Reversal of dual epigenetic repression of non-canonical Wnt-5a normalises diabetic corneal epithelial wound healing and stem cells.
Corneal stem cells
DNA methylation
Diabetic keratopathy
Epigenetics
Gene therapy
Wnt signalling
Wnt-5a
miRNA
Journal
Diabetologia
ISSN: 1432-0428
Titre abrégé: Diabetologia
Pays: Germany
ID NLM: 0006777
Informations de publication
Date de publication:
10 2023
10 2023
Historique:
received:
10
02
2023
accepted:
18
05
2023
medline:
5
9
2023
pubmed:
18
7
2023
entrez:
17
7
2023
Statut:
ppublish
Résumé
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). There was significant differential DNA methylation between diabetic and non-diabetic LEC. WNT5A promoter was hypermethylated in diabetic LEC accompanied with markedly decreased Wnt-5a protein. Treatment of diabetic LEC and organ-cultured corneas with exogenous Wnt-5a accelerated wound healing by 1.4-fold (p<0.05) and 37% (p<0.05), respectively, and increased LESC and diabetic marker expression. Wnt-5a treatment in diabetic LEC increased the phosphorylation of members of the Ca 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 ).
Identifiants
pubmed: 37460827
doi: 10.1007/s00125-023-05960-1
pii: 10.1007/s00125-023-05960-1
pmc: PMC10474199
doi:
Substances chimiques
Wnt-5a Protein
0
MicroRNAs
0
RNA, Small Interfering
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
1943-1958Subventions
Organisme : NEI NIH HHS
ID : R01 EY013431
Pays : United States
Organisme : NEI NIH HHS
ID : R01 EY025377
Pays : United States
Organisme : NCI NIH HHS
ID : R01 CA206220
Pays : United States
Organisme : NEI NIH HHS
ID : R01 EY031377
Pays : United States
Informations de copyright
© 2023. The Author(s).
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