scholarly journals The development and validation of EpiComet-Chip, a modified high-throughput comet assay for the assessment of DNA methylation status

2017 ◽  
Vol 58 (7) ◽  
pp. 508-521 ◽  
Author(s):  
Todd A. Townsend ◽  
Marcus C. Parrish ◽  
Bevin P. Engelward ◽  
Mugimane G. Manjanatha
Keyword(s):  
Dna Methylation ◽  
Comet Assay ◽  
High Throughput ◽  
Methylation Status ◽  
Development And Validation

scholarly journals Using a medium-throughput comet assay to evaluate the global DNA methylation status of single cells

2014 ◽  
Vol 5 ◽  
Author(s):  
Angélique Lewies ◽  
Etresia Van Dyk ◽  
Johannes F. Wentzel ◽  
Pieter J. Pretorius
Keyword(s):  
Dna Methylation ◽  
Comet Assay ◽  
Single Cells ◽  
Methylation Status ◽  
Global Dna Methylation

High Throughput Methylation Arrays Allow for Identification of Complex Methylation Patterns in MDS.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2437-2437
Author(s):  
Ying Jiang ◽  
Christine L. OKeefe ◽  
Andrew Dunbar ◽  
Anjali Advani ◽  
Mikkael A. Sekeres ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Tumor Suppressor ◽  
High Throughput ◽  
Tumor Suppressor Genes ◽  
Methylation Status ◽  
Epigenetic Silencing ◽  
Low Grade ◽  
High Grade ◽  
Methylation Patterns ◽  
Hypermethylated Genes

Abstract Genomic imprinting and epigenetic silencing determine tissue-specific methylation patterns. Altered methylation of CpG islands within gene promoters has been hypothesized as one pathogenetic mechanism operative in myelodysplastic syndrome (MDS). Promoter hypermethylation of various empirically selected tumor suppressor genes has been found in MDS prompting application of hypomethylating drugs in this disease. Identification of hypermethylated genes predicting response to these drugs would have a major impact on clinical practice. However, to date methylation-based prognostic algorithms have not been established. Global analysis of DNA methylation patterns may help to identify hypermethylated genes/promoters associated with the pathogenesis of MDS. Recently, microarray-based DNA methylation analysis platforms enabled a powerful, high-throughput analysis of the methylation status of hundreds of genes. The GoldenGate Methylation Cancer Panel I, spanning 1,536 independent CpG sites selected from 807 selected genes was applied to determine the methylation status in MDS patients (N=51; 21 low grade (RA, MDS-U, RARS or RCMD), 26 high grade (AML or RAEB) and 4 CMML). The methylation status was determined based on an internal reference and compared to healthy controls (N=22). Methylation values were averaged among the patients or analyzed separately for each patient in comparison to average values obtained in controls. Overall, controls showed a lesser degree of methylation than advanced MDS patients (average intensity 0.326 vs. 0.339, p<0.05). Subsequently, we concentrated on hypermethylated genes. There were no genes uniformly hypermethylated in all patients. For 70%, 50%, and 30% of patients with advanced MDS, 1, 26, and 85 loci were concordantly hypermethylated, while in 70%, 50% and 30% of low risk patients 5, 23 and 31 were hypermethylated, respectively. The most consistently hypermethylated genes (>50% of patients), included tumor suppressor genes (DCC, SLC22A18, FAT, TUSC3), genes involved in DNA repair (OGG1, DDB2, BCR, PARP1), cell cycle control (DBC1, SMARCB1), differentiation (MYOD1, TDGF1, FGF2, NOTCH4) and apoptosis (HDAC1, ALOX12, AXIN1). Despite the variability, the aberrant methylation spectrum in CMML, low grade MDS and high grade MDS showed significant overlap (for example FZD9, IL16, EVI2A, MBD2 and BCR), which suggests that these genes may relate to the common tumorigenesis in MDS. Certain genes show specific methylation correlating to the morphologic diagnosis and may serve as diagnostic markers. For example, the promoter of HDAC1 is hypomethylated in 81% of sAML/RAEB1/2 patients but hypermethylated in 81% of low risk cases. To assess the link between epigenetic changes and chromosomal abnormalities, we also investigated methylation pattern of MDS with del5q for selected genes at the 5q locus. Some genes that are involved in apoptosis (WNT1, TNF receptor) and proliferation (MAP3K8, CSF3) were found to be hypermethylated in comparison to controls, suggesting that epigenetic silencing may enhance the effect of haploinsuffciency for some of the genes. In sum, our study, the first application of a high-throughput microarray methylation assay in MDS, demonstrates that complex methylation patterns exist in MDS and may allow for identification for clinically relevant methylation markers.

