Obesity, characterized by an imbalance in energy intake and expenditure leading to excessive adipose tissue expansion, is a major risk factor for type 2 diabetes (T2D). While genetic predisposition plays a role, it doesn't fully explain individual susceptibility to T2D in obesity. Epigenetic mechanisms, such as DNA methylation, are believed to be involved in the interplay between genetic background and environmental factors. DNA methylation, the addition of a methyl group to a cytosine base, can alter gene expression without changing the underlying DNA sequence. Previous studies have linked alterations in DNA methylation to both obesity and T2D, suggesting a potential role in disease development. However, the relationship between DNA methylation and gene expression in the context of T2D development in obesity remains poorly understood. This study aimed to address this gap by investigating DNA methylation and gene expression changes in visceral adipose tissue (VAT) from obese women with and without T2D. The researchers hypothesized that specific changes in DNA methylation and gene expression patterns in VAT could underlie T2D susceptibility in obese individuals. Understanding these mechanisms could lead to the identification of new therapeutic targets and improved risk stratification for T2D.

The existing literature supports a strong association between obesity and T2D, with visceral adipose tissue (VAT) accumulation playing a particularly significant role. However, individual variations in susceptibility to obesity-related comorbidities remain challenging to explain solely based on lifestyle, environmental factors, and genetic variation. Increasing evidence suggests that epigenetic factors, such as DNA methylation, might bridge the gap between genetic predisposition and environmental influences in the development of T2D. Studies have identified altered DNA methylation patterns in obesity and T2D, implicating genes involved in glucose homeostasis. Nevertheless, the complexity of the interplay between DNA methylation and gene expression in this context necessitates further investigation. This research builds upon these previous findings by performing paired DNA methylation and gene expression analyses to gain a deeper understanding of the epigenetic mechanisms underlying T2D in obesity.

The study enrolled 19 obese women (BMI ≥35 kg/m²), including nine without T2D (OND) and ten with T2D (OD). All participants underwent bariatric surgery, and VAT biopsies were collected and stored in RNAlater. DNA and RNA were extracted from the biopsies. DNA methylation analysis was performed using Illumina HumanMethylation EPIC BeadChip Arrays (850 K). Raw data were processed using the ChAMP package in R, including filtering, normalization (beta-mixture quantile normalization), and batch effect correction (ComBat). Differentially methylated CpGs (DMCs) were identified using Limma, with a false discovery rate (FDR) < 0.05. Differentially methylated regions (DMRs) were identified using the Bumphunter algorithm (p-value < 0.05). Gene expression analysis was performed using Affymetrix Clariom S Human Microarrays. Data were preprocessed using the oligo package in R, followed by normalization (robust multi-array average) and batch effect correction (ComBat). Differentially expressed genes (DEGs) were identified using Limma (uncorrected p < 0.05 and logFC > 0.5). The correlation between DMCs/DMRs and gene expression, as well as their correlation with T2D-related traits (fasting glucose and HbA1c), were assessed using Pearson correlation with bootstrap analysis. Gene ontology analysis was performed using WebGestalt to identify enriched pathways. To validate the methylation findings, independent data from 14 Chinese and 7 German women were combined and analyzed.

The study identified 11,120 DMCs and 96 DMRs in the VAT of obese women with T2D compared to those without. The MHC locus showed the highest density of epigenomic alterations. The DMCs were associated with both previously known and novel T2D-related genes. Gene expression analysis revealed 252 DEGs. An overlap between altered methylation and expression was observed in 68 genes, including 24 genes with a significant relationship between gene expression and methylation (p<0.05). Sixteen of these genes, including ATP11A, LPL, and EHD2, also showed significant correlations with fasting glucose and HbA1c levels. Validation analysis using a combined dataset of Chinese and German women showed consistent differential methylation in a subset of CpGs. An extended multi-ethnic analysis (combining all datasets) found 9648 DMCs in 5135 genes, with substantial overlap and consistent directionality in DMCs found in the original analysis. Enrichment analyses revealed pathways involved in fatty acid metabolism, aldosterone synthesis, oxytocin signaling, GABAergic synapse, and dopaminergic synapse.

The study's findings reveal a complex interplay between DNA methylation and gene expression in the pathogenesis of T2D in obesity. The identification of numerous DMCs and DMRs, particularly in genes associated with lipid metabolism and inflammatory processes, supports the role of epigenetic modifications in adipose tissue dysfunction. The significant correlations between DNA methylation levels in specific genes (e.g., ATP11A, LPL, EHD2) and T2D-related traits suggest potential mechanisms through which these epigenetic alterations might contribute to disease development. The validation and extended multi-ethnic analyses further strengthen the findings, although ethnic differences highlight the need for more inclusive studies. The lack of a strong correlation between methylation changes and gene expression in many cases emphasizes the complexities of epigenetic regulation and the potential importance of considering alternative mechanisms. The study's strengths include the paired DNA methylation and gene expression analysis and validation with an extended multi-ethnic analysis. The results contribute to a deeper understanding of the molecular basis of T2D in obesity and potentially support DNA methylation as a biomarker for T2D risk.

This study identified novel candidate genes and pathways involved in T2D pathogenesis in obesity, highlighting the role of DNA methylation alterations in visceral adipose tissue. While the correlation between methylation and expression varied, the identified methylation changes, particularly in genes like ATP11A, LPL, and EHD2, showed significant associations with T2D-related traits. These results warrant further investigation into the functional impact of these epigenetic alterations and their potential as biomarkers for T2D risk prediction in obese individuals. Future research could focus on functional studies to validate the causal role of these epigenetic modifications, explore sex-specific differences, and investigate the influence of other epigenetic mechanisms. Moreover, larger, more diverse cohorts are needed to further establish the generalizability of these findings across different ethnic populations.

The study's limitations include its relatively small sample size and focus on a female cohort, limiting generalizability to males. The cross-sectional nature of the study prevents the determination of causality between methylation alterations and T2D development. The analysis relied primarily on correlative findings, requiring future functional studies to establish direct causal relationships. The reliance on existing databases for the extended analysis could introduce some bias due to variations in methodologies used in other studies.