Differentially Methylated Positions Research Articles

Genome-wide integrated DNA methylome and transcriptome analysis reveals a strong dysregulated myeloid component in the epigenetic landscape of Systemic Sclerosis.

Non-genetic factors influence Systemic Sclerosis (SSc) pathogenesis, underscoring epigenetics as a relevant contributor to the disease. We aimed to unravel DNA methylation abnormalities associated with SSc through an epigenome-wide association study (EWAS). We analyzed DNA methylation data from whole blood samples in 179 SSc patients and 241 unaffected individuals, to identify differentially methylated positions (DMPs) with a FDR<0.05. These results were further integrated with RNA-seq data from the same patients to assess their functional consequence. Additionally, we examined the impact of DNA methylation changes on transcription factors and analyzed the relationship between alterations of the methylation and gene expression profile and serum proteins levels. This analysis yielded 525 DMPs enriched in immune-related pathways, being leukocyte cell-cell adhesion the most significant (FDR=4.91x10-9), prioritizing integrins as they were exposed by integrating methylome and transcriptome data. Furthermore, through this integrative approach we observed an enrichment of neutrophil related pathways, highlighting this myeloid cell type as a relevant contributor in SSc pathogenesis. In addition, we uncovered novel profibrotic and proinflammatory mechanisms involved in the disease. Finally, the altered epigenetic and transcriptomic signature revealed an increased activity of CCAAT/enhancer-binding protein (CEBP) transcription factor family in SSc, which is crucial in the myeloid lineage development. Our findings uncover the impaired epigenetic regulation of the disease and its impact on gene expression, identifying new molecules for potential clinical applications and improving our understanding of SSc pathogenesis.

DNA methylation stability in cardiac tissues kept at different temperatures and time intervals

Investigating DNA methylation (DNAm) in cardiac tissues is vital for epigenetic research in cardiovascular diseases (CVDs). During cardiac surgery, biopsies may not be immediately stored due to a lack of human or technical resources at the collection site. Assessing DNAm stability in cardiac samples left in suboptimal conditions is crucial for applying DNAm analysis. We investigated the stability of DNAm in human cardiac tissues kept at 4 °C and 22 °C for periods of 1, 7, 14, and 28 days (exposed samples) using the Illumina Infinium MethylationEPIC v1.0 BeadChip Array. We observed high correlations between samples analysed immediately after tissue collection and exposed ones (R2 > 0.992). Methylation levels were measured as β-values and median absolute β-value differences (|∆β|) ranged from 0.0093 to 0.0119 in all exposed samples. Pairwise differentially methylated position (DMP) analysis revealed no DMPs under 4 °C (fridge temperature) exposure for up to 28 days and 22 °C (room temperature) exposure for one day, while 3,437, 6,918, and 3,824 DMPs were observed for 22 °C samples at 7, 14, and 28 days, respectively. This study provides insights into the stability of genome-wide DNAm, showing that cardiac tissue can be used for reliable DNAm analysis even when stored suboptimally after surgery.

A comprehensive lifestyle index and its associations with DNA methylation and type 2 diabetes among Ghanaian adults: the rodam study