Assessing the DNA methylation status of single cells with the comet assay

2010 ◽  
Vol 400 (2) ◽  
pp. 190-194 ◽  
Author(s):  
Johannes F. Wentzel ◽  
Chrisna Gouws ◽  
Cristal Huysamen ◽  
Etresia van Dyk ◽  
Gerhard Koekemoer ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Comet Assay ◽  
Single Cells ◽  
Methylation Status

DNA Methylation Status of Mash2 in Lungs of Somatic Cell Cloning Bovines*

2010 ◽  
Vol 37 (9) ◽  
pp. 960-966 ◽  
Author(s):  
Jie CHEN ◽  
Dong-Jie LI ◽  
Cui ZHANG ◽  
Ning LI ◽  
Shi-Jie LI
Keyword(s):  
Dna Methylation ◽  
Somatic Cell ◽  
Methylation Status ◽  
Cell Cloning

Development and Validation of a Prognostic Nomogram for Gastric Cancer Based on DNA Methylation-Driven Genes

2019 ◽  
Author(s):  
Yi Bai ◽  
Chunlian Wei ◽  
Yuxin Zhong ◽  
Junyu Long ◽  
Shan Huang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Dna Methylation ◽  
Development And Validation

scholarly journals Epigenetic Changes During Mechanically Induced Osteogenic Lineage Commitment

2015 ◽  
Vol 137 (2) ◽  
Author(s):  
Julia C. Chen ◽  
Mardonn Chua ◽  
Raymond B. Bellon ◽  
Christopher R. Jacobs
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Shear Stress ◽  
Fluid Shear Stress ◽  
Methylation Status ◽  
Lineage Commitment ◽  
Fluid Shear ◽  
Osteogenic Lineage ◽  
Osteogenic Markers ◽  
Heterochromatin Structure

Osteogenic lineage commitment is often evaluated by analyzing gene expression. However, many genes are transiently expressed during differentiation. The availability of genes for expression is influenced by epigenetic state, which affects the heterochromatin structure. DNA methylation, a form of epigenetic regulation, is stable and heritable. Therefore, analyzing methylation status may be less temporally dependent and more informative for evaluating lineage commitment. Here we analyzed the effect of mechanical stimulation on osteogenic differentiation by applying fluid shear stress for 24 hr to osteocytes and then applying the osteocyte-conditioned medium (CM) to progenitor cells. We analyzed gene expression and changes in DNA methylation after 24 hr of exposure to the CM using quantitative real-time polymerase chain reaction and bisulfite sequencing. With fluid shear stress stimulation, methylation decreased for both adipogenic and osteogenic markers, which typically increases availability of genes for expression. After only 24 hr of exposure to CM, we also observed increases in expression of later osteogenic markers that are typically observed to increase after seven days or more with biochemical induction. However, we observed a decrease or no change in early osteogenic markers and decreases in adipogenic gene expression. Treatment of a demethylating agent produced an increase in all genes. The results indicate that fluid shear stress stimulation rapidly promotes the availability of genes for expression, but also specifically increases gene expression of later osteogenic markers.

scholarly journals DNA methylation differs extensively between strains of the same geographical origin and changes with age in Daphnia magna

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Jack Hearn ◽  
Fiona Plenderleith ◽  
Tom J. Little
Keyword(s):  
Dna Methylation ◽  
Caloric Restriction ◽  
Daphnia Magna ◽  
Single Gene ◽  
Methylation Status ◽  
Strain Differences ◽  
Life History Trait ◽  
Food Level ◽  
Epigenetic Clock ◽  
Age Related

Abstract Background Patterns of methylation influence lifespan, but methylation and lifespan may also depend on diet, or differ between genotypes. Prior to this study, interactions between diet and genotype have not been explored together to determine their influence on methylation. The invertebrate Daphnia magna is an excellent choice for testing the epigenetic response to the environment: parthenogenetic offspring are identical to their siblings (making for powerful genetic comparisons), they are relatively short lived and have well-characterised inter-strain life-history trait differences. We performed a survival analysis in response to caloric restriction and then undertook a 47-replicate experiment testing the DNA methylation response to ageing and caloric restriction of two strains of D. magna. Results Methylated cytosines (CpGs) were most prevalent in exons two to five of gene bodies. One strain exhibited a significantly increased lifespan in response to caloric restriction, but there was no effect of food-level CpG methylation status. Inter-strain differences dominated the methylation experiment with over 15,000 differently methylated CpGs. One gene, Me31b, was hypermethylated extensively in one strain and is a key regulator of embryonic expression. Sixty-one CpGs were differentially methylated between young and old individuals, including multiple CpGs within the histone H3 gene, which were hypermethylated in old individuals. Across all age-related CpGs, we identified a set that are highly correlated with chronological age. Conclusions Methylated cytosines are concentrated in early exons of gene sequences indicative of a directed, non-random, process despite the low overall DNA methylation percentage in this species. We identify no effect of caloric restriction on DNA methylation, contrary to our previous results, and established impacts of caloric restriction on phenotype and gene expression. We propose our approach here is more robust in invertebrates given genome-wide CpG distributions. For both strain and ageing, a single gene emerges as differentially methylated that for each factor could have widespread phenotypic effects. Our data showed the potential for an epigenetic clock at a subset of age positions, which is exciting but requires confirmation.

