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Sumitra Sengupta, Jeff S Mumm; Identification of chemicals and signaling pathways that impact photoreceptor regeneration in zebrafish. Invest. Ophthalmol. Vis. Sci. 2017;58(8):350.
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© ARVO (1962-2015); The Authors (2016-present)
Irreversible loss of photoreceptors is the major cause of blindness in retinal degenerative disorders. Using an inducible zebrafish model of retinitis pigmentosa for whole-organism high-throughput drug discovery the goal of our study is to identify compounds, and corresponding signaling pathways, that promote photoreceptor regeneration. Our core hypothesis posits that a detailed understanding of the mechanisms regulating the regenerative potential of Müller glia in zebrafish will provide insights necessary to advance therapeutic strategies for stimulating retinal repair in humans.
Using an inducible degenerative disease modeling system and large-scale whole-organism drug discovery platform we have embarked upon a high-throughput screen (HTS) to identify compounds that accelerate neuronal regeneration kinetics in the zebrafish retina. Transgenic zebrafish expressing a yellow fluorescent protein-nitroreductase (YFP-NTR) fusion protein in rod photoreceptors enable prodrug-induced apoptosis of rod cells, a model of retinitis pigmentosa. To identify compounds promoting rod cell regeneration, fish larvae were treated with the prodrug metronidazole (Mtz, 10mM) for 24 hrs to induce rod cell ablation. After Mtz washout, fish were exposed to test compounds across six different concentrations over three days of recovery. YFP reporter levels were then quantified using a fluorescent plate reader to compare the extent of rod cell regeneration in drug-treated fish versus controls. Nonparametric multiple group comparisons were used to identify compounds that accelerated the kinetics of rod cell replacement; i.e., regeneration-promoting drugs.
In a pilot screen, we have identified 2 hit compounds that enhance rod cell regeneration. Suspected molecular targets of the hit compounds implicate neuroinflammation as a key regulator of the regenerative process. Further, by exposing larvae to hit compounds before or after induction of rod PR loss we have revealed stage-specific roles for immunomodulatory compounds during the regenerative process.
These results demonstrate that our whole-organism HTS-paced platform for drug discovery can successfully identify chemical modulators of retinal neuron regeneration, and reveal a novel role for the immune system in regulating the regenerative potential of Müller glia cells.
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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