Generation, transcriptome profiling, and functional validation of single cells from cone-enriched human retinal organoids
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE119343 | GEO | 2019/04/25
REPOSITORIES: GEO
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Project description:The macula of the retina has a high ratio of cones to rods and is critical for central vision and visual acuity. Macula degenerations affect vision the most and are incurable. Here we report the generation, transcriptome profiling, and functional validation of cone-enriched human retinal organoids differentiated from hESCs. Transcriptome profiling using bulk RNA-seq demonstrated that retinal differentiation in vitro recapitulated retinogenesis in vivo in the temporal expression of cell differentiation markers and retinal disease genes, as well as in mRNA alternative splicing. Single-cell RNA-seq of 8-month retinal organoids identified clusters of cone and rod photoreceptors and confirmed the cone enrichment initially revealed by immunostaining. Notably, comparisons of single-cell transcriptomes demonstrated the similarity between retinal organoids and human macula in cones and rods. Cones in retinal organoids exhibited electrophysiological functions. Collectively, we have established cone-enriched retinal organoids and a reference of transcriptomes that are rich resources for retinal studies.
2019-04-25 | GSE119274 | GEO
Project description:Cone photoreceptors are the primary initiator of visual transduction in the human retina. Dysfunction or death of rod photoreceptors precedes cone loss in many retinal and macular degenerative diseases, suggesting a rod-dependent trophic support for cone survival. Rod differentiation and homeostasis are dependent on the basic motif leucine zipper transcription factor NRL. The loss of Nrl in mice (Nrl-/-) results in a retina with predominantly S-opsin containing cones that exhibit molecular and functional characteristics of WT cones. Here we report that Nrl-/- retina undergoes a rapid but transient period of degeneration in early adulthood, with cone apoptosis, retinal detachment, alterations in retinal vessel structure, and activation and translocation of retinal microglia. However, cone degeneration stabilizes by four months of age, resulting in a thinned but intact outer nuclear layer with residual cones expressing S- and M-opsins and a preserved photopic ERG. At this stage, microglia translocate back to the inner retina and reacquire a quiescent morphology. Gene profiling analysis during the period of transient degeneration reveals misregulation of stress response and inflammation genes, implying their involvement in cone death. The Nrl-/- retina illustrates the long-term viability of cones in the absence of rods and may serve as a model for elucidating mechanisms of cone homeostasis and degeneration that would be relevant to understanding diseases of the cone-dominant human macula. Targets were generated from a pair of retinas (one Nrl-/- mouse) per biological replicate. Four biological replicates were generated for each of the five aging timepoints (1, 2, 4, 6, and 10 months post natal).
2011-11-15 | E-GEOD-33674 | biostudies-arrayexpress
Project description:Cone photoreceptors are the primary initiator of visual transduction in the human retina. Dysfunction or death of rod photoreceptors precedes cone loss in many retinal and macular degenerative diseases, suggesting a rod-dependent trophic support for cone survival. Rod differentiation and homeostasis are dependent on the basic motif leucine zipper transcription factor NRL. The loss of Nrl in mice (Nrl-/-) results in a retina with predominantly S-opsin containing cones that exhibit molecular and functional characteristics of WT cones. Here we report that Nrl-/- retina undergoes a rapid but transient period of degeneration in early adulthood, with cone apoptosis, retinal detachment, alterations in retinal vessel structure, and activation and translocation of retinal microglia. However, cone degeneration stabilizes by four months of age, resulting in a thinned but intact outer nuclear layer with residual cones expressing S- and M-opsins and a preserved photopic ERG. At this stage, microglia translocate back to the inner retina and reacquire a quiescent morphology. Gene profiling analysis during the period of transient degeneration reveals misregulation of stress response and inflammation genes, implying their involvement in cone death. The Nrl-/- retina illustrates the long-term viability of cones in the absence of rods and may serve as a model for elucidating mechanisms of cone homeostasis and degeneration that would be relevant to understanding diseases of the cone-dominant human macula.
2011-11-15 | GSE33674 | GEO
Project description:Cone photoreceptor cell death in inherited retinal diseases, such as Retinitis Pigmentosa (RP), leads to the loss of accurate and color vision and ultimately blindness. In RP, a vast number of mutations are affecting the structure and function of rod photoreceptors while cones remain mutation-free. Once majority of rods have degenerated cones are dying secondarily due to the increased oxidative stress, inflammation and loss of structural and nutritional support normally provided by rods. Here we demonstrated that secondary cone cell death in animal models for RP is governed by an increased activity of histone deacetylates (HDACs). A single intravitreal injection of an HDAC inhibitor at a late stage of the disease, when majority of rods have already degenerated, is sufficient to delay cone death and support long-term cone survival. Surviving cones are retaining functionality and are mediating light-driven ganglion cell responses. RNA-seq analysis of surviving cones demonstrated that HDAC inhibition affords multi-level protection trough regulation of different prosurvival pathways including MAPK, PI3K-AKT and autophagy. These study suggest a unique possibility for targeted pharmacological protection of both primary degenerating rods and mutation-free secondary dying cones and creates hope to maintain vision in RP patients independent of the disease stage.
