Purchase this article with an account.
Whitney Greene, Teresa A Burke, Ramesh Kaini, Heuy-Ching Hetty Wang; Expression of pro-fibrotic miRNAs in an in vitro model of Proliferative Vitreoretinopathy. Invest. Ophthalmol. Vis. Sci. 2017;58(8):5970.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Proliferative vitreoretinopathy (PVR) is the result of aberrant wound healing and fibrosis following a retinal tear or detachment. MicroRNAs (miRNAs) may promote the pathogenesis of fibrosis by regulating the expression of genes involved in TGF-β signaling, epithelial-mesenchymal transition, extracellular matrix remodeling and cell adhesion, and inflammation. An in vitro model of PVR that uses retinal pigment epithelium derived from induced pluripotent stem cells (iPS-RPE) was used to investigate the expression of pro-fibrotic miRNAs.
iPS-RPE was cultured on matrigel-coated transwells for 60 days until fully confluent and pigmented. Cell monolayers were scratched to create a wound. To recapitulate the in vivo conditions that lead to the development of PVR, one set of cells was treated with 5% fresh vitreous immediately after wounding. Unscratched cells and fresh vitreous alone were included as controls. Cells were lysed on Days 0, 3, and 12 post wounding. Total RNA including small RNA was extracted. RNA was reverse transcribed to prepare cDNA. A commercially-available human fibrosis miRNA PCR array was used to analyze expression of 84 distinct miRNAs associated with fibrosis. Data was analyzed to identify pro-fibrotic miRNAs that were differentially expressed during wound healing 4-fold or greater compared to controls.
Analysis of eleven sample sets revealed a distinctive pattern of miRNA expression. Wounded cells had increased expression of 43 pro-fibrotic miRNAs (Day 3), and 30 pro-fibrotic miRNAs (Day 12) compared to control cells. The most highly expressed miRNAs during wound healing included miR-122-5p (Day 3); and miR-101-3p, miR-150-5p, miR-223-3p (Day 12). Cells treated with vitreous immediately after wounding had increased expression of 12 pro-fibrotic miRNAs (Day 3) and 36 pro-fibrotic miRNAs (Day 12) compared to unscratched cells also treated with vitreous. The most highly expressed miRNAs during wound healing with vitreous treatment included miR-10a-5p (Day 3); and miR-10a-5p and miR-133a-3p (Day 12).
The results of this study reveal distinct patterns of miRNA expression during wound healing. The expression of pro-fibrotic miRNAs was segregated according to wounding versus control, time post-wound, and exposure to vitreous. The profiling results from this array may yield insights into the molecular mechanisms that underlie the pathogenesis of PVR.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
This PDF is available to Subscribers Only