Generation, transcriptome profiling, and functional validation of cone-rich human retinal organoids.

TitleGeneration, transcriptome profiling, and functional validation of cone-rich human retinal organoids.
Publication TypeJournal Article
Year of Publication2019
AuthorsKim, S, Lowe, A, Dharmat, R, Lee, S, Owen, LA, Wang, J, Shakoor, A, Li, Y, Morgan, DJ, Hejazi, AA, Cvekl, A, DeAngelis, MM, Z Zhou, J, Chen, R, Liu, W
JournalProc Natl Acad Sci U S A
Volume116
Issue22
Pagination10824-10833
Date Published2019 May 28
ISSN1091-6490
Abstract

Rod and cone photoreceptors are light-sensing cells in the human retina. Rods are dominant in the peripheral retina, whereas cones are enriched in the macula, which is responsible for central vision and visual acuity. Macular degenerations affect vision the most and are currently incurable. Here we report the generation, transcriptome profiling, and functional validation of cone-rich human retinal organoids differentiated from hESCs using an improved retinal differentiation system. Induced by extracellular matrix, aggregates of hESCs formed single-lumen cysts composed of epithelial cells with anterior neuroectodermal/ectodermal fates, including retinal cell fate. Then, the cysts were -passaged, attached to culture surface, and grew, forming colonies in which retinal progenitor cell patches were found. Following gentle cell detachment, retinal progenitor cells self-assembled into retinal epithelium-retinal organoid-that differentiated into stratified cone-rich retinal tissue in agitated cultures. Electron microscopy revealed differentiating outer segments of photoreceptor cells. Bulk RNA-sequencing profiling of time-course retinal organoids demonstrated that retinal differentiation in vitro recapitulated in vivo retinogenesis in temporal expression of cell differentiation markers and retinal disease genes, as well as in mRNA alternative splicing. Single-cell RNA-sequencing profiling of 8-mo retinal organoids identified cone and rod cell clusters and confirmed the cone enrichment initially revealed by quantitative microscopy. Notably, cones from retinal organoids and human macula had similar single-cell transcriptomes, and so did rods. Cones in retinal organoids exhibited electrophysiological functions. Collectively, we have established cone-rich retinal organoids and a reference of transcriptomes that are valuable resources for retinal studies.

DOI10.1073/pnas.1901572116
Alternate JournalProc. Natl. Acad. Sci. U.S.A.
PubMed ID31072937
PubMed Central IDPMC6561190
Grant ListP30 EY026878 / EY / NEI NIH HHS / United States
S10 OD018033 / OD / NIH HHS / United States
R01 EY012200 / EY / NEI NIH HHS / United States
R01 EY017353 / EY / NEI NIH HHS / United States
S10 OD023469 / OD / NIH HHS / United States
R01 EY026065 / EY / NEI NIH HHS / United States
T32 GM007491 / GM / NIGMS NIH HHS / United States
R01 EY022356 / EY / NEI NIH HHS / United States
P30 EY002520 / EY / NEI NIH HHS / United States
R01 EY018571 / EY / NEI NIH HHS / United States
R01 EY014237 / EY / NEI NIH HHS / United States
P30 CA013330 / CA / NCI NIH HHS / United States
P30 EY014800 / EY / NEI NIH HHS / United States
R01 EY022645 / EY / NEI NIH HHS / United States