Sodium channels enable fast electrical signaling and regulate phagocytosis in the retinal pigment epithelium

Julia, Johansson; Karema-Jokinen, Viivi I.; Hakanen, Satu; Jylhä, Antti; Uusitalo, Hannu; Vihinen‐Ranta, Maija; Skottman, Heli; Ihalainen, Teemu O.; Nymark, Soile

doi:10.1186/s12915-019-0681-1

Cited by 16 publications

(37 citation statements)

References 76 publications

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“…Single cells obtained by this approach were heavily pigmented and often exhibited two lobes of variable size (Figure 1C). This shape is consistent with an apical and basal cellular domain separated by a junctional actin band as observed in other studies, in which single RPE cells were isolated from native or cultured sheets [37][38][39]. FACS-purified cells were concentrated and applied to the wells of an scRNA-seq chip to create a bar-coded cDNA library for sequencing (Figure 1A).…”

Section: Preparation Of Single Rpe Cellssupporting

confidence: 85%

Single-cell RNA sequencing reveals molecular features of postnatal maturation in the murine retinal pigment epithelium

Pandey

Krebs

Bolisetty

et al. 2022

Preprint

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Transcriptomic analysis of the mammalian retinal pigment epithelium (RPE) aims to identify cellular networks that influence ocular development, maintenance, function, and disease. However, available evidence points to RPE cell heterogeneity in the native tissue, which adds complexity to transcriptomic analysis. Here, to assess cell heterogeneity, we performed single-cell RNA sequencing of RPE cells from two young adult male C57BL/6J mice. Following quality control to ensure robust transcript identification limited to cell singlets, we detected 13,858 transcripts among 2,667 and 2,846 RPE cells, respectively. Dimensional reduction by principal component analysis and uniform manifold approximation and projection revealed six distinct cell popu-lations. All clusters expressed transcripts typical of RPE cells; the smallest (C1, containing 1–2% of total cells) exhibited hallmarks of stem and/or progenitor cells. Placing C1–6 along a pseudotime axis suggested a relative decrease in melanogenesis and stem/progenitor gene expression, and a corresponding increase in visual cycle gene expression upon RPE maturation. K-means testing of all detected transcripts identified additional expression patterns that may advance understanding of RPE stem/pro-genitor cell maintenance and the evolution of cellular metabolic networks during development. This work provides new insights into the transcriptome of the mouse RPE and a baseline for identifying experimentally induced transcriptional changes in future studies of this tissue.

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Section: Preparation Of Single Rpe Cellssupporting

confidence: 85%

Single-cell RNA sequencing reveals molecular features of postnatal maturation in the murine retinal pigment epithelium

Pandey

Krebs

Bolisetty

et al. 2022

Preprint

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“…The candidate genes in this list are enriched in the ion homeostasis pathway. This is expected as previous studies implicated ion channels in POS phagocytosis such as voltage-gated sodium channels 74 and the Ltype calcium channel Ca v 1.3. 61 The list also contains known genes implicated in POS phagocytosis such as Mfge8 75 and Myl3.…”

Section: Discussionsupporting

confidence: 75%

Core circadian clock genes Per1 and Per2 regulate the rhythm in photoreceptor outer segment phagocytosis

et al. 2021

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Retinal photoreceptors undergo daily renewal of their distal outer segments, a process indispensable for maintaining retinal health. Photoreceptor outer segment (POS) phagocytosis occurs as a daily peak, roughly about 1 hour after light onset. However, the underlying cellular and molecular mechanisms which initiate this process are still unknown. Here we show that, under constant darkness, mice deficient for core circadian clock genes (Per1 and Per2) lack a daily peak in POS phagocytosis. By qPCR analysis, we found that core clock genes were rhythmic over 24 hours in both WT and Per1, Per2 double mutant whole retinas. More precise transcriptomics analysis of laser capture microdissected WT photoreceptors revealed no differentially expressed genes between time points preceding and during the peak of POS phagocytosis. In contrast, we found that microdissected WT retinal pigment epithelium (RPE) had a number of genes that were differentially expressed at the peak phagocytic time point compared to adjacent ones. We also found a number of differentially expressed genes in Per1, Per2 double mutant RPE compared to WT ones at the peak phagocytic time point. Finally, based on STRING analysis, we found a group of interacting genes that potentially drive POS phagocytosis in the RPE. This potential pathway consists of genes such as: Pacsin1, Syp, Camk2b, and Camk2d among others. Our findings indicate that Per1 and Per2 are necessary clock components for driving POS phagocytosis and suggest that this process is transcriptionally driven by the RPE.

