Filters
Technology
Platform
SRP117613
Homo sapiens
6 Downloadable Samples
Illumina HiSeq 2500
Description
We discuss the use of pluripotent stem cell lines carrying fluorescent reporters driven by retinal promoters to derive three-dimensional (3-D) retina in culture and how this system can be exploited for elucidating human retinal biology, creating disease models in a dish, and designing targeted drug screens for retinal and macular degeneration. Furthermore, we realize that stem cell investigations are labor-intensive and require extensive resources. To expedite scientific discovery by sharing of resources and to avoid duplication of efforts, we propose the formation of a Retinal Stem Cell Consortium. In the field of vision, such collaborative approaches have been enormously successful in elucidating genetic susceptibility associated with age-related macular degeneration. Overall design: CRX+ flow sorted cells from human retina derived organoids were collected at 6 time points during differentiation (day (D) 37, 48, 67, 90, 134, 220).
Publication Title
Treatment Paradigms for Retinal and Macular Diseases Using 3-D Retina Cultures Derived From Human Reporter Pluripotent Stem Cell Lines.
Sample Metadata Fields
Specimen part, Subject
View Samples- Danio rerio
- 9 Downloadable Samples
- IlluminaGenomeAnalyzerIIx
Description
Purpose: Zebrafish neurons regenerate from Müller glia following retinal lesions. Genes and signaling pathways important for retinal regeneration in zebrafish have been described, but our understanding of how Mu¨ller glial stem cell properties are regulated is incomplete. Mammalian Mu¨ller glia possess a latent neurogenic capacity that might be enhanced in regenerative therapies to treat degenerative retinal diseases. Methods: To identify transcriptional changes associated with stem cell properties in zebrafish Mu¨ller glia, we performed a comparative transcriptome analysis from isolated cells at 8 and 16 hours following an acute, photic lesion, prior to the asymmetric division that produces retinal progenitors. Results: We report a rapid, dynamic response of zebrafish Müller glia, characterized by activation of pathways related to stress, NF-kappa B signaling, cytokine signaling, immunity, prostaglandin metabolism, circadian rhythm, and pluripotency, and an initial repression of Wnt signaling. When we compared publicly available transcriptomes of isolated mouse Mu¨ller glia from two retinal degeneration models, we found that mouse Müller glia showed evidence of oxidative stress, variable responses associated with immune regulation, and repression of pathways associated with pluripotency, development, and proliferation. Conclusions: Categories of biological processes/pathways activated following photoreceptor loss in regeneration-competent zebrafish Mu¨ller glia, which distinguished them from mouse Mu¨ller glia in retinal degeneration models, included: cytokine signaling (notably NF-kappa B), prostaglandin E2 synthesis, expression of core clock genes, and pathways/metabolic states associated with pluripotency. These regulatory mechanisms are relatively unexplored as potential mediators of stem cell properties likely to be important in Müller glial cells for successful retinal regeneration. Overall design: Transcriptional profiles of 0, 8, and 16 hour post-lesion zebrafish Müller glia (in triplicate) were generated by high-throughput sequencing in an Illumina GAIIx.
Publication Title
Rapid, Dynamic Activation of Müller Glial Stem Cell Responses in Zebrafish.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 6 Downloadable Samples
Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Our data suggest that CNTF remodels the transcription profile of Mller (glial) cells leading to induction of networks associated with transcription, cell cycle regulation and inflammatory response. CNTF also appears to function as an inducer of gliosis in the retina. These studies provide new insights into the biological functions of cytokines in the retina.
Publication Title
Ciliary neurotrophic factor induces genes associated with inflammation and gliosis in the retina: a gene profiling study of flow-sorted, Müller cells.
Sample Metadata Fields
Specimen part, Treatment, Time
View Samples- Mus musculus
- 8 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Rod and cone photoreceptors in mammalian retina are generated from common pool(s) of neuroepithelial progenitors. NRL, CRX and NR2E3 are key transcriptional regulators that control photoreceptor differentiation. Mutations in NR2E3, a rod-specific orphan nuclear receptor, lead to loss of rods, increased density of S-cones, and supernormal S-cone-mediated vision in humans. To better understand its in vivo function, NR2E3 was expressed ectopically in the Nrl-/- retina, where post-mitotic precursors fated to be rods develop into functional S-cones similar to the human NR2E3 disease. Expression of NR2E3 in the Nrl-/- retina completely suppressed cone differentiation and resulted in morphologically rod-like photoreceptors, which were not functional. Gene profiling of FACS-purified photoreceptors confirmed the role of NR2E3 as a strong suppressor of cone genes and an activator of a subset of rod genes (including rhodopsin) in vivo. Ectopic expression of NR2E3 in cone precursors and differentiating S-cones of wild type retina also generates rod-like cells. The dual regulatory function of NR2E3 is not dependent upon the presence of NRL and/or CRX, but on the timing and level of its expression. Our studies reveal a critical role of NR2E3 in establishing functional specificity of post-mitotic photoreceptor precursors during retinal neurogenesis.