2020-11-04 | GSE141601 | GEO
Project description:Both rod and cone photoreceptors are critical for mammalian vision yet they have important functional differences. In this study, we investigate patterns of DNA methylation and chromatin accessibility in mouse rods and cones. Unexpectedly, we find that a substantial fraction of hypo-methylated regions in the rod methylome are in inaccessible chromatin. These regions show marks of active regulatory regions earlier in neuronal development and could remain undermethylated in adult rods due to their highly compact chromatin. We further identify rod- and cone-enriched regions of accessible chromatin that correspond with cell type-specificity of nearby photoreceptor gene expression. We predict novel DNA signatures of rod and cone specificity and show that NR2E3 function is necessary for rods to gain their complete ensemble of epigenomic features. Taken together, our data capture the epigenomic landscapes of retinal rods and cones, including differences relevant to chromatin structure and photoreceptor biology.
2016-03-07 | GSE72550 | GEO
Project description:The mechanisms underlying specification of neuronal subtypes within the human nervous system are largely unknown. The blue/S, green/M and red/L cones of the retina enable high-acuity daytime and color vision. To determine the mechanism controlling S vs. L/M fates, we studied the differentiation of human retinal organoids. Organoids and retinas have similar distributions, expression profiles, and morphologies of cone subtypes. S cones are specified first, followed by L/M cones, and thyroid hormone signaling controls this temporal switch. Dynamic expression of thyroid hormone-degrading and activating proteins within the retina ensures low signaling early to specify S cones and high signaling late to produce L/M cones. This work establishes organoids as a model for determining mechanisms of human development with promising utility for therapeutics and vision repair.
2018-10-12 | GSE119320 | GEO
Project description:The retinas of simian primates include a specialized, cone-rich, macula which regards the central visual field and mediates high acuity and colour vision. A prominent feature of the macula is the fovea centralis - a 1 mm-wide, avascular depression in the inner retinal surface that corresponds with a local absence of rods and a peak spatial density of cones in the outer photoreceptor layer. The arrangement of macular photoreceptors, and their specialized ‘midget’ circuits, are the neural substrate for high resolution vision in primates. Macular-specific photoreceptor loss and abnormal blood vessel growth within the macula are the major causes of untreatable vision loss worldwide. However, the genes that regulate specialization of the macula, and the causes of its vulnerability to degeneration, remain obscure. Microarrays were used to compare gene expression between macula and non-macular regions during a critical phase of human retinal vascular development. Keywords: Comparison of developing anatomical regions
2008-12-30 | GSE12621 | GEO
Project description:The Unfolded Protein Response is a conserved intracellular signal transduction mechanism that maintains endoplasmic reticulum homeostasis and may contribute to the pathogenesis and progression of human diseases associated with ER stress. Genetic mutations in the UPR regulator, ATF6, lead to vision loss in heritable retinal diseases. Our prior studies found that disease mutations disrupt ATF6 function, but the pathomechanism by which loss of ATF6 activity causes vision loss in people is unknown. To investigate this, we developed retinal organoids from patient iPSCs carrying ATF6 disease mutations and from hESCs bearing CRISPR-edited ATF6 null alleles. Unexpectedly, we found that retinal organoids lacking functional ATF6 failed to form cone photoreceptors while rod photoreceptors were unaffected. We used adaptive optics imaging of the retinas in patients with ATF6 mutations and saw pronounced loss of cones and preservation of rods that closely mirrored our in vitro organoid phenotypes. Last, we found that a small molecule proteostasis agonist partially restored cone photoreceptor outer segment formation and gene expression in ATF6 mutant retinal organoids. Our results reveal a surprising and novel function for ATF6 in human cone photoreceptor development. Our findings identify a potential therapeutic strategy for patients based on small molecule reprogramming of the proteostasis network in the retina. Our study suggests that the UPR guides intrinsic developmental processes in specialized metazoan cell types in addition to its role in responding to extrinsic pathologic events.
2021-06-25 | GSE153302 | GEO
Project description:In inherited retinal disorders such as retinitis pigmentosa (RP), rod photoreceptor-specific mutations cause primary rod degeneration that is followed by secondary cone death and loss of high-acuity vision. Mechanistic studies of retinal degeneration are challenging because of retinal heterogeneity. Moreover, the detection of early cone responses to rod death is especially difficult due to the paucity of cones in the retina. To resolve heterogeneity in the degenerating retina and investigate events in both types of photoreceptors during primary rod degeneration, we utilized droplet-based single-cell RNA sequencing in an RP mouse model, rd10.
2022-04-13 | GSE183206 | GEO
Project description:NRL (Neural retina leucine zipper) is a key regulator of the fate determination and gene expression of rod photoreceptors in the retina of many vertebrates. In this study, we observed a unique retinal phenotype in the nrl knockout zebrafish model characterized by reduced rods with shortened outer segments and gradually increased green-cones in adulthood. By tracing and comparing the developmental processes of WT and nrl knockout rods, we found there might be two waves of rod genesis distinguished as nrl-dependent and independent with different starting times. To reveal the underlying cellular and molecular mechanisms, bulk and single-cell RNA-seq were performed. The changes in gene expression and cell proportion of rods and cones agreed well with our histological and immunofluorescence studies. Interestingly, we found that rods exhibited noticeable heterogeneities in the gene expression patterns, and a part of rods in nrl knockout zebrafish misexpressed the green-cone genes, reflecting a hybrid status of rod and green-cone. Furthermore, we identified mafba as a novel regulator of rod genesis, which was responsible for the development of rods in nrl knockout zebrafish. Our study will largely improve the current understanding of the developmental processes and regulatory mechanisms of rods in zebrafish and probably other species, and may facilitate future studies in the fields of retinal development and retinal degeneration.
2020-10-27 | GSE160140 | GEO