Abstract
Purpose :
Emerging evidence suggest the critical role of N6-methyladenosine (m6A) in disease development. However, the implication of m6A regulators in primary open-angle glaucoma (POAG) is still largely unknown.
Methods :
Bioinformatic analysis was performed to paint the expression landscape of m6A regulators in trabecular meshwork (TM) tissues of POAG patients. H2O2 stimulation was performed to contrust oxidative stress cell model. SiRNA was used to silence gene expression. CCK-8 assay and transwell were used to investigate the proliferation and migration. RNA sequencing was performed to identify downstream targets.
Results :
19 normal, 17 POAG samples of TM tissues from the Gene Expression Omnibus (GSE27276) database were used to profile the expression of m6A regulators in POAG. Of the 25 m6A regulators, 5 exhibited a significant differential expression. The results of the Random Forest model showed that all five differentially expressed m6A regulators were characteristic genes of POAG and exhibited diagnostic potential for POAG and a nomogram was successfully established. The consensus clustering analysis for different POAG samples were achieved based on gene expression values of five important m6A regulators. As per the clustering outcomes, the POAG samples were efficaciously separated into two groups. Immune cell infiltration analysis showed that activated CD8+ T cell and regulatory T cell differed between the two m6Acluster subtypes. Notably, correlation analysis indicated that the ALKBH5 and RBM15 might correlate with the immune response. Further, human trabecular meshwork cells (HTMCs) oxidative stress cell model and TGF-β2 stimulation cell model were constructed. YTHDC2 was significantly down-regulated in both models. TGF-β2 stimulation caused a decrease in the proliferation of HTMCs and increased the rate of migration and the silence of YTHDC2 showed a reverse outcome. Finally, we silenced YTHDC2 in HTMCs and explored its potential m6A related downstream targets through bioinformatic analysis of the RNA sequencing results.
Conclusions :
M6A regulatory genes in TM tissues could serve as potential diagnostic biomarkers for POAG. The TGF-β2-YTHDC2 axis may exist in TM tissues of POAG patients, and promotes the progression of POAG by inhibiting cell activity and promoting cell migration.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.