Differentially Methylated Positions Research Articles

Epigenome-wide association study of lung cancer among never smokers in two prospective cohorts in Shanghai, China

BackgroundThe aetiology of lung cancer among individuals who never smoked remains elusive, despite 15% of lung cancer cases in men and 53% in women worldwide being unrelated to smoking. Epigenetic...

Immune cell distribution and DNA methylation signatures differ between tumor and stroma enriched compartment in pancreatic ductal adenocarcinoma

The presence of abundant tumor stroma is a prominent characteristic of pancreatic ductal adenocarcinomas (PDAC) that potentially influences disease progression and therapy response. This study aims to investigate immune cell infiltration and epigenetic profiles in tumor cell enriched (“Tumor”) and stroma cell enriched (“Stroma”) regions within human PDAC tissue samples. By comparing those regions, we identified 25,410 differentially methylated positions (DMPs) distributed across 6,963 unique genes. Pathway enrichment analysis using the top 2,000 DMPs that were either hyper- or hypomethylated indicated that immune response pathways and the estrogen receptor pathway are epigenetically dysregulated in Tumor and Stroma regions, respectively. In terms of immune cell infiltration, we observed overall low levels of T cells in both regions. In Tumor regions however, occurrence of tumor-associated macrophages (TAMs) was higher than in Stroma regions (p=0.02) concomitant with a dualistic distribution that stratifies PDAC patients into those with high and low TAM infiltration. By categorizing TAM levels into quartiles, our analysis revealed that PDAC patients with more than 1,515 TAMs per mm² exhibited significantly shorter overall survival (p=0.036). Our data suggest that variations in inflammatory characteristics between the Tumor and Stroma defined compartments of PDAC may primarily stem from the presence of macrophages rather than lymphocytes. The abundance of TAMs within regions enriched with tumor cells correlates with patient survival, underscoring the potential significance of exploring therapeutic interventions targeting TAMs. Furthermore, directing attention towards the estrogen receptor pathway may represent a promising strategy to address the stroma cell component within the PDAC tumor microenvironment.

Serum Cobalt Concentration and DNA Methylation Signatures in Women with Obesity

Obesity, a multifactorial disorder, has been associated with alterations in metal metabolismand epigenetic modifications. This pilot case–control study aimed to investigate serum cobalt concentrations and associated DNA methylation patterns in women with obesity. Serum cobalt levels were measured using inductively coupled plasma mass spectrometry (ICP-MS), revealing significantly higher cobalt concentrations in participants with normal weight than in participants with obesity. Additionally, DNA methylation analysis identified differentially methylated positions (DMPs) associated with cobalt exposure, and DMPs between groups highlighted hypomethylation in the top DMPs in individuals with obesity. Functional enrichment analysis of these DMPs unveiled potential pathways implicated in apoptosis, cancer, and metabolic signaling, warranting further investigation into the mechanistic links. This study provides preliminary insights into the interplay between cobalt exposure, DNA methylation, and potential implications for obesity management.

Lithium response in bipolar disorder: Epigenome-wide DNA methylation signatures and epigenetic aging

Lithium (Li) is the first-line treatment for bipolar disorder (BD) even though only 30 % of BD patients are considered excellent responders. The mechanisms by which Li exerts its action are not clearly understood, but it has been suggested that specific epigenetic mechanisms, such as methylation processes, may play a role. In this regard, DNA methylation patterns can be used to estimate epigenetic age (EpiAge), which is accelerated in BD patients and reversed by Li treatment. Our first aim was to compare the DNA methylation profile in peripheral blood between BD patients categorized as excellent responders to Li (Ex-Rp) and non-responders (N-Rp). Secondly, EpiAge was estimated to detect differential age acceleration between the two groups.A total of 130 differentially methylated positions (DMPs) and 16 differentially methylated regions (DMRs) between Ex-Rp (n = 26) and N-Rp (n = 37) were identified (FDR adjusted p-value < 0.05). We found 122 genes mapping the DMPs and DMRs, nine of which (HOXB6, HOXB3, HOXB-AS3, TENM2, CACNA1B, ANK3, EEF2K, CYP1A1, and SORCS2) had previously been linked to Li response. We found genes related to the GSK3β pathway to be highly represented. Using FUMA, we found enrichment in Gene Ontology Cell Component for the synapse. Gene network analysis highlighted functions related to the cell cycle, nervous system development and function, and gene expression. No significant differences in age acceleration were found between Ex-Rp and N-Rp for any of the epigenetic clocks analysed.Our findings indicate that a specific methylation pattern could determine the response to Li in BD patients. We also found that a significant portion of the differentially methylated genes are closely associated with the GSK3β pathway, reinforcing the role of this system in Li response. Future longitudinal studies with larger samples will help to elucidate the epigenetic mechanisms underlying Li response.

