Direct induction of retinal ganglion cell-like neurons
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA590512 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10497623_1.fastq.gz | Fastqsanger.gz | |||
| SRR10497623_2.fastq.gz | Fastqsanger.gz | |||
| SRR10497624_1.fastq.gz | Fastqsanger.gz | |||
| SRR10497624_2.fastq.gz | Fastqsanger.gz | |||
| SRR10497625_1.fastq.gz | Fastqsanger.gz |
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Project description:Direct induction of retinal ganglion cell-like neurons
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Project description:Retinal ganglion cells (RGCs) are the projection neurons in the retina that connect the visual sensing tissue to the brain. We found that Ascl1/Brn3b/Isl1 transcription factor combination can quickly and efficiently reprogramming mouse embryonic fibroblasts (MEFs) into retinal ganglion cell-like neurons (iRGCs). Using RNA-seq, we analyzed the transcriptomes of MEFs infected with Ascl1/Brn3b/Isl1-overexpressing viruses on day 2 or day 7 of reprogramming, or the final iRGCs on day 13 of reprogramming.
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Project description:Mouse embryonic fibroblasts were reprogrammed into retinal ganglion cell-like neurons by overexpressing Ascl1/Brn3b/Isl1 transcription factor combination in 13 days.
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Project description:During development of the central nervous system (CNS), cycling uncommitted progenitor cells give rise to a variety of distinct neuronal and glial cell types. As these different cell types are born, they progress from newly specified cells to fully differentiated neurons and glia. In order to define the developmental processes of individual cell types, single cell expression profiling was carried out on developing ganglion and amacrine cells of the murine retina. Individual cells from multiple developmental stages were isolated and profiled on Affymetrix oligonucleotide arrays. These experiments have yielded an expanded view of the processes underway in developing retinal ganglion and amacrine cells, as well as several hundred new marker genes for these cell types. In addition, this study has allowed for the definition of some of the molecular heterogeneity both between developing ganglion and amacrine cells and among subclasses of each cell type. Experiment Overall Design: Single retinal cells were isolated in tubes containing lysis buffer, their mRNAs were reverse transcribed, and the resulting cDNAs were PCR amplified for 35 cycles. Labeled cDNA samples were hybridized to Affymetrix 430 2.0 microarrays and the data was normalized using MAS5.0 software. Cells were identified post hoc as either developing retinal ganglion cells, amacrine cells or rod photoreceptor cells.
2008-06-16 | E-GEOD-9812 | biostudies-arrayexpress
Project description:Since retinal ganglion cells (RGCs) do not regenerate after injury, RGC replacement therapy could provide an approach to vision restoration in glaucoma and other optic neuropathies. Here we developed a rapid protocol for direct induction of RGC (iRGC) differentiation from human iPSCs and ESCs in less than two weeks, by overexpression of NGN2 in RGC survival-promoting medium supplemented with a Notch inhibitor. Neuronal morphology and neurite growth were observed within one week of induction. Immunostaining and qRT-PCR for characteristic RGC-specific genes confirmed identity. Calcium imaging was used to evaluate iRGCs’ electrophysiologic maturation, and demonstrated GABA-induced excitatory calcium influx characteristic of immature RGCs. Using single cell-RNA sequencing (scRNA-seq) we further delineated iRGCs’ differentiation and compared iRGC transcriptomic profiles to fetal human and retinal organoid-derived RGCs. Unbiased clustering showed high similarities of transcriptomic profiles among iRGCs, early-stage fetal human RGCs and early-stage retinal organoid-derived RGCs. However, for some markers, including BRN3a, BRN3b and NEFL, iRGCs demonstrated expression patterns more similar to fetal RGCs than to retinal organoid RGCs. After intravitreal injection into rodent eyes, transplanted iRGCs survived and migrated into host retinas where they were detected one week and one month after transplant. Prior optic nerve trauma to the recipient host did not significantly enhance or detract from iRGC integration, but iRGCs protected host RGCs from neurodegeneration. Taken together, these data demonstrate rapid iRGC generation in vitro into an immature cell with high similarity to human fetal RGCs and capacity for retinal integration after transplant, including after optic nerve injury. The simplicity of this system may benefit translational studies on human RGCs.
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Project description:RNA-seq analysis from young and pre-glaucomatous DBA/2J retinal ganglion cells and control (age and sex-matched, D2-Gpnmb+) retinal ganglion cells
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Project description:During development of the central nervous system (CNS), cycling uncommitted progenitor cells give rise to a variety of distinct neuronal and glial cell types. As these different cell types are born, they progress from newly specified cells to fully differentiated neurons and glia. In order to define the developmental processes of individual cell types, single cell expression profiling was carried out on developing ganglion and amacrine cells of the murine retina. Individual cells from multiple developmental stages were isolated and profiled on Affymetrix oligonucleotide arrays. These experiments have yielded an expanded view of the processes underway in developing retinal ganglion and amacrine cells, as well as several hundred new marker genes for these cell types. In addition, this study has allowed for the definition of some of the molecular heterogeneity both between developing ganglion and amacrine cells and among subclasses of each cell type. Keywords: Single retinal cell gene expression profiling across multiple mouse developmental stages
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Project description:Epigenetic mark deposition during embryonic development contribute to postnatal homeostasis and tissue stability. Previously, we found out that Ezh2 contributes critically to the function and postnatal cell survival in bipolar cells but not in retinal ganglion cells in the retina. (Yan et al. Postnatal onset of retinal degeneration by loss of embryonic Ezh2 repression of Six1. Sci Report. doi:10.1038/srep33887; (Cheng L, Wong LJ, Yan N, Han RC et al. Ezh2 does not mediate retinal ganglion cell homeostasis or their susceptibility to injury. PLoS One 2018;13(2):e0191853.). In this study, we used RNA-seq to define up- and down regulated genes in both Ezh2 and G9a deficient (Math5Cre; Ezh2f/fG9af/+; dKO) retinal ganglion cells (RGC) to evaluate the hypothesis of Ezh2 and G9a interaction that has been discussed in other tissues but in the retina. ChIP-Seq was applied to evaluate H3K27me3 histone marks in retinal ganglion cells in wild type (WT), Ezh2 (Math5Cre; Ezh2-/-, sKO) and combined G9a-Ezh2 (Math5Cre; G9a+/-Ezh2-/-, dKO) knockout mutant mice at P1.
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Project description:Local mRNA translation mediates the adaptive responses of axons to extrinsic signals but direct evidence that it occurs in mammalian CNS axons in vivo is scant. We developed an axon-TRAP-RiboTag approach in mouse that allows deep-sequencing analysis of ribosome-bound mRNAs in the retinal ganglion cell axons of the developing and adult retinotectal projection in vivo. The embryonic-to-postnatal axonal translatome comprises an evolving subset of enriched genes with axon-specific roles suggesting distinct steps in axon wiring, such as elongation, pruning and synaptogenesis. Adult axons, remarkably, have a complex translatome with strong links to axon survival, neurotransmission and neurodegenerative disease. Translationally co-regulated mRNA subsets share common upstream regulators, and novel sequence elements generated by alternative splicing that promote axonal mRNA translation. Our results indicate that intricate regulation of compartment-specific mRNA translation in mammalian CNS axons supports the formation and maintenance of neural circuits in vivo. The profiling of ribosome-bound mRNAs in mouse retinal ganglion cell axons at 4 different developmental stages
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