Publication Title
In vivo function of the orphan nuclear receptor NR2E3 in establishing photoreceptor identity during mammalian retinal development.
Sample Metadata Fields
Sex, Specimen part
View Samples- Homo sapiens
- 500 Downloadable Samples
Description
Age-related macular degeneration (AMD) is a complex multifactorial disease with at least 34 loci contributing to genetic susceptibility. To gain functional understanding of AMD genetics, we generated transcriptional profiles of retina from 453 individuals including both controls and cases at distinct stages of AMD. We integrated retinal transcriptomes, covering 13,662 protein-coding and 1,462 noncoding genes, with genotypes at over 9 million common single nucleotide polymorphisms (SNPs) for expression quantitative trait loci (eQTL) analysis of a tissue not included in Genotype-Tissue Expression (GTEx) and other large datasets. Cis-eQTL analysis revealed 10,474 genes under genetic regulation, including 4,541 eQTLs detected only in the retina. We then integrated the AMD-genome-wide association studies (GWAS) data with eQTLs and ascertained target genes at six loci. Furthermore, using transcriptome wide association analysis (TWAS), we identified 23 additional AMD-associated genes, including RLBP1, HIC1 and PARP12. Our studies expand the genetic landscape of AMD leading to direct targets for biological evaluation and establish the Genotype-Retina Expression (GREx) database as a resource for post-GWAS interpretation of retina-associated traits including glaucoma and diabetic retinopathy. Overall design: Retinal samples from 523 aged post-mortem human subjects from a spectrum of age-related macular degeneration (AMD) were RNA-seq profiled.
Publication Title
Improved Retinal Organoid Differentiation by Modulating Signaling Pathways Revealed by Comparative Transcriptome Analyses with Development In Vivo.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 30 Downloadable Samples
- Illumina Genome Analyzer IIx
Description
Vertebrate ancestors had only cone-like photoreceptors. The duplex retina evolved in jawless vertebrates with the advent of highly photosensitive rod-like photoreceptors. Despite cones being the arbiters of high-resolution color vision, rods emerged as the dominant photoreceptor in mammals during a nocturnal phase early in their evolution. We investigated the evolutionary and developmental origins of rods in two divergent vertebrate retinae. In mice, we discovered genetic and epigenetic vestiges of short wavelength cones in developing rods and cell lineage tracing validated the genesis of rods from S-cones. Curiously, rods did not derive from S-cones in zebrafish. Our study illuminates several questions regarding the evolution of duplex retina and supports the hypothesis that, in mammals, the S-cone lineage was recruited via the Maf-family transcription factor NRL to augment rod photoreceptors. We propose that this developmental mechanism allowed the adaptive exploitation of scotopic niches during the nocturnal bottleneck early in mammalian evolution. Overall design: GFP positive cells from Nrlp-GFP or Nrlp-GFP;Nrl-KO mouse retinas at post-natal ages P2, P4, P6, P10, P14, and P28 were isolated by flow sorting by FACSAria II (Becton Dickinson). Total RNA was extracted by Trizol LS (Life Technologies) and analyzed by 2100 Bioanalyzer (Agilent Technologies Genomics). High quality of total RNA (RIN: >7.0) was subjected to sequencing library construction using 20 ng of total RNA as input. Libraries were constructed using a stranded modification of the Illumina TruSeq mRNA (Brooks, et al. Meth Mol Biol 2012). Each library was single-end sequenced in an independent lane of a GAIIx at a length of 76 bases. Fastq files were generated from reads passing chastity filter.
Publication Title
Recruitment of Rod Photoreceptors from Short-Wavelength-Sensitive Cones during the Evolution of Nocturnal Vision in Mammals.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Mus musculus
- 27 Downloadable Samples
- Affymetrix Murine Genome U74A Version 2 Array (mgu74av2)
Description
Background: Progressive neurological dysfunction is a key aspect of human aging. Because of underlying differences in the aging of mice and humans, useful mouse models have been difficult to obtain and study. We have used gene-expression analysis and polymorphism screening to study molecular senescence of the retina and hippocampus in two rare inbred mouse models of accelerated neurological senescence (SAMP8 and SAMP10) that closely mimic human neurological aging, and in a related normal strain (SAMR1) and an unrelated normal strain (C57BL/6J).
