Fate and Rescue Effect of Human Forebrain Progenitor Cells Transplanted Into the Subretinal Space of a Rat Model of Photoreceptor Degeneration

Abstract

Purpose: : To determine the fate of donor human forebrain progenitor cells (hFPCs) injected into the subretinal space of a rat model of photoreceptor degeneration and assess their ability to rescue photoreceptors and vision.

Methods: : Prenatal hFPCs isolated from cortical tissue of a 94–day donor were grown as neurospheres in media supplemented with EGF and LIF. After >16 passages, neurospheres were dissociated into single cells and examined for cell viability using trypan blue dye exclusion prior to transplantation. 2 x 10⁴ cells were injected into the subretinal space of 21 day–old RCS rats (n>20) that were maintained on oral cyclosporine. Functional efficacy was tested by electroretinogram (ERG), threshold optomotor acuity and luminance thresholds recorded from the superior colliculus. All treated eyes were compared with sham–injected and untreated eyes. Eyes were then fixed and sectioned for staining with cresyl violet for overall retinal organization, human nuclear marker for donor cell localization and a range of cell specific antibodies.

Results: : ERG responses were significantly (p<0.01) better than shams at P90 (a–wave: 101.5±55.6 vs 5.9±9.6 µV; cone b–wave: 171.5±76.0 vs 23.0±14.7 µV). Optomotor thresholds measured at 100 days were significantly (p<0.001) improved over shams (0.50±0.06 c/d vs 0.28±0.03 c/d) with best results giving near–normal figures (0.57±0.01 c/d). Superior colliculus recordings at P100 also showed much lower luminance threshold responses in hFPC–treated eyes (average: 1.67±0.15 log units vs 3.13±0.15 log units), with some individual recordings within the normal range (0.3 log units). Histological studies in our best–performing transplant recipients revealed substantial photoreceptor rescue with a nearly fully preserved outer nuclear layer when compared to non–dystrophic animals. Donor cells formed a semi–continuous, pigmented cell layer immediately internal to host RPE and were also found scattered within the inner retina, where they remained nestin–positive and negative for retina–specific markers.

Conclusions: : Human FPCs are highly migratory, appear capable of adopting at least some characteristics of RPE cells and can achieve robust rescue of photoreceptors and visual function in this animal model of retinal degeneration.

Keywords: transplantation • retinal degenerations: hereditary • photoreceptors: visual performance