BackgroundA series of modifiable lifestyle factors, such as diet quality, physical activity, alcohol intake, and smoking, may drive the rising burden of type 2 diabetes (T2DM) among sub-Saharan Africans globally. It is unclear whether epigenetic changes play a mediatory role in the associations between these lifestyle factors and T2DM. We assessed the associations between a comprehensive lifestyle index, DNA methylation and T2DM among Ghanaian adults.MethodsWe used whole-blood Illumina 450 k DNA methylation data from 713 Ghanaians from the Research on Obesity and Diabetes among African Migrants (RODAM) study. We constructed a comprehensive lifestyle index based on established cut-offs for diet quality, physical activity, alcohol intake, and smoking status. In the T2DM-free discovery cohort (n = 457), linear models were fitted to identify differentially methylated positions (DMPs) and differentially methylated regions (DMRs) associated with the lifestyle index after adjustment for age, sex, body mass index (BMI), and technical covariates. Associations between the identified DMPs and the primary outcome (T2DM), as well as secondary outcomes (fasting blood glucose (FBG) and HbA1c), were determined via logistic and linear regression models, respectively.ResultsIn the present study population (mean age: 52 ± 10 years; male: 42.6%), the comprehensive lifestyle index showed a significant association with one DMP annotated to an intergenic region on chromosome 7 (false discovery rate (FDR) = 0.024). Others were annotated to ADCY7, SMARCE1, AHRR, LOXL2, and PTBP1 genes. One DMR was identified and annotated to the GFPT2 gene (familywise error rate (FWER) from bumphunter bootstrap = 0.036). None of the DMPs showed significant associations with T2DM; directions of effect were positive for the DMP in the AHRR and inverse for all the other DMPs. Higher methylation of the ADCY7 DMP was associated with higher FBG (p = 0.024); LOXL2 DMP was associated with lower FBG (p = 0.023) and HbA1c (p = 0.049); and PTBP1 DMP was associated with lower HbA1c (p = 0.002).ConclusionsIn this explorative epigenome-wide association study among Ghanaians, we identified one DMP and DMR associated with a comprehensive lifestyle index not previously associated with individual lifestyle factors. Based on our findings, we infer that lifestyle factors in combination, affect DNA methylation, thereby influencing the risk of T2DM among Ghanaian adults living in different contexts.

A new insight of blood vs. buccal DNA methylation in the forensic identification of monozygotic triplets

The case of the monozygotic (MZ) twin as a suspect demonstrates a practical problem in forensic casework. As the MZ twins are genetically identical, they share the same short tandem repeat (STR) profile. Many studies showed that older MZ twins have significant differences in overall content and genomic distribution of methylation between them. However, studies addressing the investigation of epigenetic MZ triplet differentiation in various forensic reference materials are lacking. Here, one triplet set of Egyptian MZ twins was used as an analog to a forensic case. The genome-wide methylation analysis was performed via the new Human Methylation EPIC BeadChip array. Following normalization methods, potential differentially methylated positions (DMPs) were discovered. This resulted in the detection of 24 potential DMPs in reference-type blood DNA and 11 potential DMPs in reference-type buccal DNA. Then, the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were performed to show the associated biological functions. Our findings revealed that the 35 potential DMPs were enriched in 283 significant GO terms. These terms are mainly enriched in the immune system. Overall, this study demonstrates the general feasibility of epigenetic MZ triplet differentiation in the forensic context and highlights that some potential DMPs identified in blood DNA were not informative in buccal DNA. This is due to various reasons, including the tissue specificity of DNA methylation.

Longitudinal changes in DNA methylation associated with clozapine use in treatment-resistant schizophrenia from two international cohorts

The second-generation antipsychotic clozapine is used as a medication for treatment-resistant schizophrenia. It has previously been associated with epigenetic changes in pre-clinical rodent models and cross-sectional studies of treatment-resistant schizophrenia. Cross-sectional studies are susceptible to confounding, however, and cannot disentangle the effects of diagnosis and medication. We therefore profiled DNA methylation in sequential blood samples (n = 126) from two independent cohorts of patients (n = 38) with treatment-resistant schizophrenia spectrum disorders who commenced clozapine after study enrolment and were followed up for up to six months. We identified significant non-linear changes in cell-type proportion estimates derived from DNA methylation data - specifically B-cells - associated with time on clozapine. Mixed effects regression models were used to identify changes in DNA methylation at specific sites associated with time on clozapine, identifying 37 differentially methylated positions (DMPs) (p < 5 × 10-5) in a linear model and 90 DMPs in a non-linear quadratic model. We compared these results to data from our previous epigenome-wide association study (EWAS) meta-analysis of psychosis, finding evidence that many previously identified DMPs associated with schizophrenia and treatment-resistant schizophrenia might reflect exposure to clozapine. In conclusion, our results indicate that clozapine exposure is associated with changes in DNA methylation and cellular composition. Our study shows that medication effects might confound many case-control studies of neuropsychiatric disorders performed in blood.

Whole genome methylation sequencing in blood from persons with mild cognitive impairment and dementia due to Alzheimer's disease identifies cognitive status.

