Fligor, None; Akshayalakshmi Sridhar, None; Valentin Sluch, None; Donald Zack, None; Daniel Suter, None; Jason Meyer, Indiana University (P), Wisconsin Alumni Research Foundation (P)
  • Footnotes
    Support  NEI R01 EY4693837
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 1370. doi:
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      Clarisse Fligor, Akshayalakshmi Sridhar, Valentin Sluch, Donald J Zack, Daniel Suter, Jason S Meyer; Extensive Axonal Outgrowth and Pathfinding from Retinal Ganglion Cells Derived From Human Pluripotent Stem Cells. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1370.

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

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Abstract

Purpose : Retinal ganglion cells (RGCs) serve as a vital connection between the eye and the brain and as such, degenerative diseases and injuries which cause the loss of RGCs often lead to blindness. Human pluripotent stem cells (hPSCs) are attractive candidates for cell replacement therapies, due to the ability to direct their differentiation into any desired cell type, including RGCs. However, in order for successful replacement of RGCs to occur, axons must extend across significant distances to reach their targets. Thus, experiments were designed to test the outgrowth and pathfinding of hPSC-derived RGCs in response to a variety of extrinsic cues.

Methods : hPSCs were differentiated into RGCs following previously established protocols. Initially, RGCs were characterized by immunocytochemistry for the expression of several RGC-associated markers. Subsequently, assays were developed to test the degree of neurite outgrowth from these RGCs. Furthermore, the ability to enhance RGC neurite outgrowth was analyzed in response to multiple factors, including varying substrates, culture media and growth factors.

Results : hPSC-derived RGCs were preliminarily identified by the expression of BRN3, which was found to co-localize with the expression of many other RGC-associated markers. Enriched populations of RGCs were isolated and plated to allow for neurite outgrowth, with significant outgrowth observed within the first 24 hours. Growth cones were rapidly observed at the leading edge of extending neurites, with these growth cones enriched for F-actin and expressing essential guidance receptors. The effects of varying substrates and media compositions were tested on hPSC-derived RGCs, with optimal neurite outgrowth at a rate of approximately 50 µm/hr, or greater than 1 mm in the first 24 hours of growth.

Conclusions : The results of this study demonstrate the robust ability of hPSC-derived RGCs to extend axons over long distances. By extension, opportunities exist by which to control the directionality of axonal outgrowth as well. Overall, these results will facilitate the replacement of RGCs following their loss due to disease and degeneration, as extensive axonal outgrowth will be critical for the development of personalized transplant therapies for optic neuropathies.

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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