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Comparative transcriptomic analysis of self-organized, in vitro generated optic tissues

ABSTRACT: Embryonic stem (ES) cells have a remarkable capacity to self-organize complex, multi-layered optic cups in vitro via a culture technique called SFEBq. During both SFEBq and in vivo optic cup development, Rax (Rx) expressing neural retina epithelial (NRE) tissues utilize Fgf and Wnt/β-catenin signalling pathways to differentiate into neural retina (NR) and retinal-pigmented epithelial (RPE) tissues, respectively. How these signaling pathways affect gene expression during optic tissue formation has remained largely unknown, especially at the transcriptome scale.

ORGANISM(S): Mus musculus

PROVIDER: GSE62432 | GEO | 2015/05/18

SECONDARY ACCESSION(S): PRJNA264087

REPOSITORIES: GEO

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Comparative transcriptomic analysis of self-organized, in vitro generated optic tissues

Project description:Embryonic stem (ES) cells have a remarkable capacity to self-organize complex, multi-layered optic cups in vitro via a culture technique called SFEBq. During both SFEBq and in vivo optic cup development, Rax (Rx) expressing neural retina epithelial (NRE) tissues utilize Fgf and Wnt/β-catenin signalling pathways to differentiate into neural retina (NR) and retinal-pigmented epithelial (RPE) tissues, respectively. How these signaling pathways affect gene expression during optic tissue formation has remained largely unknown, especially at the transcriptome scale. We generated Day 10 Rx+ optic tissue using SFEBq, exposed these tissues to either Fgf or Wnt/β-catenin stimulation, and assayed their gene expression at Days 12 and 15 using RNA-Seq. We measured gene expression in these 5 sample groups in biological triplicate using RNA-seq (Illumina HiSeq) .

2015-05-18 | E-GEOD-62432 | biostudies-arrayexpress

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Genome-wide analysis of Sox2-ablated optic cups compared with wild-type control optic cups at E16.5

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Project description:Generation of three-dimensional (3D) organoids with optic cup like structures from pluripotent stem cells has created opportunities for investigating mammalian retinal development in vitro. However, retinal organoids in culture do not completely reflect the developmental state and in vivo architecture of the rod-dominant mouse retina. To assess rod photoreceptor differentiation in retinal organoids, we took advantage of Nrl-GFP mice that show rod-specific expression of GFP directed by the promoter of leucine zipper transcription factor NRL. Using embryonic and induced pluripotent stem cells (ESCs and iPSCs, respectively) derived from the Nrl-GFP mouse, we were successful in establishing long-term retinal organoid cultures (up to day 35) using modified culture conditions (called High Efficiency Hypoxia Induced Generation of Photoreceptors in Retinal Organoids, or HIPRO). We demonstrate efficient differentiation of pluripotent stem cells to retinal structures, with over 70% of embryoid bodies forming optic vesicles at Day (D)7, >50% producing optic cups by D10, and a majority of these surviving until at least D35. The HIPRO organoids include distinct inner retina neurons in somewhat stratified architecture and mature Müller glia spanning the entire retina. Almost 70% of the cells in retinal organoids are rod photoreceptors that exhibit elongated cilia. Transcriptome profiles of GFP+ rod photoreceptors, purified from organoids at Day 25-35, demonstrate a high correlation to gene profiles of purified rods from mouse retina at postnatal day (P) 2 to 6, indicating their early state of differentiation. Our 3D retinal organoids thus closely mimic in vivo retinogenesis and provide an efficient in vitro model to investigate photoreceptor development and modeling disease pathology.

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