Transplantation of Bio-Printed Choroid-RPE Tissue into The Subretinal Space of Rats

Abstract

Purpose : Age-related macular degeneration (AMD) is the principle cause of severe, progressive and irreversible blindness among the elderly. For late stage AMD, autologous choroid (Ch)-retinal pigment epithelium (RPE) translocation has been beneficial for some patients as a last resort. In this study, we aimed to transplant a bio-printed human iPSC-derived (hiPSC) choroid (to restore the vascular bed) and Ch-RPE tissue into the rat subretinal space.

Methods : A hydrogel mixture containing hiPSC-endothelial cells, fibroblasts and pericytes was bio-printed (Organovo, CA) on an electrospun scaffold (PDLGA 50:50, 1.0 I.V.; Biosurfaces, MA) and cultured for a week. hiPSC-RPE was seeded (non-bioprinted) on the other side of the scaffold; creating a 3D multilayered tissue. The tissue was implanted after 5-7 weeks of in vitro maturation. RNU (immunocompromised) rats were anaesthetized using a mixture of Ketamine and Xylazine according to their weight (10:1 w/w; 0.13 mL/100 g). Baseline ERG (Electroretinography; Diagnosys, MA) measurements (photopic and scotopic) were recorded. Rats underwent surgery where a disc (1 mm diameter) was placed under the retina. Rats were divided into different treatment groups (Ch-RPE; Ch; RPE and empty scaffold) receiving transplants in both eyes. The animals were followed using SLO-OCT (Scanning Laser Ophthalmoscopy-Optical Coherence Tomography; Heidelberg, DE), FA-ICGA (Fluorescein Angiography-Indocyanine Green Angiography; Heidelberg) and ERG for 2 months. ERG responses were analyzed using the a-/b-wave amplitudes and latencies (MATLAB). At various time points, eyes were collected for histology.

Results : We established a technique for placing a 1 mm disc of tissue into the subretinal space of rats. A total of 33 rats were implanted with tissues. SLO-OCT and FA-ICGA analyses showed survival of the transplant and signs of integration. According to histological analyses, transplants survived for at least 2 months in the subretinal space. ERG analyses showed reduced responses two weeks after surgery, but this was regardless of treatment.

Conclusions : We successfully placed a 1 mm disc of tissue under the retina of RNU rats. We determined the functional and structural changes following surgery, non-invasively. Transplantation of bio-printed 3D RPE-Ch tissue into the subretinal space of an animal model opens the possibility of developing complex tissue therapies for retinal degenerative diseases.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.