Whole genome methylation sequencing (WGMS) in blood identifies differential DNA methylation in persons with late-onset dementia due to Alzheimer's disease (AD) but has not been tested in persons with mild cognitive impairment (MCI). We used WGMS to compare DNA methylation levels at 25,244,219 CpG loci in 382 blood samples from 99 persons with MCI, 109 with AD, and 174 who are cognitively unimpaired (CU). WGMS identified 9,756 differentially methylated positions (DMPs) in persons with MCI, including 1,743 differentially methylated genes encoding proteins in biological pathways related to synapse organization, dendrite development, and ion transport. 447 DMPs exhibit progressively increasing or decreasing DNA methylation levels between CU, MCI, and AD that correspond to cognitive status. WGMS identifies DMPs in known and newly detected genes in blood from persons with MCI and AD that support blood DNA methylation levels as candidate biomarkers of cognitive status.

Multicohort Epigenome-Wide Association Study of All-Cause Cardiovascular Disease and Cancer Incidence: A Cardio-Oncology Approach

BackgroundEmerging evidence reveals a complex relationship between cardiovascular disease (CVD) and cancer, which share common risk factors and biological pathways. ObjectivesThe aim of this study was to evaluate common epigenetic signatures for CVD and cancer incidence in 3 ethnically diverse cohorts: Native Americans from the SHS (Strong Heart Study), European Americans from the FHS (Framingham Heart Study), and European Americans and African Americans from the ARIC (Atherosclerosis Risk In Communities) study. MethodsA 2-stage strategy was used that included first conducting untargeted epigenome-wide association studies for each cohort and then running targeted models in the union set of identified differentially methylated positions (DMPs). We also explored potential molecular pathways by conducting a bioinformatics analysis. ResultsCommon DMPs were identified across all populations. In a subsequent meta-analysis, 3 and 1 of those DMPs were statistically significant for CVD only and both cancer and CVD, respectively. No meta-analyzed DMPs were statistically significant for cancer only. The enrichment analysis pointed to interconnected biological pathways involved in cancer and CVD. In the DrugBank database, elements related to 1-carbon metabolism and cancer and CVD medications were identified as potential drugs for target gene products. In an additional analysis restricted to the 950 SHS participants who developed incident CVD, the C index for incident cancer increased from 0.618 (95% CI: 0.570-0.672) to 0.971 (95% CI: 0.963-0.978) when adjusting the models for the combined cancer and CVD DMPs identified in the other cohorts. ConclusionsThese results point to molecular pathways and potential treatments for precision prevention of CVD and cancer. Screening based on common epigenetic signatures of incident CVD and cancer may help identify patients with newly diagnosed CVD at increased cancer risk.

A comprehensive map of the aging blood methylome in humans

BackgroundDuring aging, the human methylome undergoes both differential and variable shifts, accompanied by increased entropy. The distinction between variably methylated positions (VMPs) and differentially methylated positions (DMPs), their contribution to epigenetic age, and the role of cell type heterogeneity remain unclear.ResultsWe conduct a comprehensive analysis of > 32,000 human blood methylomes from 56 datasets (age range = 6–101 years). We find a significant proportion of the blood methylome that is differentially methylated with age (48% DMPs; FDR < 0.005) and variably methylated with age (37% VMPs; FDR < 0.005), with considerable overlap between the two groups (59% of DMPs are VMPs). Bivalent and Polycomb regions become increasingly methylated and divergent between individuals, while quiescent regions lose methylation more uniformly. Both chronological and biological clocks, but not pace-of-aging clocks, show a strong enrichment for CpGs undergoing both mean and variance changes during aging. The accumulation of DMPs shifting towards a methylation fraction of 50% drives the increase in entropy, smoothening the epigenetic landscape. However, approximately a quarter of DMPs exhibit anti-entropic effects, opposing this direction of change. While changes in cell type composition minimally affect DMPs, VMPs and entropy measurements are moderately sensitive to such alterations.ConclusionThis study represents the largest investigation to date of genome-wide DNA methylation changes and aging in a single tissue, providing valuable insights into primary molecular changes relevant to chronological and biological aging.

A genome-wide methylation analysis of Chinese Han patients with chronic insomnia disorder.