scholarly journals DNA methylome signatures of prenatal exposure to synthetic glucocorticoids in hippocampus and peripheral whole blood of female guinea pigs in early life

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Aya Sasaki ◽  
Margaret E. Eng ◽  
Abigail H. Lee ◽  
Alisa Kostaki ◽  
Stephen G. Matthews
Keyword(s):  
Dna Methylation ◽  
Whole Blood ◽  
Guinea Pigs ◽  
Critical Role ◽  
Methylation Status ◽  
Peripheral Tissues ◽  
Epigenetic Biomarkers ◽  
Differentially Methylated Genes ◽  
Increased Risk ◽  
Synthetic Glucocorticoids

AbstractSynthetic glucocorticoids (sGC) are administered to women at risk of preterm delivery, approximately 10% of all pregnancies. In animal models, offspring exposed to elevated glucocorticoids, either by administration of sGC or endogenous glucocorticoids as a result of maternal stress, show increased risk of developing behavioral, endocrine, and metabolic dysregulation. DNA methylation may play a critical role in long-lasting programming of gene regulation underlying these phenotypes. However, peripheral tissues such as blood are often the only accessible source of DNA for epigenetic analyses in humans. Here, we examined the hypothesis that prenatal sGC administration alters DNA methylation signatures in guinea pig offspring hippocampus and whole blood. We compared these signatures across the two tissue types to assess epigenetic biomarkers of common molecular pathways affected by sGC exposure. Guinea pigs were treated with sGC or saline in late gestation. Genome-wide modifications of DNA methylation were analyzed at single nucleotide resolution using reduced representation bisulfite sequencing in juvenile female offspring. Results indicate that there are tissue-specific as well as common methylation signatures of prenatal sGC exposure. Over 90% of the common methylation signatures associated with sGC exposure showed the same directionality of change in methylation. Among differentially methylated genes, 134 were modified in both hippocampus and blood, of which 61 showed methylation changes at identical CpG sites. Gene pathway analyses indicated that prenatal sGC exposure alters the methylation status of gene clusters involved in brain development. These data indicate concordance across tissues of epigenetic programming in response to alterations in glucocorticoid signaling.

scholarly journals DNA Methylation Based Molecular Subtypes Predict Prognosis in Breast Cancer Patients

2021 ◽  
Vol 28 ◽  
pp. 107327482098851
Author(s):  
Zeng-Hong Wu ◽  
Yun Tang ◽  
Yan Zhou
Keyword(s):  
Breast Cancer ◽  
Dna Methylation ◽  
Molecular Subtypes ◽  
Methylation Status ◽  
The Cancer Genome Atlas ◽  
Training Dataset ◽  
Consensus Clustering ◽  
Breast Cancer Patients ◽  
Cancer Subtypes ◽  
Novel Biomarkers

Background: Epigenetic changes are tightly linked to tumorigenesis development and malignant transformation’ However, DNA methylation occurs earlier and is constant during tumorigenesis. It plays an important role in controlling gene expression in cancer cells. Methods: In this study, we determining the prognostic value of molecular subtypes based on DNA methylation status in breast cancer samples obtained from The Cancer Genome Atlas database (TCGA). Results: Seven clusters and 204 corresponding promoter genes were identified based on consensus clustering using 166 CpG sites that significantly influenced survival outcomes. The overall survival (OS) analysis showed a significant prognostic difference among the 7 groups (p<0.05). Finally, a prognostic model was used to estimate the results of patients on the testing set based on the classification findings of a training dataset DNA methylation subgroups. Conclusions: The model was found to be important in the identification of novel biomarkers and could be of help to patients with different breast cancer subtypes when predicting prognosis, clinical diagnosis and management.

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