Abstract
Purpose :
Organic semiconductor polymers are encouraging materials for retinal prostheses, due to their key prosthetic features such as flexibility, light sensitivity and biocompatibility. A hindering problem remains to find the appropriate human model for pre-clinical research. Through the differentiation of human induced pluripotent stem cells (hiPSCs) to self-organized three dimensional (3D) retinal organoids (ROs), it is now possible to model the spatio-temporal development of the human retina and recapitulate in vitro the human (patho)physiology. In this context, we hypothesize that ROs can be used as human in vitro model to test the efficiency of novel organic retinal prosthesis.
Methods :
Two hiPSCs lines, one carrying a disease causing mutation on the PRPF31 gene and a healthy control; have been differentiated into ROs following a 2D/3D approach, treated with BMP4 from d6 to d15. On d30 the 3D structures with optic vesicle and lamination-like appearance were picked off and placed in suspension with retinoic acid until d100. ROs were collected at various time points and characterized through immunofluorescence, qPCR and flow cytometry for pluripotency, progenitor and photoreceptor markers. RO function was assessed via microelectrode arrays. The organic semiconductor layers were processed from solution using the spin-coating and vapor deposition techniques.
Results :
We confirmed the pluripotency of the hiPSC lines for TRA-1-60, OCT4 and NANOG and the presence of the PRPF31 deletion. Late ROs showed a decrease in progenitor markers (KI67 and PAX6), an increase in early and late photoreceptor markers (OTX2, CRX, RCVRN, OPSIN), ganglion and amacrine cell markers (HuC and Brn3b), and bipolar cell marker (PKCa). Proper retinal layer organization was observed and functional electrical firing was recorded. ROs kept in direct contact with organic materials showed no cytotoxicity by apoptosis analysis.
Conclusions :
With the discovery of iPSCs and their differentiation into 3D organoids, a new horizon of human in vitro models arose. ROs bridge the gap between animal experimental models and 2D cultures. Our results so far indicate that ROs represent a proper human in vitro model as assessed by cellular, molecular and functional analysis. Further investigation is underway to confirm their suitability as human models for organic prosthesis.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.