Retinal progenitor cells release extracellular vesicles containing developmental transcription factors, microRNA and membrane proteins

Abstract Objective Retinal degenerative diseases remain the dominant causes of blindness worldwide, and cell replacement is viewed as a promising therapeutic direction. However, the resources of seed cells are hard to obtain. To further explore this therapeutic approach, human embryonic stem extracellular vesicles (hESEVs) were extracted from human embryonic stem cells (hESCs) to inspect its effect and the possible mechanism on retinal Müller cells and retinal function. Methods hESEVs were extracted by multi-step differential centrifugation, whose morphologies and specific biomarkers (TSG101, CD9, CD63, and CD81) were observed and measured. After hESEVs were injected into the vitreous cavity of RCS rats, the retinal tissues and retinal functions of rats were assessed. The alteration of Müller cells and retinal progenitor cells was also recorded. Microvesicles (MVs) or exosomes (EXOs) were extracted from hESCs transfected with sh-HSP90 or pcDNA3.1-HSP9, and then incubated with Müller cells to measure the uptake of EVs, MVs, or EXOs in Müller cells by immunofluorescence. The retrodifferentiation of Müller cells was determined by measuring Vimentin and CHX10. qRT-PCR and western blot were used to detect HSP90 expression in MVs and evaluate Oct4 level in Müller cells, and Co-IP to inspect the interaction of HSP90 and Oct4. Results RCS rats at the postnatal 30 days had increased retinal progenitor cells which were dedifferentiated from Müller cells. hESEVs were successfully extracted from hESCs, evidenced by morphology observation and positive expressions of specific biomarkers (TSG101, CD9, CD63, and CD81). hESEVs promoted Müller cells dedifferentiated and retrodifferentiated into retinal progenitor cells evidenced by the existence of a large amount of CHX10-positive cells in the retinal inner layer of RCS rats in response to hESEV injection. The promotive role of hESEVs was exerted by MVs demonstrated by elevated fluorescence intensity of CHX10 and suppressed Vimentin fluorescence intensity in MVs rather than in EXOs. HSP90 in MVs inhibited the retrodifferentiation of Müller cells and suppressed the expression level of Oct4 in Müller cells. Co-IP revealed that HSP90 can target Oct4 in Müller cells. Conclusion hESEVs could promote the retrodifferentiation of Müller cells into retinal progenitor cells by regulating the expression of Oct4 in Müller cells by HSP90 mediation in MVs.

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2P-104 In vitro analyses of bHLH transcription factors expressed in retinal progenitor cells(The 46th Annual Meeting of the Biophysical Society of Japan)

Seibutsu Butsuri ◽

10.2142/biophys.48.s91_3 ◽

2008 ◽

Vol 48 (supplement) ◽

pp. S91

Author(s):

Osamu Hisatomi ◽

Koji Hasegawa ◽

Daisuke Kitano ◽

Tatsushi Goto ◽

Fumio Tokunaga ◽

...

Keyword(s):

Transcription Factors ◽

Progenitor Cells ◽

Annual Meeting ◽

Retinal Progenitor Cells ◽

Retinal Progenitor ◽

Biophysical Society ◽

Bhlh Transcription Factors

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Lhx2 regulates temporal changes in chromatin accessibility and transcription factor binding in retinal progenitor cells

10.1101/238279 ◽

2017 ◽

Cited By ~ 1

Author(s):

Cristina Zibetti ◽

Sheng Liu ◽

Jun Wan ◽

Jiang Qian ◽

Seth Blackshaw

Keyword(s):

Transcription Factors ◽

Progenitor Cells ◽

Late Stage ◽

Chromatin Accessibility ◽

Developmental Competence ◽

Open Chromatin ◽

Retinal Progenitor Cells ◽

Retinal Progenitor ◽

Target Sites ◽

Retinal Neurogenesis

AbstractRetinal progenitor cells (RPCs) pass through multiple stages of developmental competence, where they successively acquire and lose the ability to generate individual cell subtypes. To identify the transcriptional regulatory networks that control these transitions, we conducted epigenomic and transcriptomic profiling of early and late-stage RPCs and observed a developmentally dynamic landscape of chromatin accessibility. Open chromatin regions that showed stage-specificity, as well as those shared by early and late-stage RPCs, were selectively targeted by the homeodomain factor Lhx2, which is expressed throughout retinal neurogenesis but also regulates many stage-specific processes in RPCs. Stage-specific Lhx2 binding sites were frequently associated with target sites for transcription factors that are preferentially expressed in either early or late-stage RPCs, and which were predicted to possess pioneer activity. Lhx2 loss of function in RPCs led to a loss of chromatin accessibility at both direct Lhx2 target sites and more broadly across the genome, as well as a loss of binding by transcription factors associated with stage-specific Lhx2 target sites. These findings demonstrate a central role for Lhx2 in control of chromatin accessibility in RPCs, and identify transcription factors that may guide stage-specific target site selection by Lhx2.SummaryLhx2 is a central regulator of chromatin accessibility in retinal progenitor cells, and interacts with stage-specific transcription factors to regulate genes that are dynamically expressed during retinal neurogenesis.

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Human retinal organoids release extracellular vesicles that regulate gene expression in target human retinal progenitor cells

10.1101/2021.02.10.430690 ◽

2021 ◽

Author(s):

Jing Zhou ◽

Miguel Flores-Bellver ◽

Jianbo Pan ◽

Alberto Benito-Martin ◽

Cui Shi ◽

...

