Abstract
Purpose :
Although advances in stem cell biology will one day enable the development of cell replacement therapies to treat neurodegenerative diseases like glaucoma, technologies that promote cell survival and direct retinal ganglion cell (RGC) integration, axon growth, and synaptic connectivity are still needed. We and others have shown that electric field (EF) stimulation can direct migration of rodent neural progenitor cells as well as axon growth of differentiated rodent neurons. Given this, we hypothesize that EF simulation can be developed into a technology to direct integration of transplanted hESC-derived RGCs
Methods :
To characterize the electrotaxic response of hESC-derived RGCs to EF stimulation, an electrotaxis chamber was built onto a 100 mm tissue culture plate previously coated with Poly-L-Laminin. Purified RGCs from BRN3B-tdTomato hESCs-derived human retinal organoids were seeded onto the plate and agarose salt bridges were used to deliver current to the electrotaxis chamber. RGCs were continuously stimulated with either direct current (DC), alternative current (AC), or asymmetric charge-balanced (ACB) waveforms with various parameters (amplitude, pulse width, frequency, and duration). Cell survival and direction and velocity of axon growth were assessed via time-lapsed microscopy.
Results :
The application of an DC EF effectively guided axon growth of hESC-derived RGCs towards the cathode (-), without inflicting any tissue damage (untreated 37% +/- 0.8%; 200mV/mm 65% +/-13% of total RGC axons; n=3; p < 0.02 t-test). Based on this finding, our lab proceeded to develop an innovative ex-vivo stimulation device that generates a vertical voltage gradient across the retina that we will use to guide the integration and axon growth of RGCs into flat-mounted retinal explants.
Conclusions :
EF stimulation can guide and direct the axon growth of RGCs derived from hESCs, indicating that EF stimulation may be a viable technology to supplement other approaches to direct RGC integration for cell replacement therapies.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.