Blood DNA Methylation Analysis Reveals a Distinctive Epigenetic Signature of Vasospasm in Aneurysmal Subarachnoid Hemorrhage.

Vasospasm is a potentially preventable cause of poor prognosis in patients with aneurysmal subarachnoid hemorrhage (aSAH). Epigenetics might provide insight on its molecular mechanisms. We aimed to analyze the association between differential DNA methylation (DNAm) and development of vasospasm. We conducted an epigenome-wide association study in 282 patients with aSAH admitted to our hospital. DNAm was assessed with the EPIC Illumina chip (> 850K CpG sites) in whole-blood samples collected at hospital admission. We identified differentially methylated positions (DMPs) at the CpG level using Cox regression models adjusted for potential confounders, and then we used the DMP results to find differentially methylated regions (DMRs) and enriched biological pathways. A total of 145 patients (51%) experienced vasospasm. In the DMP analysis, we identified 31 CpGs associated with vasospasm at p-value < 10-5. One of them (cg26189827) was significant at the genome-wide level (p-value < 10-8), being hypermethylated in patients with vasospasm and annotated to SUGCT gene, mainly expressed in arteries. Region analysis revealed 13 DMRs, some of them annotated to interesting genes such as POU5F1, HLA-DPA1, RUFY1, and CYP1A1. Functional enrichment analysis showed the involvement of biological processes related to immunity, inflammatory response, oxidative stress, endothelial nitric oxide, and apoptosis. Our findings show, for the first time, a distinctive epigenetic signature of vasospasm in aSAH, establishing novel links with essential biological pathways, including inflammation, immune responses, and oxidative stress. Although further validation is required, our results provide a foundation for future research into the complex pathophysiology of vasospasm.

Methylation-driven mechanisms of allergic rhinitis during pollen and non-pollen seasons using integrated bioinformatics analysis.

Allergic rhinitis (AR) is a widespread allergic airway disease that results from a complex interplay between genetic and environmental factors and affects approximately 10%-40% of the global population. Pollen is a common allergen, and exposure to pollen can cause epigenetic changes. However, the mechanism underlying pollen-induced DNA methylation changes and their potential effects on the allergic march are still unclear. The purpose of this study was to explore the methylation-driven mechanisms of AR during the pollen and non-pollen seasons using bioinformatics analysis and to investigate their relationship with asthma. We downloaded DNA methylation and gene expression data from the GEO database (GSE50387: GSE50222, GSE50101) and identified differentially methylated positions (DMPs) and differentially expressed genes (DEGs) during the pollen and non-pollen seasons using the CHAMP and limma packages. Through correlation analysis, we identified methylation-driven genes and performed pathway enrichment analysis to annotate their functions. We incorporated external data on AR combined with asthma (GSE101720) for analysis to identify key CpGs that promote the transformation of AR to asthma. We also utilized external data on olive pollen allergy (GSE54522) for analysis to validate the methylation-driven genes. Weighted correlation network analysis (WGCNA) was employed to identify gene modules significantly correlated with pollen allergy. We extracted genes related to the key methylation-driven gene ZNF667-AS1 from the significant module and performed pathway intelligent clustering using KOBAS-i. We also utilized gene set enrichment analysis to explore the potential function of ZNF667-AS1. We identified 20 and 24 CpG-Gene pairings during the pollen and non-pollen seasons. After incorporating external data from GSE101720, we found that ZNF667-AS1 is a key gene that may facilitate the transformation of AR into asthma during the pollen season. This finding was further validated in another external dataset, GSE54522, which is associated with pollen allergy. WGCNA identified 17 modules, among which the blue module showed significant correlation with allergies. ZNF667-AS1 was located in the blue module. We performed pathway analysis on the genes correlated with ZNF667-AS1 extracted from the blue module and identified a prominent cluster of pathways in the KOBAS-i results, including Toll-like receptor (TLR) family, MyD88, MAPK, and oxidative stress. Gene set enrichment analysis around cg05508084 (paired with ZNF667-AS1) also indicated its potential involvement in initiating and modulating allergic inflammation from the perspective of TLR and MAPK signaling. We identified methylation-driven genes and their related pathways during the pollen and non-pollen seasons in patients with AR and identified key CpGs that promote the transformation of AR into asthma due to pollen exposure. This study provides new insights into the underlying molecular mechanisms of the transformation of AR to asthma.

