Abstract
Purpose :
A distinct boundary exists between the progenitor cells of the limbal epithelium and the more differentiated corneal epithelium. We have shown that reciprocal expression patterns of EphA2 and Ephrin-A1 are likely to contribute to normal limbal-corneal epithelial compartmentalization as well as playing a role in response to injury. How this signaling axis is regulated remains unclear. We have demonstrated that microRNAs (miRNAs) play critical roles in corneal epithelial wound healing and several miRNAs (e.g. miR-210) have been predicted to target Ephrins. Therefore, we investigated whether: (i) miR-210, a corneal epithelial-preferred miRNA could modify EphA2/Ephrin-A1 signaling; and (ii) such modifications have a role in corneal epithelial migration after injury.
Methods :
RNA-seq analysis was performed on human corneal epithelial cells (HCECs) treated with antago-210 and validated by real time qPCR. Retroviral transduction was used to manipulate Ephrin-A1 levels in a human corneal epithelial cell line, hTCEpi. Antago-210 and siRNA knock-down were used to study changes in scratch wound healing in hTCEpi.
Results :
Previous expression profiling experiments demonstrated that miR-210 is prominently expressed in corneal epithelial cells. Interestingly, EphA2 is a potential target for miR-210. RNA-seq data acquired from miR-210-depleted HCECs showed up-regulation of genes involved in cellular migration. In addition, miR-210 is decreased after corneal injury while EphA2 is increased. Moreover, antago-210-treated HCECs markedly enhanced wound closure in a scratch wound assay. Antago-210 treatment resulted in increased EphA2 protein levels as well as pS897-EphA2, the pro-migratory form of EphA2. As expected, Ephrin-A1 levels were reduced, while levels of a well-known target of miR-210, Ephrin-A3, were increased by antago-210 treatment. The increase in migration with antago-210 could be inhibited by Ephrin-A1 overexpression, Ephrin-A1-Fc treatment or siRNA depletion of EphA2. However, depletion of Ephrin-A3 did not have effects on antago-210-induced increase in migration. In addition, Ephrin-A1 overexpression and siEphA2 dampened EGFR signaling, which is increased by antago-210.
Conclusions :
Our data clearly demonstrate a link between miR-210 and EphA2/Ephrin-A1 signaling that regulates, in part, corneal epithelial migration. This interaction has potential in controlling the limbal-corneal epithelial boundary.
This is a 2020 ARVO Annual Meeting abstract.