A Novel Approach to Single Cell RNA-Sequence Analysis Facilitates In Silico Gene Reporting of Human Pluripotent Stem Cell-Derived Retinal Cell Types

Phillips, M. Joseph; Jiang, Peng; Howden, Sara E.; Barney, Patrick; Min, Jiyoung; York, Nathaniel W.; Chu, Li‐Fang; Capowski, Elizabeth E.; Cash, Abigail; Jain, Shivani; Barlow, Katherine; Tabassum, Tasnia; Stewart, Ron; Pattnaik, Bikash R.; Thomson, James A.; Gamm, David M.

doi:10.1002/stem.2755

Cited by 54 publications

(102 citation statements)

References 30 publications

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“…Our results suggest differential expression patterns among the seven markers. All seven rod markers are highly abundant, consistent with previous scRNA-seq studies of mouse and human retina (Macosko et al, 2015;Phillips et al, 2018). The seven markers showed differential expression patterns in the six identified rod photoreceptor clusters.…”

Section: Profiling Healthy and Degenerating Human Rod Photoreceptor Ssupporting

confidence: 87%

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A single‐cell transcriptome atlas of the adult human retina

et al. 2019

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The retina is a specialized neural tissue that senses light and initiates image processing. Although the functional organization of specific retina cells has been well studied, the molecular profile of many cell types remains unclear in humans. To comprehensively profile the human retina, we performed single‐cell RNA sequencing on 20,009 cells from three donors and compiled a reference transcriptome atlas. Using unsupervised clustering analysis, we identified 18 transcriptionally distinct cell populations representing all known neural retinal cells: rod photoreceptors, cone photoreceptors, Müller glia, bipolar cells, amacrine cells, retinal ganglion cells, horizontal cells, astrocytes, and microglia. Our data captured molecular profiles for healthy and putative early degenerating rod photoreceptors, and revealed the loss of MALAT1 expression with longer post‐mortem time, which potentially suggested a novel role of MALAT1 in rod photoreceptor degeneration. We have demonstrated the use of this retina transcriptome atlas to benchmark pluripotent stem cell‐derived cone photoreceptors and an adult Müller glia cell line. This work provides an important reference with unprecedented insights into the transcriptional landscape of human retinal cells, which is fundamental to understanding retinal biology and disease.

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Section: Profiling Healthy and Degenerating Human Rod Photoreceptor Ssupporting

confidence: 87%

“…Correlation matrix to benchmark hiPSC‐derived cone photoreceptors (week 15 and week 20; Welby et al , ), fetal cone photoreceptors (Welby et al , ), adult retina (Phillips et al , ), and the human Müller glia cell line MIO‐M1 against all retinal cell types identified in this human neural retina atlas.…”

Section: Resultsmentioning

confidence: 99%

A single‐cell transcriptome atlas of the adult human retina

et al. 2019

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“…Our results suggest differential expression patterns among the 7 markers. All 7 rod markers are highly abundant, consistent with previous scRNA-seq studies of mouse and human retina (Phillips et al, 2018;Macosko et al, 2015). The 7 markers showed differential expression patterns in the 6 identified rod photoreceptor clusters.…”

Section: Profiling Healthy and Degenerating Human Rod Photoreceptor Ssupporting

confidence: 81%

“…To demonstrate the use of our dataset as a benchmarking reference, we compared the scRNA-seq profiles of distinct cell types generated using alternative methods, including fetal human cone photoreceptors, human induced pluripotent stem cell derived-cone photoreceptors (hiPSC-cone; (Welby et al, 2017), and a sample of adult human retina with 139 cells (Phillips et al, 2018). Correlation analysis demonstrated that the adult human retina sample showed highest similarity to rod photoreceptor (0.63, Supplementary figure 10), which is expected as rod photoreceptors represent the majority of the cells in the retina.…”

Section: Using the Human Neural Retina Transcriptome Atlas For Benchmmentioning

confidence: 99%

“…Recently, single cell transcriptomics was used to analyse the human retina. Phillips et al have profiled a total of 139 adult retina cells using the C1 Fluidigm platform (Phillips et al, 2018), but the limited number of profiled cells presents challenges in the annotation and accurate identification of individual retinal cell types. Moreover, a flow cytometry approach was used to isolate 65 human fetal cone photoreceptors followed by scRNA-seq profiling (Welby et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

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Generation of human neural retina transcriptome atlas by single cell RNA sequencing

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Sharov

et al. 2018

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The retina is a highly specialized neural tissue that senses light and initiates image processing. Although the functional organisation of specific cells within the retina has been well-studied, the molecular profile of many cell types remains unclear in humans. To comprehensively profile cell types in the human retina, we performed single cell RNA-sequencing on 20,009 cells obtained post-mortem from three donors and compiled a reference transcriptome atlas. Using unsupervised clustering analysis, we identified 18 transcriptionally distinct clusters representing all known retinal cells: rod photoreceptors, cone photoreceptors, Müller glia cells, bipolar cells, amacrine cells, retinal ganglion cells, horizontal cells, retinal astrocytes and microglia. Notably, our data captured molecular profiles for healthy and early degenerating rod photoreceptors, and revealed a novel role of MALAT1 in putative rod degeneration. We also demonstrated the use of this retina transcriptome atlas to benchmark pluripotent stem cell-derived cone photoreceptors and an adult Müller glia cell line. This work provides an important reference with unprecedented insights into the transcriptional landscape of human retinal cells, which is fundamental to our understanding of retinal biology and disease.

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3D Microstructured Scaffolds to Support Photoreceptor Polarization and Maturation

et al. 2018

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Blinding disorders of the outer retina involve dysfunction and degeneration of photoreceptors. One potential approach to treat these forms of blindness is to repopulate the outer retina via a simple bolus injection of donor photoreceptors. However, this may not be ideal due to the highly polarized organization of photoreceptors that include apical light sensing photopigments and basal axon terminals. Furthermore, bolus injections create uncertainty with regard to the area, density, and retention of donor cells. Here, a novel and robust microfabrication process is developed to create 3D, micrometer-sized complex structures in ultrathin and biocompatible elastomer films (nonbiodegradable polydimethylsiloxane and biodegradable poly(glycerol-sebacate)) that can serve as polarizable photoreceptor delivery scaffolds, consisting of an array of cup-shaped photoreceptor capture wells that funnel into a microchannel. This "wine glass" scaffold design promotes efficient capture of human pluripotent stem-cell-derived photoreceptor cell bodies and guidance of basal axon extensions, ultimately achieving a uniform level of organization and polarization that is not possible with bolus injections or previously described scaffolds. In addition to future therapeutic applications, our scaffold design and materials provide a platform to generate reproducible and scalable in vitro models of photoreceptor-based diseases.

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A Novel Approach to Single Cell RNA-Sequence Analysis Facilitates In Silico Gene Reporting of Human Pluripotent Stem Cell-Derived Retinal Cell Types

Cited by 54 publications

References 30 publications

A single‐cell transcriptome atlas of the adult human retina

A single‐cell transcriptome atlas of the adult human retina

Generation of human neural retina transcriptome atlas by single cell RNA sequencing

3D Microstructured Scaffolds to Support Photoreceptor Polarization and Maturation

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