Abstract
To determine the safety, survival, and functionality of human embryonic stem cell-derived RPE (hESC-RPE) cells seeded on a polymeric substrate (rCPCB-RPE1 implant) and implanted into the subretinal (SR) space of Royal College of Surgeons (RCS) rats. Monolayers of hESC-RPE cells cultured on parylene membrane were transplanted into the SR space of 4-week-old RCS rats. Group 1 (n = 46) received vitronectin-coated parylene membrane without cells (rMSPM+VN), group 2 (n = 59) received rCPCB-RPE1 implants, and group 3 (n = 13) served as the control group. Animals that are selected based on optical coherence tomography screening were subjected to visual function assays using optokinetic (OKN) testing and superior colliculus (SC) electrophysiology. At approximately 25 weeks of age (21 weeks after surgery), the eyes were examined histologically for cell survival, phagocytosis, and local toxicity. Eighty-seven percent of the rCPCB-RPE1-implanted animals showed hESC-RPE survivability. Significant numbers of outer nuclear layer cells were rescued in both group 1 (rMSPM+VN) and group 2 (rCPCB-RPE1) animals. A significantly higher ratio of rod photoreceptor cells to cone photoreceptor cells was found in the rCPCB-RPE1-implanted group. Animals with rCPCB-RPE1 implant showed hESC-RPE cells containing rhodopsin-positive particles in immunohistochemistry, suggesting phagocytic function. Superior colliculus mapping data demonstrated that a significantly higher number of SC sites responded to light stimulus at a lower luminance threshold level in the rCPCB-RPE1-implanted group. Optokinetic data suggested both implantation groups showed improved visual acuity. These results demonstrate the safety, survival, and functionality of the hESC-RPE monolayer transplantation in an RPE dysfunction rat model.
Highlights
Title Survival and Functionality of hESC-Derived Retinal Pigment Epithelium Cells Cultured as a Monolayer on Polymer Substrates Transplanted in Royal College of Surgeons (RCS) Rats
Significant numbers of outer nuclear layer cells were rescued in both group 1 and group 2 animals
A significantly higher ratio of rod photoreceptor cells to cone photoreceptor cells was found in the rCPCB-RPE1–implanted group
Summary
Monolayers of hESC-RPE cells cultured on parylene membrane were transplanted into the SR space of 4-week-old RCS rats. Group 1 (n 1⁄4 46) received vitronectin-coated parylene membrane without cells (rMSPMþVN), group 2 (n 1⁄4 59) received rCPCB-RPE1 implants, and group 3 (n 1⁄4 13) served as the control group. Royal College of Surgeons rat pups (27–29 days old) were divided into an rMSPMþVN (parylene substrate coated with vitronectin)–implanted group (group 1, n 1⁄4 46), an rCPCBRPE1–implanted group (group 2, n 1⁄4 59), and a nonimplanted control group (group 3, n 1⁄4 13). The isolated RPE-like cells were dissociated by trypLE (Life Technologies, Grand Island, NY, USA) and cultured in human VN (BD Biosciences, Franklin Lakes, NJ, USA)-coated plates with XVIVO 10 medium. The parylene substrates used for both rMSPMþVN and rCPCB-RPE1 implants consisted of an ultrathin membrane of parylene (0.30-lm thick) with a 6lm-thick supporting mesh. For group 1 studies, rMSPMs were coated with 10 lg/mL VN for 2 hours at room
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