Structural and functional assessment after subretinal transplantation of human iPSC-derived photoreceptor precursors in a canine model of end stage retinal degeneration

Abstract

Purpose : Human photoreceptor precursor cells (hPRPCs) survive and integrate following subretinal injection (SRI) in degenerating retinas of dogs placed under systemic immunosuppression. Here we evaluated the value of the RHO^T4R/T4R dog, a light-inducible model of end stage retinal degeneration, to assess structural and vision restoration following SRI of iPSC-derived hPRPCs.

Methods : 12 days after a bilateral light damage that causes acute rod loss, two RHO^T4R/T4R dogs received in the left eye a 50 µL SRI of ~5 million hPRPCs derived from day 72 retinal organoids. Photoreceptor structure and function were assessed by color fundus photography, cSLO/OCT, ff and mfERG. Visual cortical and geniculate activities to monocular stimulation were evaluated by fMRI using a monocular, photopic 16 Hz flicker. Visual behavior was evaluated in an obstacle avoidance course under scotopic (0.003 Lux) to photopic (646 Lux) illumination, and in a forced 2-choice Y maze with bright white light stimuli.

Results : Following successful local SRI, the entire retinas detached as early as 48 hrs post-injection (PI). At 8 wks PI the retina of one dog remained detached with no evidence of retained PRPCs, while in the second dog retinal reattachment was near complete with retention of PRPCs covering ~ 19 mm² above the area centralis. There were no clinical signs of rejection up to termination at 45 wks PI. IHC showed loss of host RPE under a multilayered donor graft (>18 rows of iPRP cells) composed primarily of rod opsin- and cone arrestin-positive cells, along with rare infiltration of the graft with CD18-, CD3-, C20-positive cells. There was close contact between PKCa-positive canine rod bipolar cell dendrites and hPRPCs expressing presynaptic markers such as synaptophysin, VGLUT1, and CASK1. However, neither ffERG, mfERG, fMRI, nor obstacle course revealed improved function that could be attributed to the transplant. Intriguingly, the Y maze test showed impaired performance when testing the transplanted vs. the untreated eye.

Conclusions : These results confirm the long-term survival of hPRPCs in the immunosuppressed canine retina. However, damage to the RPE (described in this light damage model) may hamper retinal reattachment and prevent functional restoration. This model may thus be better suited to assess dual RPE:PRPC replacement instead of PRPCs alone.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.