Conserved methylation signatures associate with the tumor immune microenvironment and immunotherapy response

BackgroundAberrant DNA methylation is a major characteristic of cancer genomes. It remains unclear which biological processes determine epigenetic reprogramming and how these processes influence the variants in the cancer methylome, which can further impact cancer phenotypes.MethodsWe performed pairwise permutations of 381,900 loci in 569 paired DNA methylation profiles of cancer tissue and matched normal tissue from The Cancer Genome Atlas (TCGA) and defined conserved differentially methylated positions (DMPs) based on the resulting null distribution. Then, we derived independent methylation signatures from 2,465 cancer-only methylation profiles from the TCGA and 241 cell line-based methylation profiles from the Genomics of Drug Sensitivity in Cancer (GDSC) cohort using nonnegative matrix factorization (NMF). We correlated DNA methylation signatures with various clinical and biological features, including age, survival, cancer stage, tumor immune microenvironment factors, and immunotherapy response. We inferred the determinant genes of these methylation signatures by integrating genomic and transcriptomic data and evaluated the impact of these signatures on cancer phenotypes in independent bulk and single-cell RNA/methylome cohorts.ResultsWe identified 7,364 differentially methylated positions (2,969 Hyper-DMPs and 4,395 Hypo-DMPs) in nine cancer types from the TCGA. We subsequently retrieved three highly conserved, independent methylation signatures (Hyper-MS1, Hypo-MS1, and Hypo-MS4) from cancer tissues and cell lines based on these Hyper and Hypo-DMPs. Our data suggested that Hypo-MS4 activity predicts poor survival and is associated with immunotherapy response and distant tumor metastasis, and Hypo-MS4 activity is related to TP53 mutation and FOXA1 binding specificity. In addition, we demonstrated a correlation between the activities of Hypo-MS4 in cancer cells and the fractions of regulatory CD4 + T cells with the expression levels of immunological genes in the tumor immune microenvironment.ConclusionsOur findings demonstrated that the methylation signatures of distinct biological processes are associated with immune activity in the cancer microenvironment and predict immunotherapy response.

Green space exposure and blood DNA methylation at birth and in childhood – A multi-cohort study

Green space exposure has been associated with improved mental, physical and general health. However, the underlying biological mechanisms remain largely unknown. The aim of this study was to investigate the association between green space exposure and cord and child blood DNA methylation.Data from eight European birth cohorts with a total of 2,988 newborns and 1,849 children were used. Two indicators of residential green space exposure were assessed: (i) surrounding greenness (satellite-based Normalized Difference Vegetation Index (NDVI) in buffers of 100 m and 300 m) and (ii) proximity to green space (having a green space ≥ 5,000 m2 within a distance of 300 m). For these indicators we assessed two exposure windows: (i) pregnancy, and (ii) the period from pregnancy to child blood DNA methylation assessment, named as cumulative exposure. DNA methylation was measured with the Illumina 450 K or EPIC arrays. To identify differentially methylated positions (DMPs) we fitted robust linear regression models between pregnancy green space exposure and cord blood DNA methylation and between cumulative green space exposure and child blood DNA methylation. Two sensitivity analyses were conducted: (i) without adjusting for cellular composition, and (ii) adjusting for air pollution. Cohort results were combined through fixed-effect inverse variance weighted meta-analyses. Differentially methylated regions (DMRs) were identified from meta-analysed results using the Enmix-combp and DMRcate methods.There was no statistical evidence of pregnancy or cumulative exposures associating with any DMP (False Discovery Rate, FDR, p-value < 0.05). However, surrounding greenness exposure was inversely associated with four DMRs (three in cord blood and one in child blood) annotated to ADAMTS2, KCNQ1DN, SLC6A12 and SDK1 genes. Results did not change substantially in the sensitivity analyses.Overall, we found little evidence of the association between green space exposure and blood DNA methylation. Although we identified associations between surrounding greenness exposure with four DMRs, these findings require replication.

Abstract 7011: The DNA methylation landscape across the TCR loci genes in patients with acute myeloid leukemia

