Abstract
Purpose :
Leber congenital amaurosis (LCA) is a group of early-onset retinal diseases, characterized by dysfunction and/or death of photoreceptors. Almost 25% of LCA is caused by mutations in the centrosomal/ciliary protein of 290 kDa (CEP290), which is critical for the biogenesis of primary cilia. The outer segment of a photoreceptor represents a modified primary cilium and is structurally and functionally abnormal in CEP290-LCA. This study aims to identify disease-associated molecular features of patient induced pluripotent stem cell (iPSC)-derived retinal organoids, which can be used as a promising in vitro platform to model disease pathogenesis and develop treatments.
Methods :
We established iPSC lines from fibroblasts of CEP290-LCA patients harboring the most common mutation and the respective familial controls. These iPSC lines were differentiated into retinal organoids, which were harvested at different developmental time points for immunohistochemistry, transmission electron microscopy and transcriptome analysis.
Results :
iPSC lines could be differentiated into retinal organoids, with major cell types self-patterning into a polarized, laminated architecture mimicking human retina morphology. The ciliary axoneme of photoreceptors started to elongate in organoids derived from control iPSCs as early as differentiation day (D) 120. By D180, control photoreceptors in culture exhibited extensive outer segment-like structures, with rudimentary disc-like structures and transport of Rhodopsin to the outer segment. However, CEP290-LCA organoids demonstrated aberrant ciliogenesis from D120, with compromised photoreceptor cilia development and opsin mis-localization. We have now performed transcriptome analysis of differentiating control and patient organoids to define molecular mechanisms and identify therapeutic targets.
Conclusions :
CEP290-LCA patient iPSC-derived retinal organoids displayed developmental defects mimicking disease pathology. Therefore, retinal organoids can serve as a valuable in vitro platform to model blinding ciliopathies and evaluation of therapeutic candidates.
This is a 2020 ARVO Annual Meeting abstract.