Novel FBXW7 hypermethylation in a carboxymethyl lysine- driven retinal co-culture model: An epigenetic biomarker with translational potential for diabetic retinopathy.

Diabetic retinopathy, a microvascular complication of diabetes, is a primary cause of vision loss. Elevated levels of Nε-(Carboxymethyl) lysine (CML), the predominant advanced glycation end product, are associated with its progression. To delve into the critical aspects of CML-induced epigenetic modulation, this study analysed DNA methylation profiles in a co-culture model following CML exposure. Human Retinal Endothelial Cells (HREC) and Human Retinal Pericytes (HRP) were co-cultured using a 6-well insert model and treated with 3.81 nM of CML under normoglycemic conditions (5.5 mM Glucose) for 24 h. DNA methylation profiling was performed using the Infinium EPIC 935K array and validated by qPCR. Comparative analysis of Illumina Infinium Human Methylation 935K EPIC BeadChip has revealed seven commonly hypermethylated and five hypomethylated genes between HREC and HRP. Markedly, FBXW7 was significantly hypermethylated (Diff Score +13), while SF3B1 and DIXDC1 were hypomethylated (Diff Score -13). Furthermore, protein-protein interaction analysis identified TP53 as a central hub among hypermethylated genes. CpG island mapping showed locus-specific methylation heterogeneity. Gene expression analysis confirmed methylation-associated transcriptional changes, with FBXW7 expression strikingly reduced in the serum of PDR patients (8.169 ± 1.25, n = 21, p value < 0.033). FBXW7 was identified as a significantly differentially methylated marker in the co-culture model, underscoring its potential role as an epigenetic biomarker and therapeutic target. Validation in site-specific biological samples, such as tear fluid, could unveil its application as a predictive marker for early detection.