Abstract The T-cell receptor (TCR) repertoire is partly shaped by epigenetic modifications, including DNA methylation. Hypomethylating agents like decitabine are used to treat patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). The precise DNA methylation patterns within the TCR loci and their dynamics associated with HMAs treatments remain underexplored. Here, we analyzed methylation patterns across TCR alpha and beta gene loci in T cells from GSE175758 and GSE67170 datasets, comparing patterns between patients with AML and healthy controls. We also investigated the impact of decitabine on the methylation patterns of TCR gene loci in patients with AML and MDS (GSE175758 and GSE80762).The overall levels of methylation between T cells from healthy donors and patients with AML were significantly different within the TRAV (0.55 vs 0.42, P &amp;lt; 0.001) and TRBV regions (0.70 vs 0.59, P &amp;lt; 0.001) with 46 TRAV and 33 TRBV sites differentially methylated positions, respectively. However, methylation patterns in HSCs (GSE63409), T cells from healthy donors and patients with AML and those in AML cells were strongly correlated (R &amp;gt; 0.67, range 0.67-0.99, P &amp;lt; 0.001).Because decitabine preferentially acts on leukemic cells and given the high correlation levels between AML cells and T cells across datasets we analyzed the effects of decitabine on TCR DNA methylation patterns in AML cells. In the leukemic cells decitabine treatment led to significant demethylation across 33 and 30 CpG sites within the TRAV and TRBV genes, respectively. In the pre-treatment AML samples higher methylation beta values were observed in differentially methylated positions (DMPs) compared with non-DMPs (TRAV: 0.847 vs 0.407; P &amp;lt; 0.001; TRBV: 0.771 vs 0.538; P = 0.049). In samples largely enriched of myeloid leukemic blasts, we found that TRAV and TRBV loci methylation patterns differ significantly between AML patients with good, moderate, and poor risk patients (P &amp;lt; 0.001). We found TRBV6-1 position 1 and 2 methylation was lower in patients with good risk, compared with intermediate risk (pos1: 0.07 vs 0.13, P=0.04; pos2: 0.1 vs 0.2, P = 0.002) and poor risk (pos 1: 0.07 vs 0.21, P &amp;lt; 0.001; pos 2: 0.1 vs 0.32, P &amp;lt; 0.001) and was lower in patients with intermediate risk compared with poor risk (pos1: 0.13 vs 0.21, p=0.007; pos 2: 0.21 vs 0.32, P = 0.001). The high correlation of TCR DNA methylation patterns between the T cells and the myeloid cells, suggests the need for similar analyses in the context of AML T cells. The study highlights the presence of a conserved TCR loci methylated signatures which is highly correlated between myeloid leukemic cells and T cells and is associated with clinical outcomes, offering a potential avenue for therapeutic intervention. Hypomethylating agents, such as decitabine, can modulate TCR loci methylation patterns. Citation Format: Mateusz Pospiech, John Beckford, Advaith M. Kumar, Mukund Tamizharasan, Jaqueline Brito, Gangning Liang, Serghei Mangul, Houda Alachkar. The DNA methylation landscape across the TCR loci genes in patients with acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7011.

Associations of environmental cadmium exposure with kidney damage: Exploring mediating DNA methylation sites in Chinese adults

Environmental exposure is widely recognized as the primary sources of Cadmium (Cd) in the human body, and exposure to Cd is associated with kidney damage in adults. Nevertheless, the role of DNA methylation in Cd-induced kidney damage remains unclear. This study aimed to investigate the epigenome-wide association of environmental Cd-related DNA methylation changes with kidney damage. We included 300 non-smoking adults from the China in 2019. DNA methylation profiles were measured with Illumina Infinium MethylationEPIC BeadChip array. Linear mixed-effect model was employed to estimate the effects of urinary Cd with DNA methylation. Differentially methylated positions (DMPs) associated with urinary Cd were then tested for the association with kidney damage indicators. The mediation analysis was further applied to explore the potential DNA methylation based mediators. The prediction model was developed using a logistic regression model, and used 1000 bootstrap resampling for the internal validation. We identified 27 Cd-related DMPs mapped to 20 genes after the adjustment of false-discovery-rate for multiple testing among non-smoking adults. 17 DMPs were found to be associated with both urinary Cd and kidney damage, and 14 of these DMPs were newly identified within the Chinese. Mediation analysis revealed that DNA methylation of cg26907612 and cg16848624 mediated the Cd-related reduced kidney damage. In addition, ten variables were selected using the LASSO regression analysis and were utilized to develop the prediction model. It found that the nomogram model predicted the risk of kidney damage caused by environmental Cd with a corrected C-index of 0.779. Our findings revealed novel DMPs associated with both environmental Cd exposure and kidney damage among non-smoking adults, and developed an easy-to-use nomogram-illustrated model using these novel DMPs. These findings could provide a theoretical basis for formulating prevention and control strategies for kidney damage from the perspective of environmental pollution and epigenetic regulation.

Epigenome-Wide Association Studies of COPD and Lung Function: A Systematic Review.

