Single-cell RNA sequencing reveals transcriptional changes of human choroidal and retinal pigment epithelium cells during fetal development, in healthy adult and intermediate age-related macular degeneration

Collin, Joseph; Hasoon, Megan; Zerti, Darin; Hammadi, Sarah; Dorgau, Birthe; Clarke, Lucy; Steel, David; Hussain, Rafiqul; Coxhead, Jonathan; Lisgo, Steven; Lako, Majlinda

doi:10.1093/hmg/ddad007

Cited by 12 publications

(13 citation statements)

References 68 publications

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“…Given the near-ubiquitous Emcn mRNA expression we observed in the mouse choroidal pericyte cluster, we were surprised that EMCN was absent from human pericytes within a previously published choroidal cell atlas, despite expression in choroidal fibroblasts that was similar to our observation in mouse (17). This finding was confirmed by exploring other published human datasets accessible via Spectacle (27), including data from Voigt et al 2022 and Collin et al, 2023 (28,29). Consistent with findings from our initial analysis, limited EMCN expression was detected in pericytes within these datasets, while fibroblast expression appeared widespread.…”

Section: Discussionsupporting

confidence: 82%

Non-endothelial expression of Endomucin in the mouse and human choroid

Brookins,

Serrano,

Yacu

et al. 2024

Preprint

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Endomucin (EMCN) is a 261 AA transmembrane glycoprotein that is highly expressed by venous and capillary endothelial cells where it plays a role in VEGF-mediated angiogenesis and regulation of immune cell recruitment. However, it is better known as a histological marker, where it has become widespread due to the commercial availability of high-quality antibodies that work under a wide range of conditions and in many tissues. The specificity of EMCN staining has been well-validated in retinal vessels, but while it has been used extensively as a marker in other tissues of the eye, including the choroid, the pattern of expression has not been described in detail. Here, in addition to endothelial expression in the choriocapillaris and deeper vascular layers, we characterize a population of EMCN-positive perivascular cells in the mouse choroid that did not co-localize with cells expressing other endothelial markers such as PECAM1 or PODXL. To confirm that these cells represented a new population of EMCN-expressing stromal cells, we then performed single cell RNA sequencing in choroids from adult wild-type mice. Analysis of this new dataset confirmed that, in addition to endothelial cells,EmcnmRNA expression was present in choroidal pericytes and a subset of fibroblasts, but not vascular smooth muscle cells. BesidesEmcn, no known endothelial gene expression was detected in these cell populations, confirming that they did not represent endothelial-stromal doublets, a common technical artifact in single cell RNA seq datasets. Instead, choroidalEmcn-expressing fibroblasts exhibited high levels of chemokine and interferon signaling genes, whileEmcn-negative fibroblasts were enriched in genes encoding extracellular matrix proteins.Emcnexpressing fibroblasts were also detected in published datasets from mouse brain and human choroid, suggesting that stromalEmcnexpression was not unique to the choroid and was evolutionarily conserved. Together, these findings highlight unique fibroblast and pericyte populations in the choroid and provide new context for the role of EMCN in angiogenesis and immune cell recruitment.

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Section: Discussionsupporting

confidence: 82%

Non-endothelial expression of Endomucin in the mouse and human choroid

Brookins,

Serrano,

Yacu

et al. 2024

Preprint

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“…The RPE dataset included >10,000 single nucleus transcriptomes from peripheral samples. Their transcriptomes generally resembled those reported in recent single cell studies (15,(17)(18)(19)(55)(56)(57), although our dataset is currently the largest. Along with known markers, we found several novel markers that are selectively expressed by RPE even when compared to all known ocular cell types (Fig.…”

Section: Pigmented Cellssupporting

confidence: 76%

“…They include WFDC1, CXCL14, and CALCB enriched in macular RPE, and PMEL and IGFBP5 enriched in peripheral RPE. Many of these genes were also reported to be differentially expressed in previous studies (15,(17)(18)(19)(55)(56)(57)(58).…”

