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 en bloc-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.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 10824-10833 |
| Number of pages | 10 |
| Journal | Proceedings of the National Academy of Sciences of the United States of America |
| Volume | 166 |
| Issue number | 22 |
| DOIs | |
| State | Published - May 28 2019 |
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Keywords
- Cone and rod photoreceptor
- Retinal differentiation
- Retinal organoid
- RNA-seq
- Single cell
ASJC Scopus subject areas
- General
Cite this
Generation, transcriptome profiling, and functional validation of cone-rich human retinal organoids. / Kim, Sangbae; Lowe, Albert; Dharmat, Rachayata; Lee, Seunghoon; Owen, Leah A.; Wang, Jun; Shakoor, Akbar; Li, Yumei; Morgan, Denise J.; Hejazi, Andre A.; Cvekl, Ales; DeAngelis, Margaret M.; Jimmy Zhou, Z.; Chen, Rui; Liu, Wei.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 166, No. 22, 28.05.2019, p. 10824-10833.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Generation, transcriptome profiling, and functional validation of cone-rich human retinal organoids
AU - Kim, Sangbae
AU - Lowe, Albert
AU - Dharmat, Rachayata
AU - Lee, Seunghoon
AU - Owen, Leah A.
AU - Wang, Jun
AU - Shakoor, Akbar
AU - Li, Yumei
AU - Morgan, Denise J.
AU - Hejazi, Andre A.
AU - Cvekl, Ales
AU - DeAngelis, Margaret M.
AU - Jimmy Zhou, Z.
AU - Chen, Rui
AU - Liu, Wei
PY - 2019/5/28
Y1 - 2019/5/28
N2 - 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 en bloc-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.
AB - 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 en bloc-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.
KW - Cone and rod photoreceptor
KW - Retinal differentiation
KW - Retinal organoid
KW - RNA-seq
KW - Single cell
UR - http://www.scopus.com/inward/record.url?scp=85066287373&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85066287373&partnerID=8YFLogxK
U2 - 10.1073/pnas.1901572116
DO - 10.1073/pnas.1901572116
M3 - Article
C2 - 31072937
AN - SCOPUS:85066287373
VL - 166
SP - 10824
EP - 10833
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 22
ER -