As the most common sleep disorder, chronic insomnia disorder (CID) has become a global health burden to the public. However, it remains unclear about the pathogenesis of this disease. Epigenetic changes may provide important insights into the gene-environment interaction in CID. Therefore, this study was conducted to investigate the DNA methylation pattern in CID and reveal the epigenetic mechanism of this disease. In this study, whole blood DNA was extracted from 8 CID patients (the CID group) and 8 healthy controls (the control group), respectively. Besides, genome-wide DNA methylation was detected by Illumina Human Methylation 850K Beadchip. Moreover, the sleep quality and insomnia severity were evaluated by the Pittsburgh Sleep Quality Index (PSQI) and Insomnia Severity Index (ISI), respectively. A total of 369 differentially methylated positions (DMPs) and 23 differentially methylated regions (DMRs) were identified between the CID and control groups. LHX6 was identified as the most important differentially methylated gene (DMG). The Gene Ontology (GO) analysis results corroborated that DMPs were significantly enriched in 105 GO terms, including cell signaling, homogenous cell adhesion of plasma membrane adhesion molecules, nervous system development, cell adhesion, and calcium ion binding. In addition, it was demonstrated that DMPs were significantly enriched in the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, including the hippo signaling pathway, Ras signaling pathway, and vitamin B6 metabolism. The DMR-related GO analysis results revealed the positive regulation of protein kinase activities. DNA methylation plays a critical role in the development of CID, and LHX6 is validated to be an important DMG.

An epigenome-wide study of selenium status and DNA methylation in the Strong Heart Study

BackgroundSelenium (Se) is an essential nutrient linked to adverse health endpoints at low and high levels. The mechanisms behind these relationships remain unclear and there is a need to further understand the epigenetic impacts of Se and their relationship to disease. We investigated the association between urinary Se levels and DNA methylation (DNAm) in the Strong Heart Study (SHS), a prospective study of cardiovascular disease (CVD) among American Indians adults. MethodsSelenium concentrations were measured in urine (collected in 1989–1991) using inductively coupled plasma mass spectrometry among 1,357 participants free of CVD and diabetes. DNAm in whole blood was measured cross-sectionally using the Illumina MethylationEPIC BeadChip (850 K) Array. We used epigenome-wide robust linear regressions and elastic net to identify differentially methylated cytosine-guanine dinucleotide (CpG) sites associated with urinary Se levels. ResultsThe mean (standard deviation) urinary Se concentration was 51.8 (25.1) μg/g creatinine. Across 788,368 CpG sites, five differentially methylated positions (DMP) (hypermethylated: cg00163554, cg18212762, cg11270656, and hypomethylated: cg25194720, cg00886293) were significantly associated with Se in linear regressions after accounting for multiple comparisons (false discovery rate p-value: 0.10). The top hypermethylated DMP (cg00163554) was annotated to the Disco Interacting Protein 2 Homolog C (DIP2C) gene, which relates to transcription factor binding. Elastic net models selected 425 hypo- and hyper-methylated DMPs associated with urinary Se, including three sites (cg00163554 [DIP2C], cg18212762 [MAP4K2], cg11270656 [GPIHBP1]) identified in linear regressions. ConclusionsUrinary Se was associated with minimal changes in DNAm in adults from American Indian communities across the Southwest and the Great Plains in the United States, suggesting that other mechanisms may be driving health impacts. Future analyses should explore other mechanistic biomarkers in human populations, determine these relationships prospectively, and investigate the potential role of differentially methylated sites with disease endpoints.

Folate-mediated transgenerational inheritance of sperm DNA methylation patterns correlate with spinal axon regeneration