Section: Pigmented Cellsmentioning

confidence: 57%

“…In previous studies, we used high-throughput single-cell and single-nucleus RNA sequencing (scRNA-seq and snRNA-seq) to catalog and molecularly characterize the cell types of the human anterior segment (9,10). We and others also used scRNA-seq and snRNA-seq to characterize the adult human neural retina and RPE (11)(12)(13)(14)(15)(16)(17)(18)(19). Here, to complete a "v.1" cell atlas of the human eye, we used snRNA-seq to analyze the extra-retinal tissues of the posterior segment.…”

Section: Introductionmentioning

confidence: 99%

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Transcriptomic Analysis of the Ocular Posterior Segment Completes a Cell Atlas of the Human Eye

Monaverfeshani

Yan

Pappas

et al. 2023

Preprint

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Although the visual system extends through the brain, most vision loss originates from defects in the eye. Its central element is the neural retina, which senses light, processes visual signals, and transmits them to the rest of the brain through the optic nerve (ON). Surrounding the retina are numerous other structures, conventionally divided into anterior and posterior segments. Here we used high-throughput single nucleus RNA sequencing (snRNA-seq) to classify and characterize cells in the extraretinal components of the posterior segment: ON, optic nerve head (ONH), peripheral sclera, peripapillary sclera (PPS), choroid, and retinal pigment epithelium (RPE). Defects in each of these tissues are associated with blinding diseases, for example, glaucoma (ONH and PPS), optic neuritis (ON), retinitis pigmentosa (RPE), and age-related macular degeneration (RPE and choroid). From ~151,000 single nuclei, we identified 37 transcriptomically distinct cell types, including multiple types of astrocytes, oligodendrocytes, fibroblasts, and vascular endothelial cells. Our analyses revealed a differential distribution of many cell types among distinct structures. Together with our previous analyses of the anterior segment and retina, the new data complete a Version 1 cell atlas of the human eye. We used this atlas to map the expression of >180 genes associated with the risk of developing glaucoma, which is known to involve ocular tissues in both anterior and posterior segments as well as neural retina. Similar methods can be used to investigate numerous additional ocular diseases, many of which are currently untreatable.

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“…The presence of NLRP3 inflammasome in RPE cells has been debated in age-related macular degeneration research. Investigations of NLRP3 expression in single-cell human data for the RPE-choroid tissues show low expression of NLRP3 compared to macrophages [ 91 , 92 ]. In support of this finding, a study of human RPE cell lines found no mRNA, protein, or evidence of NLRP3 inflammasome activation in the RPE [ 93 ].…”

Section: Ocular Expression Of Tlrs and Nlrp3mentioning

confidence: 99%

The contribution of pattern recognition receptor signalling in the development of age related macular degeneration: the role of toll-like-receptors and the NLRP3-inflammasome

Brandli,

Vessey,

Fletcher

2024

J Neuroinflammation

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Age-related macular degeneration (AMD) is a leading cause of irreversible vision loss, characterised by the dysfunction and death of the photoreceptors and retinal pigment epithelium (RPE). Innate immune cell activation and accompanying para-inflammation have been suggested to contribute to the pathogenesis of AMD, although the exact mechanism(s) and signalling pathways remain elusive. Pattern recognition receptors (PRRs) are essential activators of the innate immune system and drivers of para-inflammation. Of these PRRs, the two most prominent are (1) Toll-like receptors (TLR) and (2) NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3)-inflammasome have been found to modulate the progression of AMD. Mutations in TLR2 have been found to be associated with an increased risk of developing AMD. In animal models of AMD, inhibition of TLR and NLRP3 has been shown to reduce RPE cell death, inflammation and angiogenesis signalling, offering potential novel treatments for advanced AMD. Here, we examine the evidence for PRRs, TLRs2/3/4, and NLRP3-inflammasome pathways in macular degeneration pathogenesis.

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Single-cell RNA sequencing reveals transcriptional changes of human choroidal and retinal pigment epithelium cells during fetal development, in healthy adult and intermediate age-related macular degeneration

Cited by 12 publications

References 68 publications

Non-endothelial expression of Endomucin in the mouse and human choroid

Non-endothelial expression of Endomucin in the mouse and human choroid

Transcriptomic Analysis of the Ocular Posterior Segment Completes a Cell Atlas of the Human Eye

The contribution of pattern recognition receptor signalling in the development of age related macular degeneration: the role of toll-like-receptors and the NLRP3-inflammasome

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