Subretinal transplantation of human induced pluripotent stem cell-derived retinal pigment epithelium (hiPS-RPE) into pigs.

Abstract

Purpose : To determine the safety, survival, and distribution of human induced pluripotent stem cell-derived RPE (hiPS-RPE) [protocol Tanaka, European Patent - EP2383333 B1; US Patent - US9249390 B2] implanted as a cell suspension into the subretinal (SR) space of domestic pigs (German Landrace).

Methods : In 10 pigs (age: 9 - 15 weeks) subretinal hiPS-RPE cell transplantation (3-port standard vitrectomy, subretinal injection with a 32 gauges cannula, RPE cell suspensions with different cell counts, 150.000 to 400.000 cells/100µl, right eye) was performed under general anesthesia. RPE cells were generated by a patented protocol described in Zhu et al. (2013). One to four weeks after transplantation, pigs were euthanatized. Experimental eyes were enucleated, SD-OCT imaged and paraformaldehyde-fixed. Next, serial cryo-sections were prepared for histological and immunhistochemical analysis.

Results : A protocol for the injection of iPSC-derived RPE cells to the subretinal space of domestic pigs was developed. Creating a sub-retinal bleb with 20-50µl PBS prior transplantation of the cell suspension improved donor cell delivery and circumvented reflux into the vitreous. Improved transplantation outcomes were further received by increasing the number of donor RPE cells in the suspension to 400.000 cells. Hyper-pigmented areas were detectable with SD-OCT within experimental retinas and immunhistochemical stainings using human- and RPE-specific markers revealed location of iPSC-derived RPE clusters within the subretinal space of the host. Results show a large variation of cell amount and grade of subretinal pigmentation after transplantation.

Conclusions : Sub-retinal transplantation of human iPSC-derived RPE into the pig eye was developed. Improved protocols allowed safe donor cell delivery as no side effects were observed in the treated eyes. However, the surgical process for subretinal cell injections might have to be further optimized to allow uniform distribution of cells and robust outcomes. This proof-of-concept study serves as a starting point to analyse in more detail the interactions between human RPE cells and endogenous pig photoreceptors following transplantation.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.