Keyword(s):

Gene Expression ◽

Progenitor Cells ◽

Extracellular Vesicles ◽

Noncoding Rna ◽

Tissue Characterization ◽

Retinal Development ◽

Retinal Progenitor Cells ◽

Retina Development ◽

Retinal Tissue ◽

Retinal Progenitor

AbstractThe mechanisms underlying retinal development have not been completely elucidated. Extracellular vesicles (EVs) are novel essential mediators of cell-to-cell communication with emerging roles in developmental processes. Nevertheless, the identification of EVs in human retinal tissue, characterization of their cargo, and analysis of their potential role in retina development has not been accomplished. Three-dimensional retinal tissue derived from human induced pluripotent stem cells (hiPSC) provide an ideal developmental system to achieve this goal. Here we report that hiPSC-derived retinal organoids release exosomes and microvesicles with small noncoding RNA cargo. EV miRNA cargo-predicted targetome correlates with GO pathways involved in mechanisms of retinogenesis relevant to specific developmental stages corresponding to hallmarks of native human retina development. Furthermore, uptake of EVs by human retinal progenitor cells leads to changes in gene expression correlated with EV miRNA cargo predicted gene targets, and mechanisms involved in retinal development, ganglion cell and photoreceptor differentiation and function.

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Mesenchymal Stem Cells: Signaling Pathways in Transdifferentiation Into Retinal Progenitor Cells

Basic and Clinical Neuroscience Journal ◽

10.32598/bcn.9.10.510 ◽

2021 ◽

Vol 12 (1) ◽

pp. 29-42

Author(s):

Hamid Aboutaleb Kadkhodaeian ◽

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Transcription Factors ◽

Signaling Pathways ◽

Progenitor Cells ◽

Retinal Pigment ◽

Host Cells ◽

Retinal Cell ◽

Retinal Progenitor Cells ◽

Retinal Progenitor

Several signaling pathways and transcription factors control the cell fate in its in vitro development and differentiation. The orchestrated use of these factors results in cell specification. In coculture methods, many of these factors secrete from host cells but control the process. Today, transcription factors required for retinal progenitor cells are well known, but the generation of these cells from mesenchymal stem cells is an ideal goal. The purpose of the paper is to review novel methods for retinal progenitor cell production and selecting a set of signaling molecules in the presence of adult retinal pigment epithelium and extraocular mesenchyme acting as inducers of retinal cell differentiation.

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Human Retinal Progenitor Cells Derived Small Extracellular Vesicles Delay Retinal Degeneration: A Paradigm for Cell-free Therapy

Frontiers in Pharmacology ◽

10.3389/fphar.2021.748956 ◽

2021 ◽

Vol 12 ◽

Author(s):

Min Chen ◽

Chunge Ren ◽

Bangqi Ren ◽

Yajie Fang ◽

Qiyou Li ◽

...

Keyword(s):

Retinal Degeneration ◽

Progenitor Cells ◽

Extracellular Vesicles ◽

Target Genes ◽

Outer Nuclear Layer ◽

Subretinal Space ◽

Retinal Progenitor Cells ◽

Retinal Progenitor ◽

Rcs Rats

Retinal degeneration is a leading cause of irreversible vision impairment and blindness worldwide. Previous studies indicate that subretinal injection of human retinal progenitor cells (hRPCs) can delay the progression of retinal degeneration, preserve retinal function, and protect photoreceptor cells from death, albeit the mechanism is not well understood. In this study, small extracellular vesicles derived from hRPCs (hRPC-sEVs) were injected into the subretinal space of retinal dystrophic RCS rats. We find that hRPC-sEVs significantly preserve the function of retina and thickness of the outer nuclear layer (ONL), reduce the apoptosis of photoreceptors in the ONL, and suppress the inflammatory response in the retina of RCS rats. In vitro, we have shown that hRPC-sEV treatment could significantly reserve the low-glucose preconditioned apoptosis of photoreceptors and reduce the expression of pro-inflammatory cytokines in microglia. Pathway analysis predicted the target genes of hRPC-sEV microRNAs involved in inflammation related biological processes and significantly enriched in processes autophagy, signal release, regulation of neuron death, and cell cycle. Collectively, our study suggests that hRPC-sEVs might be a favorable agent to delay retinal degeneration and highlights as a new paradigm for cell-free therapy.

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Human Retinal Organoids Release Extracellular Vesicles That Regulate Gene Expression in Target Human Retinal Progenitor Cells

10.21203/rs.3.rs-275959/v1 ◽

2021 ◽

Author(s):

Jing Zhou ◽

Miguel Flores-Bellver ◽

Jianbo Pan ◽

Alberto Benito-Martin ◽

Cui Shi ◽

...

Keyword(s):

Gene Expression ◽

Progenitor Cells ◽

Extracellular Vesicles ◽

Noncoding Rna ◽

Tissue Characterization ◽

Retinal Development ◽

Retinal Progenitor Cells ◽

Retina Development ◽

Retinal Tissue ◽

Retinal Progenitor

Abstract The mechanisms underlying retinal development have not been completely elucidated. Extracellular vesicles (EVs) are novel essential mediators of cell-to-cell communication with emerging roles in developmental processes. Nevertheless, the identification of EVs in human retinal tissue, characterization of their cargo, and analysis of their potential role in retina development has not been accomplished. Three-dimensional retinal tissue derived from human induced pluripotent stem cells (hiPSC) provide an ideal developmental system to achieve this goal. Here we report that hiPSC-derived retinal organoids release exosomes and microvesicles with small noncoding RNA cargo. EV miRNA cargo-predicted targetome correlates with GO pathways involved in mechanisms of retinogenesis relevant to specific developmental stages corresponding to hallmarks of native human retina development. Furthermore, uptake of EVs by human retinal progenitor cells leads to changes in gene expression correlated with EV miRNA cargo predicted gene targets, and mechanisms involved in retinal development, ganglion cell and photoreceptor differentiation and function.

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