Publication Title
Mechanisms of aging in senescence-accelerated mice.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 23 Downloadable Samples
- IlluminaGenomeAnalyzerIIx
Description
To define molecular mechanisms underlying rod and cone differentiation, we generated H9 human embryonic stem cell line carrying a GFP reporter that is controlled by the promoter of cone-rod homeobox (CRX) gene, the first known marker of post-mitotic photoreceptor precursors. CRXp-GFP reporter in H9 line replicates endogenous CRX expression when induced to form self-organizing 3-D retina-like tissue. We define temporal transcriptome dynamics of developing photoreceptors during the establishment of cone and rod cell fate. Our studies provide an essential framework for delineating molecules and cellular pathways that guide human photoreceptor development and should assist in chemical screening and cell-based therapies of retinal degeneration. Overall design: Undifferentiated CRXp-GFP HP hES cells and 3D-neural retina were collected at days 37, 47, 67 and 90 and dissociated into single cells. Cells were sorted at 4°C and by FACSAria (Becton Dickinson). GFP+ and GFP- cells were separately collected. Total RNA was extracted by RNA purification kit (Norgen Biotek) and analyzed by 2100 Bioanalyzer (Agilent Technologies Genomics). High quality of total RNA (RIN: 7.7-9.2) was subjected to libraries construction using 40-60 ng of total RNA as input. Libraries were constructed using a stranded modification of the Illumina TruSeq mRNA (Brooks, et al. Meth Mol Biol 2012). Each library was single-end sequenced in an independent lane of a GAIIx at a length of 76 bases. Fastq files were generated from reads passing chastity filter.
Publication Title
Transcriptome Dynamics of Developing Photoreceptors in Three-Dimensional Retina Cultures Recapitulates Temporal Sequence of Human Cone and Rod Differentiation Revealing Cell Surface Markers and Gene Networks.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 20 Downloadable Samples
- Illumina Genome Analyzer IIx
Description
Leber congenital amaurosis (LCA) includes congenital or early-onset blinding diseases, characterized by vision loss together with nystagmus and nonrecordable electroretinogram (ERG). At least 19 genes are associated with LCA. While most LCA is recessive, mutations in the homeodomain transcription factor gene CRX lead to autosomal dominant LCA. The mechanism of CRX-LCA is not understood. Here, we report a new spontaneous mouse mutant carrying a frameshift mutation in Crx (CrxRip). We show that, unlike Crx-/- mouse retina, the dominant Crx c.763del1 mutation in CrxRip results in congenital blindness with complete loss of ERG, yet the photoreceptors do not degenerate. Dominant CRX frameshift mutations associated with LCA mimic the CrxRip phenotype that can be rescued by Crx. RNA-Seq profiling reveals progressive and complete loss of rod differentiation factor Nrl in CrxRip, while residual Nrl remains in Crx-/- retina. Moreover, Nrl partially restores the rod phenotype in CrxRip/+ mice. We show that the binding of Otx2 to Nrl promoter is obliterated in CrxRip mutant, and ectopic Otx2 can rescue the rod phenotype. Therefore, Otx2 is required to maintain Nrl expression in developing rods to consolidate rod fate. Our studies provide the mechanism of congenital blindness caused by dominant CRX mutations and should assist in therapeutic design. Overall design: Retinal samples were harvested from WT, CrxRip/+, CrxRip/Rip, Crx-/- and Nrl-/- retina at postnatal days 2 and 21 for whole transcriptome sequencing (RNAseq). Each sample included 2 independent frozen retina and experiments were performed in duplicates. RNA-seq transcriptome libraries were constructed from 1 µg of total RNA.
Publication Title
OTX2 loss causes rod differentiation defect in CRX-associated congenital blindness.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 24 Downloadable Samples
- Affymetrix Murine Genome U74A Version 2 Array (mgu74av2)
Description
The rod photoreceptor-specific neural retina leucine zipper protein Nrl is essential for rod differentiation and plays a critical role in regulating gene expression. In the mouse retina, rods account for 97% of the photoreceptors; however, in the absence of Nrl (Nrl-/-), no rods are present and a concomitant increase in cones is observed.
Publication Title
Expression profiling of the developing and mature Nrl-/- mouse retina: identification of retinal disease candidates and transcriptional regulatory targets of Nrl.
Sample Metadata Fields
Specimen part
View Samples