Unique methylation signature of painful diabetic neuropathy

Aims: Epigenetic modifications, such as DNA methylation, regulate gene expression without altering the DNA sequence. Methylation, primarily occurring at CpG sites, is a reversible mechanism influencing gene transcription. CpG islands, regions rich in CpG sites near genes, play a crucial role in gene expression regulation. Genome-wide DNA methylation patterns have been studied in diabetic peripheral neuropathy and Charcot neuro-osteoarthropathy. However, this is the first genome-wide DNA methylation study investigating the epigenetic signature of neuropathic pain in type 2 diabetes mellitus (T2DM).

Methods: Two independent European cohorts were analyzed: PROPGER (German Diabetes Center, Düsseldorf) with 72 patients with painful diabetic neuropathy (PDN) and 67 with painless diabetic neuropathy (PLDN); and PROPENG (University of Manchester) with 27 PDN and 65 PLDN patients. Diabetic neuropathy was diagnosed using the Toronto Consensus guidelines, and intraepidermal nerve fiber density (IENF/mm) was measured. PDN classification followed the Treede et al. criteria (Neurology 2008;70:1630–5). Pain frequency and severity were assessed via the pain intensity numerical rating scale (PI-NRS).

Whole-genome methylation was evaluated using the Illumina Infinium Methylation EPIC v1.0 BeadChip. Several analyses were conducted: Differential Methylation Analysis (DMA) identified CpG sites with significant methylation differences between PDN and PLDN; Pathway Enrichment Analysis (PEA) explored functional pathways using KEGG and OMIM databases; Cluster Analysis applied Principal Component Analysis (PCA) and Multidimensional Scaling (MDS) to determine whether methylation signatures could distinguish PDN and PLDN; stepwise gene selection identified candidate genes with the most robust differentially methylated CpGs based on significance thresholds and methylation differences.

Results: Whole-genome methylation analysis revealed significant differences between PDN and PLDN, with specific genes exhibiting hypomethylation or hypermethylation patterns related to pain. Three genes were particularly noteworthy: 1) GCH1 (GTP Cyclohydrolase 1), involved in tetrahydrobiopterin (BH4) biosynthesis, a key regulator of pain-related phenotypes. It was significantly hypomethylated in PDN; 2) MYT1L (Myelin Transcription Factor 1-like) associated with neuronal function, fibromyalgia, and neuropsychiatric disorders. Three CpG sites were significantly hypomethylated in PDN. 3) MED16 (Mediator Complex Subunit 16) a transcriptional regulatory complex component, which was hypomethylated in PDN. This is the first reported association between MED16 and PDN. Pathway Enrichment Analysis identified TGF-beta signaling, purine metabolism, and circadian rhythm regulation as key pathways involved in pain perception, inflammation, and diabetes progression. PCA and MDS revealed distinct epigenetic signatures between PDN and PLDN, although these were not entirely predictive.

Conclusions: Distinct DNA methylation signatures differentiate PDN and PLDN phenotypes in T2DM. The GCH1, MYT1L, and MED16 genes were identified as potential therapeutic targets for neuropathic pain.

Comments: This study provides evidence of unique methylation patterns distinguishing painful from painless diabetic neuropathy, suggesting epigenetic modifications contribute to neuropathic pain susceptibility. The identified CpG sites may serve as potential therapeutic targets.

Although these epigenetic markers enhance the understanding of pain mechanisms, they are not yet suitable for clinical diagnosis. Further studies are needed to validate these findings and assess their predictive value. Future longitudinal research should explore whether these methylation changes can predict pain development and inform personalized treatments. Additionally, this study paves the way for investigating other chronic pain conditions such as cancer, surgical, and transplant-related pain, using a similar epigenetic approach.

Fabiana Picconi

Reference: Kwiatkowska KM, Garagnani P, Bonafé M, Bacalini MG, Sala C, Castellani G, Gentilini D, Calzari L, Ziegler D, Gerrits MM, Faber CG, Malik RA, Marchi M, Salvi E, Lauria G, Pirazzini C. High-resolution whole-genome DNA methylation revealed unique signatures of painful diabetic neuropathy. Diabetes. 2025 Jan 8:db240930. doi: 10.2337/db24-0930. Epub ahead of print. PMID: 39774670.

🔗 https://diabetesjournals.org/diabetes/article/doi/10.2337/db24-0930/157700/High-resolution-whole-genome-DNA-methylation