Chronic Obstructive Pulmonary Disease (COPD) results from gene-environment interactions over the lifetime. These interactions are captured by epigenetic changes, such as DNA methylation. This systematic review synthesizes evidence from epigenome-wide association studies (EWAS) related to COPD and lung function. Systematic literature search on PubMed, Embase and CINAHL databases, identified 1947 articles that investigated epigenetic changes associated with COPD/lung function; 17 of them met our eligibility criteria from which data was manually extracted. Differentially methylated positions (DMPs) and/or annotated genes, were considered replicated if identified by ≥2 studies with a p<1 x 10-4. Ten studies profiled DNA methylation changes in blood and 7 in respiratory samples, including surgically resected lung tissue (n=3), small airways epithelial brushings (n=2), bronchoalveolar lavage (n=1) and sputum (n=1). Main results showed: (1) high variability in study design, covariates and effect sizes, which prevented a formal meta-analysis; (2) in blood samples, 51 DMPs were replicated in relation to lung function and 12 related to COPD; (3) in respiratory samples, 42 DMPs were replicated in relation to COPD but none in relation to lung function; and, (4) in COPD vs. control studies, 123 genes (2.6% of total) were shared between ≥1 blood and ≥1 respiratory sample and associated with chronic inflammation, ion transport and coagulation. There is high heterogeneity across published COPD/lung function EWAS studies. A few genes (n=123; 2.6%) were replicated in blood and respiratory samples, suggesting that blood can recapitulate some changes in respiratory tissues. These findings have implications for future research.

Epigenome-wide association study of dietary fatty acid intake

BackgroundDietary intake of n-3 polyunsaturated fatty acids (PUFA) may have a protective effect on the development of cardiovascular diseases, diabetes, depression and cancer, while a high intake of n-6 PUFA was often reported to be associated with inflammation-related traits. The effect of PUFAs on health outcomes might be mediated by DNA methylation (DNAm). The aim of our study is to identify the impact of PUFA intake on DNAm in the Cooperative Health Research in the Region of Augsburg (KORA) FF4 cohort and the Leiden Longevity Study (LLS).ResultsDNA methylation levels were measured in whole blood from the population-based KORA FF4 study (N = 1354) and LLS (N = 448), using the Illumina MethylationEPIC BeadChip and Illumina HumanMethylation450 array, respectively. We assessed associations between DNAm and intake of eight and four PUFAs in KORA and LLS, respectively. Where possible, results were meta-analyzed.Below the Bonferroni correction threshold (p < 7.17 × 10–8), we identified two differentially methylated positions (DMPs) associated with PUFA intake in the KORA study. The DMP cg19937480, annotated to gene PRDX1, was positively associated with docosahexaenoic acid (DHA) in model 1 (beta: 2.00 × 10–5, 95%CI: 1.28 × 10–5-2.73 × 10–5, P value: 6.98 × 10–8), while cg05041783, annotated to gene MARK2, was positively associated with docosapentaenoic acid (DPA) in our fully adjusted model (beta: 9.80 × 10–5, 95%CI: 6.25 × 10–5-1.33 × 10–4, P value: 6.75 × 10–8). In the meta-analysis, we identified the CpG site (cg15951061), annotated to gene CDCA7L below Bonferroni correction (1.23 × 10–7) associated with eicosapentaenoic acid (EPA) intake in model 1 (beta: 2.00 × 10–5, 95% CI: 1.27 × 10–5–2.73 × 10–5, P value = 5.99 × 10–8) and we confirmed the association of cg19937480 with DHA in both models 1 and 2 (beta: 2.07 × 10–5, 95% CI: 1.31 × 10–5–2.83 × 10–5, P value = 1.00 × 10–7 and beta: 2.19 × 10–5, 95% CI: 1.41 × 10–5–2.97 × 10–5, P value = 5.91 × 10–8 respectively).ConclusionsOur study identified three CpG sites associated with PUFA intake. The mechanisms of these sites remain largely unexplored, highlighting the novelty of our findings. Further research is essential to understand the links between CpG site methylation and PUFA outcomes.

Blood epigenome-wide association studies of suicide attempt in adults with bipolar disorder

Suicide attempt (SA) risk is elevated in individuals with bipolar disorder (BD), and DNA methylation patterns may serve as possible biomarkers of SA. We conducted epigenome-wide association studies (EWAS) of blood DNA methylation associated with BD and SA. DNA methylation was measured at >700,000 positions in a discovery cohort of n = 84 adults with BD with a history of SA (BD/SA), n = 79 adults with BD without history of SA (BD/non-SA), and n = 76 non-psychiatric controls (CON). EWAS revealed six differentially methylated positions (DMPs) and seven differentially methylated regions (DMRs) between BD/SA and BD/non-SA, with multiple immune-related genes implicated. There were no epigenome-wide significant differences when BD/SA and BD/non-SA were each compared to CON, and patterns suggested that epigenetics differentiating BD/SA from BD/non-SA do not differentiate BD/non-SA from CON. Weighted gene co-methylation network analysis and trait enrichment analysis of the BD/SA vs. BD/non-SA contrast further corroborated immune system involvement, while gene ontology analysis implicated calcium signalling. In an independent replication cohort of n = 48 BD/SA and n = 47 BD/non-SA, fold changes at the discovery cohort’s significant sites showed moderate correlation across cohorts and agreement on direction. In both cohorts, classification accuracy for SA history among individuals with BD was highest when methylation at the significant CpG sites as well as information from clinical interviews were combined, with an AUC of 88.8% (CI = 83.8–93.8%) and 82.1% (CI = 73.6–90.5%) for the combined epigenetic-clinical classifier in the discovery and replication cohorts, respectively. Our results provide novel insight to the role of immune system functioning in SA and BD and also suggest that integrating information from multiple levels of analysis holds promise to improve risk assessment for SA in adults with BD.

