Genome-wide Deoxyribonucleic Acid Methylation Research Articles

Study of association of global deoxyribonucleic acid methylation in women with polycystic ovary syndrome

Polycystic ovary syndrome (PCOS), a common endocrine disorder affecting 5%-10% of reproductive age women worldwide, associated with various metabolic morbidities. One potential molecular mechanism could be epigenetic modifications, such as deoxyribonucleic acid (DNA) methylation. The aim is to determine the association of global DNA methylation in peripheral blood leucocyte (PBL) cells and PCOS women. Also to assess abnormal lipid profile, insulin resistance, gonadotropins and reproductive markers in them. The study design involves a hospital-based prospective case-control study. Fifty women with PCOS, diagnosed as per Rotterdam criteria and the rest 50 without PCOS or any disease, attending outpatient department were recruited. Serum biochemical markers and Global DNA methylation assay were done by using standardised kit. Data were compared using Independent t-test or Mann-Whitney U test using IBM SPSS version 26.0. P < 0.05 was considered statistically significant. Majority, 72% of PCOS and 82% non-PCOS women were between 20 and 25 years. Most common presenting symptom was menstrual irregularity. Women with PCOS have high serum cholesterol and triglyceride level, elevated serum luteinising hormone (LH), follicle-stimulating hormone (FSH), LH/FSH ratio and testosterone but low estradiol levels as compared to non-PCOS. Statistically significant high mean Global DNA methylation percentage was found in PBLs of women with PCOS. Despite study limitations, this study provided insight into Global DNA methylation in PBLs was associated with PCOS. It requires further research to better understand the influence of epigenetic factors including genome-wide DNA methylation profiling in PCOS development.

Differences of DNA methylation patterns in the placenta of large for gestational age infant

To investigate the molecular mechanisms of later metabolic health changes in large for gestational age (LGA) newborns by analyzing deoxyribonucleic acid (DNA) methylation patterns in the placenta of LGA and appropriate for gestational age (AGA) newborns.A total of 6 placentas of LGA and 6 placentas of AGA newborns were enrolled as LGA group and AGA group. DNA methylation was analyzed using the Illumina Infinium Human MethylationEPIC BeadChip microarrays and verified via pyrosequencing and reverse transcription-quantitative real-time polymerase chain reaction. Functional enrichment analysis were constructed by gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis based on the differentially methylated regions between LGA and AGA groups.Clinical investigation showed that LGA newborns had significantly lower hemoglobin and blood glucose compared to AGA newborns. Birth weight was negatively correlated to hemoglobin and blood glucose. Genome-wide DNA methylation analysis identified 17 244 methylation variable positions achieving genome-wide significance (adjusted P < .05). 34% methylation variable positions were located in the gene promoter region. A total of 117 differentially methylated regions were revealed by bump hunting analysis, which mapped to 107 genes. Function analysis showed 13 genes enriched in adhesion and infection process, endocrine and other factor-regulated calcium reabsorption, calcium signaling pathway and transmembrane transport. Four genes linked to type II diabetes mellitus. Among the 13 genes, we selected GNAS and calcium voltage-gated channel subunit alpha1 G for independent verification of pyrosequencing, and the messenger ribonucleic acid levels of guanine nucleotide binding protein, calcium voltage-gated channel subunit alpha1 G, DECR1, and FK506 binding protein 11 were verified by reverse transcription-quantitative real-time polymerase chain reaction.DNA methylation variation and gene expression differences in placental samples were associated with LGA newborns, which linking the effect of intrauterine environment to regulation of the offspring's gene expression. Furthermore, pathway analysis suggested that intrauterine environment affecting fetal growth might had a functional impact on multiple signaling pathways involved in fetal growth, metabolism, and inflammation. Further studies were required to understand the differences of methylation patterns.

