Survival of Transplanted Retinal Ganglion Cell in Human Donor Eyes under Elevated Pressure

Glaucoma is a group of optic neuropathies characterized by visual field loss, classically due to increased intraocular pressure (IOP) and retinal ganglion cell (RGC) degeneration. Current treatment options reduce IOP, but progressive RGC degeneration persists. The ability to reprogram de novo RGCs from human corneal keratocytes provides a valuable tool to potentially restore vision in patients with late-stage disease when most RGCs are irreversibly damaged. We investigate the survival of these human induced pluripotent stem cell (hiPSC) derived RGCs after culturing them in human donor eyes under conditions of elevated and normal IOP using the pressurized ocular translaminar autonomous system (TAS) chamber. The hiPSCs were generated by reprogramming human donor keratocytes using Sendai viral vectors with Yamanaka factors. The hiPSCs were then differentiated into retinal organoids (ROs) from which RGCs were obtained. The RGCs were transduced with AAV2-CBA-EGFP (Adeno-Associated Virus serotype 2- Chicken Beta Actin- Enhanced Green Fluorescent Protein) and successfully transplanted into donor human eyes obtained from individuals having non-ocular history. They were pressurized for 5-7 days, with the left eye maintained at normal IOP and right eye at high IOP. Viability was measured by expression levels of pro-survival pathways via qRT-PCR, immunohistochemistry staining, and electroretinography (ERG) for retinal function. After RGC transplantation, increased expression of pro-survival and decreased inflammatory and apoptotic markers were identified in normal IOP conditions compared to high IOP. In conclusion, survival of RGCs was more conducive under normal IOP conditions with significantly increased degeneration observed at high IOP. Thus, suggesting that elevated IOP could potentially create a microenvironment that would significantly inhibit successful transplantation of de novo RGCs.