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Updated: Jul 13, 2026
MicroRNA Expression Profiles of Human iPS Cells, Retinal Pigment Epithelium Derived From iPS, and Fetal Retinal Pigment Epithelium
Published on: June 24, 2014
Small Extracellular Vesicle-Associated MiRNAs in Polarized Retinal Pigmented Epithelium
Belinda J Hernandez1, Madison Strain2, Maria Fernanda Suarez1
1Department of Ophthalmology, Duke University School of Medicine, Durham, North Carolina, United States.
Purpose:
Oxidative stress in the retinal pigmented epithelium (RPE) has been implicated in age-related macular degeneration by impacting endocytic trafficking, including the formation, content, and secretion of extracellular vesicles (EVs). Using our model of polarized primary porcine RPE (pRPE) cells under chronic subtoxic oxidative stress, we tested the hypothesis that RPE miRNAs packaged into EVs are secreted in a polarized manner and contribute to maintaining RPE homeostasis.
Methods:
Small EVs (sEVs) enriched for exosomes were isolated from apical and basal conditioned media from pRPE cells grown for up to four weeks with or without low concentrations of hydrogen peroxide using two sEV isolation methods, leading to eight experimental groups. The sEV miRNA expression was profiled using miRNA-Seq with Illumina MiSeq, followed by quality control and bioinformatics analysis for differential expression using the R computing environment. Expression of selected miRNAs were validated using qRT-PCR.
Results:
We identified miRNA content differences carried by sEVs isolated using two ultracentrifugation-based methods. Regardless of the sEV isolation method, miR-182 and miR-183 were enriched in the cargo of apically secreted sEVs, and miR-122 in the cargo of basally secreted sEVs from RPE cells during normal homeostatic conditions. After oxidative stress, miR-183 levels were significantly decreased in the cargo of apically released sEVs from stressed RPE cells.
Conclusions:
We curated RPE sEV miRNA datasets based on cell polarity and oxidative stress. Unbiased miRNA analysis identified differences based on polarity, stress, and sEV isolation methods. These findings suggest that miRNAs in sEVs may contribute to RPE homeostasis and function in a polarized manner.
Insights
Retinal pigmented epithelium (RPE) cells secrete extracellular vesicles (EVs) containing specific microRNAs (miRNAs) in a polarized manner. Oxidative stress alters this secretion, impacting RPE homeostasis and potentially age-related macular degeneration.
Area of Science:
- Cell Biology
- Molecular Biology
- Ophthalmology
Background:
- Oxidative stress in retinal pigmented epithelium (RPE) is linked to age-related macular degeneration (AMD).
- Endocytic trafficking and extracellular vesicle (EV) secretion are crucial for RPE function.
- Polarized secretion of EVs by RPE cells is not fully understood.
Purpose of the Study:
- To investigate if RPE cells secrete microRNAs (miRNAs) packaged into EVs in a polarized manner.
- To determine if RPE-secreted EVs contribute to maintaining RPE homeostasis under oxidative stress.
- To test the hypothesis that RPE miRNAs in EVs are secreted directionally.
Main Methods:
- Primary porcine RPE (pRPE) cells were cultured under chronic oxidative stress conditions.
- Small EVs (sEVs) were isolated from apical and basal conditioned media using two ultracentrifugation methods.
- sEV miRNA content was analyzed using miRNA-Seq and validated with qRT-PCR.
Main Results:
- Distinct miRNA profiles were identified in apically and basally secreted sEVs.
- miR-182 and miR-183 were enriched in apical sEVs; miR-122 was enriched in basal sEVs under homeostatic conditions.
- Oxidative stress led to a significant decrease in miR-183 levels within apically released sEVs.
Conclusions:
- RPE cells exhibit polarized secretion of specific miRNAs via sEVs.
- Oxidative stress alters the miRNA cargo of secreted sEVs.
- Polarized miRNA secretion in EVs may play a role in RPE homeostasis and AMD pathogenesis.