Deoxyribonucleic acid methylation signatures in sperm deoxyribonucleic acid fragmentation

To evaluate Deoxyribonucleic acid (DNA) methylation patterns in sperm from men with differential levels of sperm DNA fragmentation index (DFI). Prospective study. University-affiliated reproductive medicine center. A total of 278 male patients consulting for couple infertility were recruited from the First Affiliated Hospital of Wenzhou Medical University. None. Genome-wide DNA methylation analysis was performed using Infinium MethylationEPIC BeadChip on spermatozoal DNA from 20 male patients. Differentially methylated regions (DMRs) were identified and validated using targeted bisulfite amplicon sequencing in spermatozoal DNA from 266 males. Unsupervised hierarchical clustering analysis revealed three main clusters corresponding to sperm DFI levels (low, medium, or high). Between-cluster comparisons identified 959 (medium-low), 738 (high-medium), and 937 (high-low) DMRs. Sixty-six DMRs were validated in the 266-sample cohort, of which nine CpG fragments corresponding to nine genes (BLCAP, DIRAS3, FAM50B, GNAS, MEST, TSPAN32, PSMA8, SYCP1, and TEX12) exhibited significantly altered methylation in those with high DFI (≥25%) compared with those with low DFI (<25%). We identified and validated a distinct DNA methylation signature associated with sperm DNA damage in a large, unselected cohort. These results indicate that sperm DNA damage may affect DNA methylation patterns in human sperm.

Differences of DNA methylation patterns in the placenta of large for gestational age infant

To investigate the molecular mechanisms of later metabolic health changes in large for gestational age (LGA) newborns by analyzing deoxyribonucleic acid (DNA) methylation patterns in the placenta of LGA and appropriate for gestational age (AGA) newborns.A total of 6 placentas of LGA and 6 placentas of AGA newborns were enrolled as LGA group and AGA group. DNA methylation was analyzed using the Illumina Infinium Human MethylationEPIC BeadChip microarrays and verified via pyrosequencing and reverse transcription-quantitative real-time polymerase chain reaction. Functional enrichment analysis were constructed by gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis based on the differentially methylated regions between LGA and AGA groups.Clinical investigation showed that LGA newborns had significantly lower hemoglobin and blood glucose compared to AGA newborns. Birth weight was negatively correlated to hemoglobin and blood glucose. Genome-wide DNA methylation analysis identified 17 244 methylation variable positions achieving genome-wide significance (adjusted P < .05). 34% methylation variable positions were located in the gene promoter region. A total of 117 differentially methylated regions were revealed by bump hunting analysis, which mapped to 107 genes. Function analysis showed 13 genes enriched in "adhesion and infection process, endocrine and other factor-regulated calcium reabsorption, calcium signaling pathway and transmembrane transport". Four genes linked to type II diabetes mellitus. Among the 13 genes, we selected GNAS and calcium voltage-gated channel subunit alpha1 G for independent verification of pyrosequencing, and the messenger ribonucleic acid levels of guanine nucleotide binding protein, calcium voltage-gated channel subunit alpha1 G, DECR1, and FK506 binding protein 11 were verified by reverse transcription-quantitative real-time polymerase chain reaction.DNA methylation variation and gene expression differences in placental samples were associated with LGA newborns, which linking the effect of intrauterine environment to regulation of the offspring's gene expression. Furthermore, pathway analysis suggested that intrauterine environment affecting fetal growth might had a functional impact on multiple signaling pathways involved in fetal growth, metabolism, and inflammation. Further studies were required to understand the differences of methylation patterns.

In utero Bisphenol A Exposure Is Linked with Sex Specific Changes in the Transcriptome and Methylome of Human Amniocytes.

