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Qun Zeng, Yao Li, Eyup Karahan, Stephen H Tsang, Tongalp H Tezel; LACK OF VERTICAL STABILITY RESULTS IN EPITELIal-MESENCHYMAL TRANSITION OF THE IPS-DERIVED RETINAL PIGMENT EPITHELIAL CELL. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3935. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To test whether vertical stabilization of the scaffolds can prevent epithelial-mesenchymal transition (EMT) of human iPS-derived retinal pigment epithelial cell (hiPS-RPE) grafts.
RPE cells from 5 different hiPS-RPE lines and ARPE-19 cells were cultured to confluence on human Bruch’s membrane explants and tissue culture inserts with a 10µm-thick transparent polyester membrane. One week after the confluence, 6.0 mm grafts were cut and transferred to another dish. These circular grafts were either stabilized on a photopolymerizable biogel or left unstabilized within the culture medium. Grafts were maintained within a culture medium alone or in the presence of 10 ng/ml of TGF-b for another week. At the end of the incubation period, Western blotting, immunofluorescence and reverse transcription quantitative PCR was used to determine the expression epithelial and EMT markers (E-cadherin, N-cadherin, ZO-1, cytokeratin-18, ezrin, RPE 65, Na+/K+-ATPase).
Both ARPE-19 cells and hiPS-RPE grafts on free-floating Bruch’s membrane and polyester membranes exhibited EMT in vitro. Vertical stabilization of the scaffolds using the biomimetic hydrogel suppressed he EMT and maintained epithelial characteristics (18% more E-cadherin expression compared to unstabilized grafts). Presence of TGF- b results in more mesenchymal characteristics (36% less E-cadherin expression; 3.15 x more N-cadherin expression). As a result, cells become fusiform and migratory which reflected itself with a 2.86 x increase in cell perimeter. Induction of EMT with TGF- b was suppressed with the vertical stabilization of the graft.
Vertical stabilization of the hiRPE grafts can suppress EMT in vitro. Vertical stabilization of the RPE grafts should be an integral part of any RPE transplantation attempts to maintain functional RPE sheets in the subretinal space.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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