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“…The candidate genes in this list are enriched in the ion homeostasis pathway. This is expected as previous studies implicated ion channels in POS phagocytosis such as voltage gated sodium channels [69] and the L-type calcium channel Cav1.3 [55]. The list also contains known genes implicated in POS phagocytosis such as Mfge8 [70] and Myl3 [71].…”

Section: Discussionmentioning

confidence: 54%

Core Circadian Clock GenesPer1andPer2regulate the Rhythm in Photoreceptor Outer Segment Phagocytosis

Milićević

Hakkari

Bagchi

et al. 2020

Preprint

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Retinal photoreceptors undergo daily renewal of their distal outer segments, a process indispensable for maintaining retinal health. Photoreceptor Outer Segment (POS) phagocytosis occurs as a daily peak, roughly about one hour after light onset. However, the underlying cellular and molecular mechanisms which initiate this process are still unknown. Here we show that, under constant darkness, mice deficient for core circadian clock genes (Per1 and Per2), lack a daily peak in POS phagocytosis. By qPCR analysis we found that core clock genes were rhythmic over 24h in both WT and Per1, Per2 double mutant whole retinas. More precise transcriptomics analysis of laser capture microdissected WT photoreceptors revealed no differentially, expressed genes between time-points preceding and during the peak of POS phagocytosis. By contrast, we found that microdissected WT retinal pigment epithelium (RPE) had a number of genes that were differentially expressed at the peak phagocytic time-point compared to adjacent ones. We also found a number of differentially expressed genes in Per1, Per2 double mutant RPE compared to WT ones at the peak phagocytic time-point. Finally, based on STRING analysis we found a group of interacting genes which potentially drive POS phagocytosis in the RPE. This potential pathway consists of genes such as: Pacsin1, Syp, Camk2b and Camk2d among others. Our findings indicate that Per1 and Per2 are necessary clock components for driving POS phagocytosis and suggest that this process is transcriptionally driven by the RPE.DeclarationsFundingThis project has been funded with support from the NeuroTime Erasmus+ grant (European Union), Rotterdamse Stichting Blindenbelangen (Netherlands), Nelly Reef fund (Netherlands), Stichting voor Ooglijders (Netherlands), Stichting tot Verbetering van het Lot der Blinden (Netherlands) and Retina France (France).Conflicts of interest/Competing interestsThe authors declare no competing interests.Availability of data and materialData supporting the conclusions of this article are included within the article and are available from the corresponding authors on reasonable request.Code availabilityThe R code for analysis is available from the corresponding authors on reasonable request.Ethics approvalAll experimental procedures were performed in accordance with the Association for Research in Vision and Ophthalmology Statement on Use of Animals in Ophthalmic and Vision Research, as well as with the European Union Directive (2010/63/EU).Consent to participateNot applicableConsent for publicationAll authors read and approve of the contents of this manuscript.Author contributionsN.M. performed experiments, analysis, prepared figures, wrote the manuscript and obtained funding. O.A.-H.H. performed experiments, data analysis, prepared figures and obtained funding. P.D.M. and A.J. performed bioinformatics analysis and edited the manuscript. U.B., J.B.t.B. and C.S. provided technical assistance, performed experiments, prepared figures and edited the manuscript. D.H., A.A.B. and M.-P.F.-S. conceptualized and directed the project, obtained funding, provided resources, performed analysis and edited the manuscript.

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Sodium channels enable fast electrical signaling and regulate phagocytosis in the retinal pigment epithelium

Cited by 16 publications

References 76 publications

Single-cell RNA sequencing reveals molecular features of postnatal maturation in the murine retinal pigment epithelium

Single-cell RNA sequencing reveals molecular features of postnatal maturation in the murine retinal pigment epithelium

Core circadian clock genes Per1 and Per2 regulate the rhythm in photoreceptor outer segment phagocytosis

Core Circadian Clock GenesPer1andPer2regulate the Rhythm in Photoreceptor Outer Segment Phagocytosis

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