Prenatal exposure to bisphenol A (BPA) is linked to obesity and diabetes but the molecular mechanisms driving these phenomena are not known. Alterations in deoxyribonucleic acid (DNA) methylation in amniocytes exposed to BPA in utero represent a potential mechanism leading to metabolic dysfunction later in life. To profile changes in genome-wide DNA methylation and expression in second trimester human amniocytes exposed to BPA in utero. A nested case-control study was performed in amniocytes matched for offspring sex, maternal race/ethnicity, maternal age, gestational age at amniocentesis, and gestational age at birth. Cases had amniotic fluid BPA measuring 0.251 to 23.74 ng/mL. Sex-specific genome-wide DNA methylation analysis and RNA-sequencing (RNA-seq) were performed to determine differentially methylated regions (DMRs) and gene expression changes associated with BPA exposure. Ingenuity pathway analysis was performed to identify biologically relevant pathways enriched after BPA exposure. In silico Hi-C analysis identified potential chromatin interactions with DMRs. There were 101 genes with altered expression in male amniocytes exposed to BPA (q < 0.05) in utero, with enrichment of pathways critical to hepatic dysfunction, collagen signaling and adipogenesis. Thirty-six DMRs were identified in male BPA-exposed amniocytes and 14 in female amniocyte analysis (q < 0.05). Hi-C analysis identified interactions between DMRs and 24 genes with expression changes in male amniocytes and 12 in female amniocytes (P < 0.05). In a unique repository of human amniocytes exposed to BPA in utero, sex-specific analyses identified gene expression changes in pathways associated with metabolic disease and novel DMRs with potential distal regulatory functions.

Zebrafish as an In Vivo Model to Assess Epigenetic Effects of Ionizing Radiation.

Exposure to ionizing radiations (IRs) is ubiquitous in our environment and can be categorized into “targeted” effects and “non-targeted” effects. In addition to inducing deoxyribonucleic acid (DNA) damage, IR exposure leads to epigenetic alterations that do not alter DNA sequence. Using an appropriate model to study the biological effects of radiation is crucial to better understand IR responses as well as to develop new strategies to alleviate exposure to IR. Zebrafish, Danio rerio, is a scientific model organism that has yielded scientific advances in several fields and recent studies show the usefulness of this vertebrate model in radiation biology. This review briefly describes both “targeted” and “non-targeted” effects, describes the findings in radiation biology using zebrafish as a model and highlights the potential of zebrafish to assess the epigenetic effects of IR, including DNA methylation, histone modifications and miRNA expression. Other in vivo models are included to compare observations made with zebrafish, or to illustrate the feasibility of in vivo models when the use of zebrafish was unavailable. Finally, tools to study epigenetic modifications in zebrafish, including changes in genome-wide DNA methylation, histone modifications and miRNA expression, are also described in this review.

Aberrant sperm DNA methylation predicts male fertility status and embryo quality

To evaluate whether male fertility status and/or embryo quality during in vitro fertilization (IVF) therapy can be predicted based on genomewide sperm deoxyribonucleic acid (DNA) methylation patterns. Retrospective cohort study. University-based fertility center. Participants were 127 men undergoing IVF treatment (where any major female factor cause of infertility had been ruled out), and 54 normozoospermic, fertile men. The IVF patients were stratified into 2 groups: patients who had generally good embryogenesis and a positive pregnancy (n = 55), and patients with generally poor embryogenesis (n = 72; 42 positive and 30 negative pregnancies) after IVF. Genomewide sperm DNA methylation analysis was performed to measure methylation at >485,000 sites across the genome. A comparison was made of DNA methylation patterns of IVF patients vs. normozoospermic, fertile men. Predictive models proved to be highly accurate in classifying male fertility status (fertile or infertile), with 82% sensitivity, and 99% positive predictive value. Hierarchic clustering identified clusters enriched for IVF patient samples and for poor-quality-embryo samples. Models built to identify samples within these groups, from neat samples, achieved positive predictive value ≥ 94% while identifying >one fifth of all IVF patient and poor-quality-embryo samples in each case. Using density gradient prepared samples, the same approach recovered 46% of poor-quality-embryo samples with no false positives. Sperm DNA methylation patterns differ significantly and consistently for infertile vs. fertile, normozoospermic men. In addition, DNA methylation patterns may be predictive of embryo quality during IVF.