ABSTRACT In mammals, the molecular mechanisms underlying transgenerational inheritance of phenotypic traits in serial generations of progeny after ancestral environmental exposures, without variation in DNA sequence, remain elusive. We’ve recently described transmission of a beneficial trait in rats and mice, in which F0 supplementation of methyl donors, including folic acid, generates enhanced axon regeneration after sharp spinal cord injury in untreated F1 to F3 progeny linked to differential DNA methylation levels in spinal cord tissue. To test whether the transgenerational effect of folic acid is transmitted via the germline, we performed whole-genome methylation sequencing on sperm DNA from F0 mice treated with either folic acid or vehicle control, and their F1, F2, and F3 untreated progeny. Transgenerational differentially methylated regions (DMRs) are observed in each consecutive generation and distinguish folic acid from untreated lineages, predominate outside of CpG islands and in regions of the genome that regulate gene expression, including promoters, and overlap at both the differentially methylated position (DMP) and gene levels. These findings indicate that molecular changes between generations are caused by ancestral folate supplementation. In addition, 29,719 DMPs exhibit serial increases or decreases in DNA methylation levels in successive generations of untreated offspring, correlating with a serial increase in the phenotype across generations, consistent with a ‘wash-in’ effect. Sibship-specific DMPs annotate to genes that participate in axon- and synapse-related pathways.

Blood DNA methylation in post-acute sequelae of COVID-19 (PASC): a prospective cohort study

DNA methylation integrates environmental signals with transcriptional programs. COVID-19 infection induces changes in the host methylome. While post-acute sequelae of COVID-19 (PASC) is a long-term complication of acute illness, its association with DNA methylation is unknown. No universal blood marker of PASC, superseding single organ dysfunctions, has yet been identified. In this single centre prospective cohort study, PASC, post-COVID without PASC, and healthy participants were enrolled to investigate their symptoms association with peripheral blood DNA methylation data generated with state-of-the-art whole genome sequencing. PASC-induced quality-of-life deterioration was scored with a validated instrument, SF-36. Analyses were conducted to identify potential functional roles of differentially methylated loci, and machine learning algorithms were used to resolve PASC severity. 103 patients with PASC (22.3% male, 77.7% female), 15 patients with previous COVID-19 infection but no PASC (40.0% male, 60.0% female), and 27 healthy volunteers (48.1% male, 51.9% female) were enrolled. Whole genome methylation sequencing revealed 39 differentially methylated regions (DMRs) specific to PASC, each harbouring an average of 15 consecutive positions, that differentiate patients with PASC from the two control groups. Motif analyses of PASC-regulated DMRs identify binding domains for transcription factors regulating circadian rhythm and others. Some DMRs annotated to protein coding genes were associated with changes of RNA expression. Machine learning support vector algorithm and random forest hierarchical clustering reveal 28unique differentially methylated positions (DMPs) in the genome discriminating patients with better and worse quality of life. Blood DNA methylation levels identify PASC, stratify PASC severity, and suggest that DNA motifs are targeted by circadian rhythm-regulating pathways in PASC. This project has been funded by the following agencies: NIH-AI173035 (A. Jaitovich and R. Alisch); and NIH-AG066179 (R. Alisch).

Transcriptome- and DNA methylation-based cell-type deconvolutions produce similar estimates of differential gene expression and differential methylation

BackgroundChanging cell-type proportions can confound studies of differential gene expression or DNA methylation (DNAm) from peripheral blood mononuclear cells (PBMCs). We examined how cell-type proportions derived from the transcriptome versus the methylome (DNAm) influence estimates of differentially expressed genes (DEGs) and differentially methylated positions (DMPs).MethodsTranscriptome and DNAm data were obtained from PBMC RNA and DNA of Kenyan children (n = 8) before, during, and 6 weeks following uncomplicated malaria. DEGs and DMPs between time points were detected using cell-type adjusted modeling with Cibersortx or IDOL, respectively.ResultsMost major cell types and principal components had moderate to high correlation between the two deconvolution methods (r = 0.60–0.96). Estimates of cell-type proportions and DEGs or DMPs were largely unaffected by the method, with the greatest discrepancy in the estimation of neutrophils.ConclusionVariation in cell-type proportions is captured similarly by both transcriptomic and methylome deconvolution methods for most major cell types.