Integrated DNA methylation analysis reveals a potential role for PTPRN2 in Marfan syndrome scoliosis.

Marfan syndrome (MFS) is a rare genetic disorder caused by mutations in the Fibrillin-1 gene (FBN1) with significant clinical features in the skeletal, cardiopulmonary, and ocular systems. To gain deeper insights into the contribution of epigenetics in the variability of phenotypes observed in MFS, we undertook the first analysis of integrating DNA methylation and gene expression profiles in whole blood from MFS and healthy controls (HCs). The Illumina 850K (EPIC) DNA methylation array was used to detect DNA methylation changes on peripheral blood samples of seven patients with MFS and five HCs. Associations between methylation levels and clinical features of MFS were analyzed. Subsequently, we conducted an integrated analysis of the outcomes of the transcriptome data to analyze the correlation between differentially methylated positions (DMPs) and differentially expressed genes (DEGs) and explore the potential role of methylation-regulated DEGs (MeDEGs) in MFS scoliosis. The weighted gene co-expression network analysis was used to find gene modules with the highest correlation coefficient with target MeDEGs to annotate their functions in MFS. Our study identified 1253 DMPs annotated to 236 genes that were primarily associated with scoliosis, cardiomyopathy, and vital capacity. These conditions are typically associated with reduced lifespan in untreated MFS. We calculated correlations between DMPs and clinical features, such as cobb angle to evaluate scoliosis and FEV1% to assess pulmonary function. Notably, cg20223687 (PTPRN2) exhibited a positive correlation with cobb angle of scoliosis, potentially playing a role in ERKs inactivation. Taken together, our systems-level approach sheds light on the contribution of epigenetics to MFS and offers a plausible explanation for the complex phenotypes that are linked to reduced lifespan in untreated MFS patients.

P1193 NOD2 and Beyond: Unmasking Tobacco-Induced Epigenetic Changes in Crohn's Disease

Abstract Background Smoking has been shown to have a very negative impact on Crohn's disease (CD). Indeed, this harmful habit induces the early onset of the disease and doubles the risk of postoperative recurrence. This research project involves a thorough analysis of omics data, with a primary focus on DNA methylation, one of the most prevalent epigenetic modifications. Our hypothesis is that tobacco-induced alterations in gene methylation pattern of subcutaneous adipose tissue (SAT), particularly within key pathways, contribute to a more adverse disease profile in smokers. Methods A comprehensive epigenome-wide analysis was conducted using the Illumina EPIC/450k array to investigate subcutaneous adipose tissue in patients with CD, encompassing both smokers and non-smokers. Utilizing tools such as dmpfinder and Bumphunter (minfi), differentially methylated positions (DMPs) and regions (DMRs) were identified. Additionally, gene expression analysis for key genes was performed to establish correlations between methylation and subsequent gene expression, not only in SAT but also in other fat depots such as visceral adipose tissue (VAT) or peripheral blood mononuclear cells (PBMCs). Results A distinctive DNA methylation pattern in the SAT of smokers compared to non-smoker CD patients was identified (Fig. 1A). Over 27,270 DMPs associated with significant biological processes related to DNA damage, inflammation, or immunity were uncovered (Fig. 1B-C). Additionally, more than 13,000 DMRs located in the promoter region were identified (Fig. 1C). The relationship between methylation levels in the promoter regions and subsequent gene expression on SAT was validated for selected genes. Specifically, NOD2 exhibited higher DNA methylation in the promoter region among smokers compared to non-smokers CD patients (p value=0.04), resulting in reduced NOD2 gene expression on SAT (p value=0.04) (Fig.1D). Consistent results were observed in other tissues such as VAT and PBMCs (Fig. 1E). Finally, we observe the implication of NOD2 in pathways related to immune response, inflammation or apoptosis and a trend indicating that lower NOD2 expression is associated with higher levels of C-reactive protein (CRP)(Fig. 1F). Conclusion Our study reveals that smoking produces DNA methylation changes on SAT of CD patients and affects the expression of several key genes involved in the pathogeny of CD. Particularly, our results unveil a new link between tobacco-smoking and lower NOD2 expression and associate this to an elevated levels of PCR, paving the way for a fresh perspective in exploring disease prognosis.

