Investigating the role of epithelial to mesenchymal transition (EMT) regulators TCF4 abd TCF-beta in Fuchs corneal dystrophy
Authors: Judy Yan1, Keya Patel1, Narisa Dhupar2, Ness Little2, Stephan Ong Tone 2. 1Sunnybrook Research Institute, 2Sunnybrook Research Institute, University of Toronto.
Author Disclosures: J. Yan: None. K. Patel: None. N. Dhupar: None. N. Little: None. S. Ong Tone: Any direct financial payments including receipt of honoraria; Name of for-profit or not-for-profit organization(s); RxRenewal. Any direct financial payments including receipt of honoraria; Description of relationship(s); Consultant. Funded grants or clinical trials; Name of for-profit or not-for-profit organization(s); Fighting Blindness Canada, Eye Bank Association of America, TD Pooler Charitable Grant. Funded grants or clinical trials; Description of relationship(s); Principal Investigator Research Grants, Principal Investigator Research Grants, Principal Investigator Research Grants.
Abstract Body:
Purpose: Fuchs endothelial corneal dystrophy (FECD) is a complex ocular disease characterized by the progressive decline and morphological changes of corneal endothelial cells (CECs) that leads to corneal edema. The most common mutation in FECD is a trinucleotide CTG repeat expansion in the third intron of the transcription factor 4 (TCF4) gene. Studies on the effects of CTG repeats on TCF4 gene expression in FECD have shown inconsistent results, but recent evidence supports increased TCF4 isoform expression. Similarly, increased levels of transforming growth factor-β (TGF-β) 1 and 2 in FECD patients have been reported. Both TCF4 and TGF-β are involved in regulation of EMT and have been implicated in FECD pathogenesis. However, the interplay between TCF4, TGF-β and EMT in FECD remains to be elucidated. This study aims to determine whether TCF4 overexpression and TGF-β stimulation are sufficient to cause an FECD-like phenotype in normal CECs.
Study Design: We generated stable corneal endothelial cell lines derived from normal donors or FECD patients that overexpressed different TCF-4 isoforms (-A, -B, -C).
Methods: Western blot was performed to probe for EMT markers. Filamentous-actin staining was used to examine morphological changes. Scratch assay was performed to measure wound closure rates.
Results: We found that overexpression of TCF4 isoforms, even in the context of increased TGF-β1/-β2, did not increase EMT markers (fibronectin, ZEB1), change morphology or alter wound closure rates in normal cells. However, overexpression of TCF4-B isoform, but not TCF4-A or TCF4-C isoforms, in FECD cells resulted in increased cellular migration speeds compared to controls (p<0.05).
Conclusions: Overexpression of TCF4 in the presence of increased TGF-β1/-β2 is insufficient to induce an FECD-like phenotype in normal cells, suggesting that additional dysregulated pathways are likely required to induce FECD. In contrast, dysregulation of TCF4 in FECD cells led to altered cellular migration. Future studies will investigate the mechanistic link between TCF4-B and cell migration, and provide further insight into FECD pathogenesis.