DNA methylation profile of inflammatory breast cancer and its impact on prognosis and outcome

BackgroundInflammatory breast cancer (IBC) is a rare disease characterized by rapid progression, early metastasis, and a high mortality rate.MethodsGenome-wide DNA methylation analysis (EPIC BeadChip platform, Illumina) and somatic gene variants (105 cancer-related genes) were performed in 24 IBCs selected from a cohort of 140 cases.ResultsWe identified 46,908 DMPs (differentially methylated positions) (66% hypomethylated); CpG islands were predominantly hypermethylated (39.9%). Unsupervised clustering analysis revealed three clusters of DMPs characterized by an enrichment of specific gene mutations and hormone receptor status. The comparison among DNA methylation findings and external datasets (TCGA-BRCA stages III-IV) resulted in 385 shared DMPs mapped in 333 genes (264 hypermethylated). 151 DMPs were associated with 110 genes previously detected as differentially expressed in IBC (GSE45581), and 68 DMPs were negatively correlated with gene expression. We also identified 4369 DMRs (differentially methylated regions) mapped on known genes (2392 hypomethylated). BCAT1, CXCL12, and TBX15 loci were selected and evaluated by bisulfite pyrosequencing in 31 IBC samples. BCAT1 and TBX15 had higher methylation levels in triple-negative compared to non-triple-negative, while CXCL12 had lower methylation levels in triple-negative than non-triple-negative IBC cases. TBX15 methylation level was associated with obesity.ConclusionsOur findings revealed a heterogeneous DNA methylation profile with potentially functional DMPs and DMRs. The DNA methylation data provided valuable insights for prognostic stratification and therapy selection to improve patient outcomes.Graphical

O-306 Sperm DNA methylome changes in testicular cancer patients following chemotherapy treatment

Abstract Study question Does chemotherapy (CT) treatment for testicular cancer induce sperm methylome changes? Summary answer The analysis of 27 pre- and post-CT samples from testicular cancer survivors identified 65 differentially methylated regions. What is known already Research has shown that cancer treatment can cause DNA damage and aneuploidy in sperm, even years after the treatment. However, there is limited information available on the impact of CT on the sperm epigenome, and most of the information has been obtained from rodent models. Two recent studies, one involving one patient and the other involving three, have investigated this topic in humans. Both studies have suggested that cancer survivors may experience changes in sperm’s DNA methylation following oncological treatment. Study design, size, duration This is a multicenter prospective study. We included 27 patients [31.23 years (CI 95% 29.29-33.17)] diagnosed with testicular cancer who cryopreserved semen before CT (2009-2020). After receiving oncological treatment, these patients recovered their spermatogenesis. Patients who agreed to participate in this IRB-approved study, donated a second semen sample after their recovery, which was also frozen. Participants/materials, setting, methods Semen samples were thawed; sperm DNA was isolated and subsequently hybridized using the Human Infinitum DNA Methylation 850K EPIC BeadChip array platform (Illumina). A paired differential methylation analysis was performed between pre-CT and post-CT samples from the same patient, fitting a linear model of the methylated positions and regions. Minfi and DMRcate R packages were used to find differentially methylated positions (DMPs) and regions (DMRs). Subsequently, the functional impact of involved genes was evaluated. Main results and the role of chance DNA methylation data were available for the 54 sperm samples (27 collected before CT and 27 after CT treatment). Results were controlled by age and time elapsed between the cryopreservation of pre-CT and post-CT samples, which was 4.01 years (CI 95%: 3.42-5.64). After filtering for initial QC, the assessment was done on 445.000 CpGs per sample. The analysis revealed that a total of 228 CpG positions showed a tendency towards statistical significance. DMRcate identified 65 DMRs (40 hypomethylated and 25 hypermethylated) that reached a smoothed FDR p-value &amp;lt; 0.05 when compared post-CT samples to pre-CT within the same patient. We observed that 61 genes were located within DMRs, being TBC1D20, EN1, HOXB1 and SCNN1A the ones within the most significant DMRs (min-smoothed-FDR &amp;lt; 0.001). The functional enrichment analysis of putatively affected genes revealed that the biological processes more affected were: Regulation of macromolecule biosynthetic process, hormone receptor binding, signaling pathways regulating pluripotency of stem cells and pathways in cancer. Limitations, reasons for caution Epigenetics may be affected during cryopreservation and thawing. To ensure comparability, we also froze the samples after undergoing the CT process. Due to the high-throughput nature of the study and the limited access to samples, we only achieved marginal significance. Therefore, our results should be further validated. Wider implications of the findings This is the largest study suggesting distinct sperm DNA methylation signatures in testicular cancer survivors. Our findings emphasize the importance of counseling cancer patients in pretreatment sample collection and may explain the reported increase in birth defects after treatment. Trial registration number NOT APPLICABLE

DNA methylation is associated with hair trace elements in female adolescents from two vulnerable populations in the Colombian Caribbean.