Alterations in leukocyte DNA methylome are associated to immunosuppression in severe clinical phenotypes of septic patients.

Sepsis patients experience a complex interplay of host pro- and anti-inflammatory processes which compromise the clinical outcome. Despite considering the latest clinical and scientific research, our comprehension of the immunosuppressive events in septic episodes remains incomplete. Additionally, a lack of data exists regarding the role of epigenetics in modulating immunosuppression, subsequently impacting patient survival. To advance the current understanding of the mechanisms underlying immunosuppression, in this study we explored the dynamics of DNA methylation using the Infinium Methylation EPIC v1.0 BeadChip Kit in leukocytes from patients suffering from sepsis, septic shock, and critically ill patients as controls, within the first 24 h after admission in the Intensive Care Unit of a tertiary hospital. Employing two distinct analysis approaches (DMRcate and mCSEA) in comparing septic shock and critically ill patients, we identified 1,256 differentially methylated regions (DMRs) intricately linked to critical immune system pathways. The examination of the top 100 differentially methylated positions (DMPs) between septic shock and critically ill patients facilitated a clear demarcation among the three patient groups. Notably, the top 6,657 DMPs exhibited associations with organ dysfunction and lactate levels. Among the individual genes displaying significant differential methylation, IL10, TREM1, IL1B, and TNFAIP8 emerged with the most pronounced methylation alterations across the diverse patient groups when subjected to DNA bisulfite pyrosequencing analysis. These findings underscore the dynamic nature of DNA methylation profiles, highlighting the most pronounced alterations in patients with septic shock, and revealing their close association with the disease.

Genome-wide DNA methylation risk scores for schizophrenia derived from blood and brain tissues further explain the genetic risk in patients stratified by polygenic risk scores for schizophrenia and bipolar disorder

BackgroundGenetic and environmental factors contribute to the pathogenesis of schizophrenia (SZ) and bipolar disorder (BD). Among genetic risk groups stratified by combinations of Polygenic Risk Score (PRS) deciles for SZ,...

Methylation and transcriptomic expression profiles of HUVEC in the oxygen and glucose deprivation model and its clinical implications in AMI patients.

The obstructed coronary artery undergoes a series of pathological changes due to ischemic-hypoxic shocks during acute myocardial infarction (AMI). However, the altered DNA methylation levels in endothelial cells under these conditions and their implication for the etiopathology of AMI have not been investigated in detail. This study aimed to explore the relationship between DNA methylation and pathologically altered gene expression profile in human umbilical vein endothelial cells (HUVECs) subjected to oxygen-glucose deprivation (OGD), and its clinical implications in AMI patients. The Illumina Infinium MethylationEPIC BeadChip assay was used to explore the genome-wide DNA methylation profile using the Novaseq6000 platform for mRNA sequencing in 3 pairs of HUVEC-OGD and control samples. GO and KEGG pathway enrichment analyses, as well as correlation, causal inference test (CIT), and protein-protein interaction (PPI) analyses identified 22 hub genes that were validated by MethylTarget sequencing as well as qRT-PCR. ELISA was used to detect four target molecules associated with the progression of AMI. A total of 2,524 differentially expressed genes (DEGs) and 22,148 differentially methylated positions (DMPs) corresponding to 6,642 differentially methylated genes (DMGs) were screened (|Δβ|>0.1 and detection p < 0.05). After GO, KEGG, correlation, CIT, and PPI analyses, 441 genes were filtered. qRT-PCR confirmed the overexpression of VEGFA, CCL2, TSP-1, SQSTM1, BCL2L11, and TIMP3 genes, and downregulation of MYC, CD44, BDNF, GNAQ, RUNX1, ETS1, NGFR, MME, SEMA6A, GNAI1, IFIT1, and MEIS1. DNA fragments BDNF_1_ (r = 0.931, p < 0.0001) and SQSTM1_2_NEW (r = 0.758, p = 0.0043) were positively correlated with the expressions of corresponding genes, and MYC_1_ (r = -0.8245, p = 0.001) was negatively correlated. Furthermore, ELISA confirmed TNFSF10 and BDNF were elevated in the peripheral blood of AMI patients (p = 0.0284 and p = 0.0142, respectively). Combined sequencing from in vitro cellular assays with clinical samples, aiming to establish the potential causal chain of the causal factor (DNA methylation) - mediator (mRNA)-cell outcome (endothelial cell ischemic-hypoxic injury)-clinical outcome (AMI), our study identified promising OGD-specific genes, which provided a solid basis for screening fundamental diagnostic and prognostic biomarkers of coronary endothelial cell injury of AMI. Moreover, it furnished the first evidence that during ischemia and hypoxia, the expression of BNDF was regulated by DNA methylation in endothelial cells and elevated in peripheral blood.

