June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Characterization of miRNA cargo in extracellular vesicles released by polarized human retinal pigment epithelium cells
Author Affiliations & Notes
  • Conan Chen
    CellSight Ocular Stem Cell and Regeneration Program, Department of Ophthalmology, Sue Anschutz-Rodgers Eye Center, University of Colorado - Anschutz Medical Campus, Aurora, Colorado, United States
  • Silvia Aparicio-Domingo
    CellSight Ocular Stem Cell and Regeneration Program, Department of Ophthalmology, Sue Anschutz-Rodgers Eye Center, University of Colorado - Anschutz Medical Campus, Aurora, Colorado, United States
  • Kang Li
    CellSight Ocular Stem Cell and Regeneration Program, Department of Ophthalmology, Sue Anschutz-Rodgers Eye Center, University of Colorado - Anschutz Medical Campus, Aurora, Colorado, United States
  • Robert Dubin
    Center for Epigenomics, Computational Genomics Core, Albert Einstein College of Medicine, Bronx, New York, United States
  • Timothy Domashevich
    CellSight Ocular Stem Cell and Regeneration Program, Department of Ophthalmology, Sue Anschutz-Rodgers Eye Center, University of Colorado - Anschutz Medical Campus, Aurora, Colorado, United States
  • Stephen Redenti
    Biology Doctoral Program, The Graduate School and University Center, Lehman College, Bronx, New York, United States
  • Valeria Canto Soler
    CellSight Ocular Stem Cell and Regeneration Program, Department of Ophthalmology, Sue Anschutz-Rodgers Eye Center, University of Colorado - Anschutz Medical Campus, Aurora, Colorado, United States
  • Miguel Flores-Bellver
    CellSight Ocular Stem Cell and Regeneration Program, Department of Ophthalmology, Sue Anschutz-Rodgers Eye Center, University of Colorado - Anschutz Medical Campus, Aurora, Colorado, United States
  • Footnotes
    Commercial Relationships   Conan Chen None; Silvia Aparicio-Domingo None; Kang Li None; Robert Dubin None; Timothy Domashevich None; Stephen Redenti None; Valeria Canto Soler None; Miguel Flores-Bellver None
  • Footnotes
    Support  The Gates Frontiers Fund The Solich Found Sue Anschutz-Rodgers Fund CellSight Fund unrestricted Research Award from Research to Prevent Blindness National Institute of General Medical Sciences (5SC3GM113782) National Eye Institute (5R21EY026752-02)
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 3914 – A0116. doi:
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    • Get Citation

      Conan Chen, Silvia Aparicio-Domingo, Kang Li, Robert Dubin, Timothy Domashevich, Stephen Redenti, Valeria Canto Soler, Miguel Flores-Bellver; Characterization of miRNA cargo in extracellular vesicles released by polarized human retinal pigment epithelium cells. Invest. Ophthalmol. Vis. Sci. 2022;63(7):3914 – A0116.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : The retinal pigment epithelium (RPE) plays an essential physiological role between the photoreceptor cell layer and the choroid. This interaction is critical in the maintenance of an internal equilibrium within the retina. Several retinal diseases occur as a consequence of RPE dysfunction. Our team has established a differentiation protocol that is efficient in producing pure populations of fully differentiated, functionally mature RPE cells from human induced pluripotent stem cells (hiPSC). The purpose of this study was to analyze and characterize the population of extracellular vesicles (EVs) released by hiPSC-derived RPE cells. EVs are cell-derived nanoscale particles containing a range of molecular cargo including small RNA with emerging roles as essential mediators of cell-to-cell communication in developmental processes.

Methods : We established a differentiation protocol that is efficient in producing functionally mature RPE cells from hiPSC. To characterize EVs secreted by hiPSC-RPE cells, we analyzed the release rate, concentration, morphology and miRNAs content of EVs obtained from hiPSC-RPE. Transcriptomic analysis of miRNA EV cargo provided a comprehensive analysis of this signaling system, including miRNA cargo-predicted targetome correlated with Gene Ontology pathways.

Results : Characteristic EV morphology and size (30-170 nm) was observed by TEM. The average concentration of released EVs corresponded to 7.76 x 1010 particles/ml (apical) and 5.40 x 1010 particles/ml (basal). Transcriptomic analysis identified a total of 208 high confidence miRNAs; 17 detected only in RPE-EVs (1 only found in apical and 5 only in basal). Target gene prediction of differentially expressed miRNAs was carried out against a retinal and choroid database. To gain functional insight on RPE-EV transcriptomic cargo, we conducted over representation analysis and determined several biological processes enricheded in EV-associated miRNAs, including mechanisms of cell communication, signal transduction, cell growth and maintenance, metabolism, oxidative stress, immune response, inflammation, neurogenesis and phagocytosis.

Conclusions : This study represents the first transcriptomic profile from EVs released by hiPSC-derived RPE. EV content provides a snapshot of the originating cell’s phenotype by revealing molecular cargo associated with mechanisms regulating human retinal physiology.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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