Exposure to trace elements (TEs) influences DNA methylation patterns, which may be associated with disease development. Vulnerable populations, such as adolescents undergoing maturity, are susceptible to the effects of TE exposure. The aim of this study was to analyze the association of hair TE concentration with DNA methylation in a sample from female adolescents living in two communities in the Colombian Caribbean coast. Hair and blood samples were obtained from 45 females, between 13 and 16 years of age. Seventeen TEs were quantified in hair samples. DNA methylation was measured in leukocytes using the Infinium MethylationEPIC BeadChip. Linear models were employed to identify differentially methylated positions (DMPs) adjusting for age, body mass index, mother's education, and cell type composition. Among the tested elements, vanadium, chromium, nickel, copper, zinc, yttrium, tin, and barium were significantly associated with DMPs (false discovery rate < 0.05), registering 225, 1, 2, 184, 1, 209 189, and 104 hits, respectively. Most of the DMPs were positively associated with TEs and located in open sea regions. The greatest number of DMPs was annotated to the HOXA3 and FOXO3 genes, related to regulation of gene expression and oxidative stress, respectively. These findings suggest that DNA methylation may be involved in linking exposure to TEs among female adolescents to downstream health risks.

DNA methylation and stroke prognosis: an epigenome-wide association study

Background and aimsStroke is the leading cause of adult-onset disability. Although clinical factors influence stroke outcome, there is a significant variability among individuals that may be attributed to genetics and epigenetics, including DNA methylation (DNAm). We aimed to study the association between DNAm and stroke prognosis.Methods and resultsTo that aim, we conducted a two-phase study (discovery-replication and meta-analysis) in Caucasian patients with ischemic stroke from two independent centers (BasicMar [discovery, N = 316] and St. Pau [replication, N = 92]). Functional outcome was assessed using the modified Rankin Scale (mRS) at three months after stroke, being poor outcome defined as mRS > 2. DNAm was determined using the 450K and EPIC BeadChips in whole-blood samples collected within the first 24 h. We searched for differentially methylated positions (DMPs) in 370,344 CpGs, and candidates below p-value < 10–5 were subsequently tested in the replication cohort. We then meta-analyzed DMP results from both cohorts and used them to identify differentially methylated regions (DMRs).After doing the epigenome-wide association study, we found 29 DMPs at p-value < 10–5 and one of them was replicated: cg24391982, annotated to thrombospondin-2 (THBS2) gene (p-valuediscovery = 1.54·10–6; p-valuereplication = 9.17·10–4; p-valuemeta-analysis = 6.39·10–9). Besides, four DMRs were identified in patients with poor outcome annotated to zinc finger protein 57 homolog (ZFP57), Arachidonate 12-Lipoxygenase 12S Type (ALOX12), ABI Family Member 3 (ABI3) and Allantoicase (ALLC) genes (p-value < 1·10–9 in all cases).DiscussionPatients with poor outcome showed a DMP at THBS2 and four DMRs annotated to ZFP57, ALOX12, ABI3 and ALLC genes. This suggests an association between stroke outcome and DNAm, which may help identify new stroke recovery mechanisms.

DNA methylation signatures provide novel diagnostic biomarkers and predict responses of immune therapy for breast cancer.

