Reversal of dual epigenetic repression of non-canonical Wnt-5a normalises diabetic corneal epithelial wound healing and stem cells.
Link: https://doi.org/10.1007/s00125-023-05960-1
Authors: Shah, Ruchi; Spektor, Tanya M; Weisenberger, Daniel J; Ding, Hui; Patil, Rameshwar; Amador, Cynthia; Song, Xue-Ying; Chun, Steven T; Inzalaco, Jake; Turjman, Sue; Ghiam, Sean; Jeong-Kim, Jiho; Tolstoff, Sasha; Yampolsky, Sabina V; Sawant, Onkar B; Rabinowitz, Yaron S; Maguen, Ezra; Hamrah, Pedram; Svendsen, Clive N; Saghizadeh, Mehrnoosh; Ljubimova, Julia Y; Kramerov, Andrei A; Ljubimov, Alexander V
Abstract: Diabetes is associated with epigenetic modifications including DNA methylation and miRNA changes. Diabetic complications in the cornea can cause persistent epithelial defects and impaired wound healing due to limbal epithelial stem cell (LESC) dysfunction. In this study, we aimed to uncover epigenetic alterations in diabetic vs non-diabetic human limbal epithelial cells (LEC) enriched in LESC and identify new diabetic markers that can be targeted for therapy to normalise corneal epithelial wound healing and stem cell expression. Human LEC were isolated, or organ-cultured corneas were obtained, from autopsy eyes from non-diabetic (59.87±20.89 years) and diabetic (71.93±9.29 years) donors. The groups were not statistically different in age. DNA was extracted from LEC for methylation analysis using Illumina Infinium 850K MethylationEPIC BeadChip and protein was extracted for Wnt phospho array analysis. Wound healing was studied using a scratch assay in LEC or 1-heptanol wounds in organ-cultured corneas. Organ-cultured corneas and LEC were transfected with WNT5A siRNA, miR-203a mimic or miR-203a inhibitor or were treated with recombinant Wnt-5a (200 ng/ml), DNA methylation inhibitor zebularine (1-20 µmol/l) or biodegradable nanobioconjugates (NBCs) based on polymalic acid scaffold containing antisense oligonucleotide (AON) to miR-203a or a control scrambled AON (15-20 µmol/l). We provide the first account of epigenetic changes in diabetic corneas including dual inhibition of WNT5A by DNA methylation and miRNA action. Overall, Wnt-5a is a new corneal epithelial wound healing stimulator that can be targeted to improve wound healing and stem cells in the diabetic cornea. The DNA methylation dataset is available from the public GEO repository under accession no. GSE229328 ( https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE229328 ).