Deoxyribonucleic acid methylation profiling of single human blastocysts by methylated CpG-island amplification coupled with CpG-island microarray

ObjectiveTo study whether methylated CpG-island (CGI) amplification coupled with microarray (MCAM) can be used to generate DNA (deoxyribonucleic acid) methylation profiles from single human blastocysts.DesignA pilot microarray study with methylated CpG-island amplification applied to human blastocyst genomic DNA and hybridized on CpG-island microarrays.SettingUniversity research laboratory.Patient(s)Five cryopreserved sibling 2-pronuclear zygotes that were surplus to requirements for clinical treatment by in vitro fertilization were donated with informed consent from a patient attending Bourn Hall Clinic, Cambridge, United Kingdom.Intervention(s)None.Main Outcome Measure(s)Successful generation of genome-wide DNA methylation profiles at CpG islands from individual human blastocysts, with common genomic regions of DNA methylation identified between embryos.Result(s)Between 472 and 734 CpG islands were methylated in each blastocyst, with 121 CpG islands being commonly methylated in all 5 blastocysts. A further 159 CGIs were commonly methylated in 4 of the 5 tested blastocysts. Methylation was observed at a number of CGIs within imprinted-gene, differentially methylated regions (DMRs), including placental and preimplantation-specific DMRs.Conclusion(s)The MCAM method is capable of providing comprehensive DNA methylation data in individual human blastocysts.

Abstract 1357: Clinical, pathological and prognostic value of LINE-1 methylation in hepatocellular carcinoma

Abstract Objectives:Cancer cells exhibit 2 types of deoxyribonucleic acid (DNA) methylation alterations: genome-wide DNA hypomethylation and site-specific CpG island promoter hypermethylation. Genome-wide DNA hypomethylation plays a role in genomic instability and carcinogenesis. As long interspersed nucleotide element-1 (LINE-1) retrotransposon constitutes a substantial portion (approximately 17%) of the humangenome, the level of LINE-1 methylation is regarded to be a surrogate marker of genome-wide DNA methylation. Importantly, LINE-1 hypomethylation is associated with poor prognosis in several type of cancers, suggesting the potencial role of the LINE-1 methylation level as a useful marker for predicting cancer prognosis. However, clinical, pathological and prognostic significance of LINE-1 hypomethylaiton in hepatocellular carcinoma (HCC) remains unclear. Methods: Using 160 HCC resected specimens, we quantified the LINE-1 methylation by utilizing the bisulfite pyrosequencing technology. This assay amplifies a region of LINE-1 element (position 305-331 in accession no. X58075), which includes 4 CpG cites. The average of the relative amounts of C in the 4 CpG sites was used as the overall LINE-1 methylation level.Results: We first examined the LINE-1 methylation level in 85 HCC tissues and matched normal hepatic parenchyma samples. The cancer tissues exhibited significantly lower levels of LINE-1 methylation (median, 64.5 ; range, 99.1-21.5) than the matched normal hepatic parenchyma (median, 81.0 ; range, 97.2-35.8) (P &amp;lt; 0.0001 by the paired t test). The LINE-1 methylation level of HCC was not associated with any of theclinical, epidemiological, or pathological characteristics, including sex, age, AFP, PIVKA-II, infection of hepatitis B virus and hepatitis C virus, ICG-R15, stage of fibrosis, differentiation, tumor size, tumor type, vascular invasion and stage (all P &amp;gt; 0.09). In the Kaplan-Meier analysis, LINE-1 hypomethylators (methylation level &amp;lt; 74.4) experienced significantly shorter disease-free survival than those with hypermethylators (log-rank P=0.029). Over-all survival is not associated with LINE-1 methylation ( log-rank P=0.16). Conclusions: Genome-wide DNA hypomethylation, as measured in LINE-1, is associated with disease-free survival among patients with HCC, suggesting that LINE-1 hypomethylation may be a biomarker that can be used to identify patients who will experience an inferior outcome. Citation Format: kazuto harada, Yoshirumi Baba, Akira Chikamoto, Takatsugu Ishimoto, Keisuke Kosumi, Asuka Murata, Naoya Yoshida, Toru Beppu, Hideo Baba. Clinical, pathological and prognostic value of LINE-1 methylation in hepatocellular carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1357. doi:10.1158/1538-7445.AM2014-1357