Breast cancer (BRCA) is one of the most common malignant tumors affecting women worldwide. DNA methylation modifications can influence oncogenic pathways and provide potential diagnostic and therapeutic targets for precision oncology. In this study, we used non-parametric permutation tests to identify differentially methylated positions (DMPs) between paired tumor and normal BRCA tissue samples from the Cancer Genome Atlas (TCGA) database. Then, we applied non-negative matrix factorization (NMF) to the DMPs to derive eight distinct DNA methylation signatures. Among them, signatures Hyper-S3 and Hypo-S4 signatures were associated with later tumor stages, while Hyper-S1 and Hypo-S3 exhibited higher methylation levels in earlier stages. Signature Hyper-S3 displayed an effect on overall survival. We further validated the four stage-associated signatures using an independent BRCA DNA methylation dataset from peripheral blood samples. Results demonstrated that 24 commonly hypomethylated sites in Hypo-S4 showed lower methylation in BRCA patients compared to healthy individuals, suggesting its potential as an early diagnostic biomarker. Furthermore, we found that methylation of 23 probes from four stage-related signatures exhibited predictive power for immune therapy response. Notably, methylation levels of all three probes from the Hypo-S4 and activity of the Hypo-S4 demonstrated highly positive relevance to PD-L1 gene expression, implying their significant predictive values for immunotherapy outcomes. GO and KEGG pathway enrichment analysis revealed that genes with these 23 immunotherapy-related methylation probes are mainly involved in glycan degradation or protein deglycosylation. These methylation signatures and probes may serve as novel epigenetic biomarkers for predicting tumor immunotherapy response. Our findings provide new insights into precision oncology approaches for BRCA.

Smoking-Induced DNA Hydroxymethylation Signature Is Less Pronounced than True DNA Methylation: The Population-Based KORA Fit Cohort.

Despite extensive research on 5-methylcytosine (5mC) in relation to smoking, there has been limited exploration into the interaction between smoking and 5-hydroxymethylcytosine (5hmC). In this study, total DNA methylation (5mC+5hmC), true DNA methylation (5mC) and hydroxymethylation (5hmC) levels were profiled utilizing conventional bisulphite (BS) and oxidative bisulphite (oxBS) treatment, measured with the Illumina Infinium Methylation EPIC BeadChip. An epigenome-wide association study (EWAS) of 5mC+5hmC methylation revealed a total of 38,575 differentially methylated positions (DMPs) and 2023 differentially methylated regions (DMRs) associated with current smoking, along with 82 DMPs and 76 DMRs associated with former smoking (FDR-adjusted p < 0.05). Additionally, a focused examination of 5mC identified 33 DMPs linked to current smoking and 1 DMP associated with former smoking (FDR-adjusted p < 0.05). In the 5hmC category, eight DMPs related to current smoking and two DMPs tied to former smoking were identified, each meeting a suggestive threshold (p < 1 × 10-5). The substantial number of recognized DMPs, including 5mC+5hmC (7069/38,575, 2/82), 5mC (0/33, 1/1), and 5hmC (2/8, 0/2), have not been previously reported. Our findings corroborated previously established methylation positions and revealed novel candidates linked to tobacco smoking. Moreover, the identification of hydroxymethylated CpG sites with suggestive links provides avenues for future research.

Genome-Wide DNA Methylation in Early-Onset-Dementia Patients Brain Tissue and Lymphoblastoid Cell Lines.

Epigenetics, a potential underlying pathogenic mechanism of neurodegenerative diseases, has been in the scope of several studies performed so far. However, there is a gap in regard to analyzing different forms of early-onset dementia and the use of Lymphoblastoid cell lines (LCLs). We performed a genome-wide DNA methylation analysis on sixty-four samples (from the prefrontal cortex and LCLs) including those taken from patients with early-onset forms of Alzheimer's disease (AD) and frontotemporal dementia (FTD) and healthy controls. A beta regression model and adjusted p-values were used to obtain differentially methylated positions (DMPs) via pairwise comparisons. A correlation analysis of DMP levels with Clariom D array gene expression data from the same cohort was also performed. The results showed hypermethylation as the most frequent finding in both tissues studied in the patient groups. Biological significance analysis revealed common pathways altered in AD and FTD patients, affecting neuron development, metabolism, signal transduction, and immune system pathways. These alterations were also found in LCL samples, suggesting the epigenetic changes might not be limited to the central nervous system. In the brain, CpG methylation presented an inverse correlation with gene expression, while in LCLs, we observed mainly a positive correlation. This study enhances our understanding of the biological pathways that are associated with neurodegeneration, describes differential methylation patterns, and suggests LCLs are a potential cell model for studying neurodegenerative diseases in earlier clinical phases than brain tissue.