Targeted bisulfite sequencing analysis of Alzheimer’s disease candidate genes reveals differential methylation across neurofibrillary tangle burden in the prefrontal cortex.

AbstractBackgroundRecent epigenome‐wide association studies (EWAS) identified several loci in specific genes showing robust and reproducible alterations in DNA methylation in Alzheimer’s disease (AD) brain samples. Standardly, assessing methylation in EWAS is done using microarrays, which target a limited number of sites in each gene. Here, we performed targeted bisulfite sequencing of candidate genes associated with AD to determine the exact extent of methylation changes across neurofibrillary tangle burden (NFT) within these loci.MethodPrefrontal cortex brain samples from 58 individuals were grouped by Braak stage (control 0‐II; intermediate III‐IV; AD V‐VI). Following DNA extraction, 30 genomic regions were captured using Agilent SureSelect target baits. After next‐generation bisulfite sequencing, reads were aligned and the methylation status of cytosine residues called using the Bismark program. Differentially methylated positions (DMPs) were analyzed across the three groups. Furthermore, the presence of differentially methylated regions (DMRs), made up of several DMPs was assessed. Methylation levels in genomic features, such as promoters and gene bodies, of the target regions were also compared.ResultMethylation levels were quantified for each group. Linear regression controlled for co‐variation before differences were examined using a one‐way ANOVA, with Tukey’s post‐hoc test. Sites in several genes showed stepwise increases in DNA methylation across the groups. Interestingly, amongst the most robust sites, differences in methylation in the intermediate group when compared to the control and AD samples were observed. DMRs were observed in the groups, as were methylation differences between promotors and gene bodies in several targeted regions.ConclusionThis work provides further evidence that dysregulation of methylation is associated with pathological changes in AD prefrontal cortex. Differential methylation with intermediate NFT pathology, at sites showing no difference between control and AD samples, potentially indicates early pathological changes. As DNA methylation is reversible, this could present potential targets for pharmacological intervention.

Amyloid‐beta immunotherapy induces novel methylomic changes in Alzheimer’s disease brain

AbstractBackgroundPost‐mortem neuropathological examinations following the first active immunotherapy strategy (AN‐1792, Elan Pharmaceuticals, 2000) for Alzheimer’s disease (AD) have evidenced amyloid‐β (Aβ) plaque clearance and increased microglial phagocytic activity in immunised individuals. This study characterises the epigenetic profiles of individuals who underwent Aβ immunotherapy with the aim of discovering novel therapeutic targets and biomarkers.MethodDNA was isolated from post‐mortem prefrontal cortex tissue of 16 immunised AD cases who received varying doses (ug) and number of doses during the trial period, including placebos. DNA methylation was quantified using methylation arrays and the raw intensity values processed and normalised for subsequent statistical analysis to identify differentially methylated positions (DMPs) across the genome associated with Aβ immunotherapy. Additional analyses included regions of enrichment analysis for identification of differentially methylated regions (DMRs) and weighted gene co‐expression network analysis to identify gene clusters with highly correlated methylation levels.ResultAfter correcting for known (age, sex, cell type composition) and unknown variables, a DMP located within Chromosome 19 Open Reading Frame 12 (C19Orf12) at the genome‐wide significance level (P &lt; 9.00E−08) was associated with Aβ immunisation. 10 DMRs were further associated with immunisation, with three regions consisting of ≥ 3 CpG sites and having a Sidak‐corrected P &lt; 0.05. These regions were located within the CRISP2 (chr6; 49681178:49681391 [8 probes], Sidak‐corrected P = 8.29E‐05), CTTNA1 (chr5; 138210906:138211184 [8 probes], Sidak‐corrected P = 5.96E‐06), and RNF39 (chr6; 30038859:30039025 [12 probes], Sidak‐Corrected P = 2.60E‐05) genes.ConclusionIndicating better inflammatory markers can help inform of effective preventative care strategies for elders. This study provides evidence for altered epigenetic processes in the pathophysiology of AD and identifies novel processes specific to Aβ immunotherapy. The next step for this project includes analysing RNA sequencing data, to determine if observed methylomic changes correlate with gene expression, and genotyping array data.