Colorimetric and electrochemical quantification of global DNA methylation using a methyl cytosine-specific antibody

The Analyst ◽  
2017 ◽  
Vol 142 (11) ◽  
pp. 1900-1908 ◽  
Author(s):  
Md. Hakimul Haque ◽  
Ripon Bhattacharjee ◽  
Md. Nazmul Islam ◽  
Vinod Gopalan ◽  
Nam-Trung Nguyen ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Specific Antibody ◽  
Electrochemical Method ◽  
Global Dna Methylation ◽  
Global Methylation ◽  
Naked Eye ◽  
Methyl Cytosine

We report a simple colorimetric (naked-eye) and electrochemical method for the rapid, sensitive and specific quantification of global methylation levels using only 25 ng of input DNA.

scholarly journals Oxidative damage, inflammation, genotoxic effect, and global DNA methylation caused by inhalation of formaldehyde and the purpose of melatonin

2020 ◽  
Vol 9 (6) ◽  
pp. 778-789
Author(s):  
Letícia Bernardini ◽  
Eduardo Barbosa ◽  
Mariele Feiffer Charão ◽  
Gabriela Goethel ◽  
Diana Muller ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Bone Marrow ◽  
Oxidative Damage ◽  
Bronchoalveolar Lavage ◽  
Bone Marrow Cells ◽  
Histological Study ◽  
Global Dna Methylation ◽  
Protective Effects ◽  
Global Methylation ◽  
Protein Thiols

Abstract Formaldehyde (FA) exposure has been proven to increase the risk of asthma and cancer. This study aimed to evaluate for 28 days the FA inhalation effects on oxidative stress, inflammation process, genotoxicity, and global DNA methylation in mice as well as to investigate the potential protective effects of melatonin. For that, analyses were performed on lung, liver and kidney tissues, blood, and bone marrow. Bronchoalveolar lavage was used to measure inflammatory parameters. Lipid peroxidation (TBARS), protein carbonyl (PCO), non-protein thiols (NPSH), catalase activity (CAT), comet assay, micronuclei (MN), and global methylation were determined. The exposure to 5-ppm FA resulted in oxidative damage to the lung, presenting a significant increase in TBARS and NO levels and a decrease in NPSH levels, besides an increase in inflammatory cells recruited for bronchoalveolar lavage. Likewise, in the liver tissue, the exposure to 5-ppm FA increased TBARS and PCO levels and decreased NPSH levels. In addition, FA significantly induced DNA damage, evidenced by the increase of % tail moment and MN frequency. The pretreatment of mice exposed to FA applying melatonin improved inflammatory and oxidative damage in lung and liver tissues and attenuated MN formation in bone marrow cells. The pulmonary histological study reinforced the results observed in biochemical parameters, demonstrating the potential beneficial role of melatonin. Therefore, our results demonstrated that FA exposure with repeated doses might induce oxidative damage, inflammatory, and genotoxic effects, and melatonin minimized the toxic effects caused by FA inhalation in mice.

scholarly journals Influence of tissue, age, and environmental quality on DNA methylation in Alligator mississippiensis

Reproduction ◽  
2014 ◽  
Vol 147 (4) ◽  
pp. 503-513 ◽  
Author(s):  
Benjamin B Parrott ◽  
John A Bowden ◽  
Satomi Kohno ◽  
Jessica A Cloy-McCoy ◽  
Matthew D Hale ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Whole Blood ◽  
Epigenetic Modification ◽  
Alligator Mississippiensis ◽  
Sentinel Species ◽  
Global Dna Methylation ◽  
American Alligator ◽  
Global Methylation ◽  
Contaminant Exposure ◽  
Liquid Chromatography Tandem Mass

Epigenetic modifications are key mediators of the interactions between the environment and an organism's genome. DNA methylation represents the best-studied epigenetic modification to date and is known to play key roles in regulating transcriptional activity and promoting chromosome stability. Our laboratory has previously demonstrated the utility of the American alligator (Alligator mississippiensis) as a sentinel species to investigate the persistent effects of environmental contaminant exposure on reproductive health. Here, we incorporate a liquid chromatography–tandem mass spectrometry method to directly measure the total (global) proportion of 5-methyl-2′-deoxycytidine (5mdC) in ovarian and whole blood DNA from alligators. Global DNA methylation in ovaries was significantly elevated in comparison with that of whole blood. However, DNA methylation appeared similar in juvenile alligators reared under controlled laboratory conditions but originating from three sites with dissimilar environmental qualities, indicating an absence of detectable site-of-origin effects on persistent levels of global 5mdC content. Analyses of tissues across individuals revealed a surprising lack of correlation between global methylation levels in blood and ovary. In addition, global DNA methylation in blood samples from juvenile alligators was elevated compared with those from adults, suggesting that age, as observed in mammals, may negatively influence global DNA methylation levels in alligators. To our knowledge, this is the first study examining global levels of DNA methylation in the American alligator and provides a reference point for future studies examining the interplay of epigenetics and environmental factors in a long-lived sentinel species.

scholarly journals A maternal effect regulates global DNA methylation patterns

2020 ◽  
Author(s):  
Remco Loos ◽  
Valeria Carola ◽  
Enrica Audero ◽  
Elena Brini ◽  
Luisa Lo Iacono ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Maternal Effect ◽  
Cpg Island ◽  
Inbred Mouse ◽  
Mouse Strains ◽  
Global Dna Methylation ◽  
Global Methylation ◽  
Genome Methylation ◽  
Genetic And Environmental Factors ◽  
Methylation Patterns

AbstractVariation in DNA methylation between individuals has been shown to be influenced by both genetic and environmental factors. However, the relative impact of genetic and non-genetic factors on DNA methylation patterns across the mammalian genome has not been systematically studied. We performed whole-genome methylation analysis in two inbred mouse strains, revealing striking differences in the global distribution of DNA methylation. Although global methylation patterns were indistinguishable for most genomic features, a significant increase in the number of unmethylated CpG-island promoters and first exons was observed between strains. Experiments using F1 reciprocal hybrid strains demonstrated that the genotype of the mother dictated global DNA methylation patterns. Cross-fostering experiments ruled out a postnatal maternal effect on these differences and suggested that they were driven by a prenatal maternal effect, possibly via differential deposition of maternal gene products into the oocyte or uterine environment. These data demonstrate that maternal effects have a major impact on global DNA methylation patterns.

scholarly journals The Long-Term Impact of Mild Traumatic Brain Injuries on Multiple Functional Outcomes and Epigenetics: A Pilot Study with College Students

2020 ◽  
Author(s):  
Hyunhwa Lee ◽  
Sungchul Lee ◽  
Ipuna Black ◽  
Laura Salado ◽  
Jonica Estrada ◽  
...  
Keyword(s):  
College Students ◽  
Dna Methylation ◽  
Pilot Study ◽  
Visual Function ◽  
Postural Balance ◽  
Sleep Disturbances ◽  
Global Dna Methylation ◽  
Balance Performance ◽  
Global Methylation

People who suffer a mild traumatic brain injury (mTBI) have heterogeneous symptoms and disease trajectories, which make it difficult to precisely diagnose and assess complications long-term. Insufficient information is available regarding how to precisely diagnose and assess mTBI. This study identified and compared deficits in cognitive, psychosocial, visual functions, and balance performance between college students with and without histories of mTBI. Global DNA methylation ratio (5-mC%) in blood was also compared as a peripheral epigenetic marker. Twenty-five volunteers participated in this pilot study, including 11 mTBI cases (27.3% females; mean age of 28.7 years, SD=5.92) and 14 healthy controls (64.3% females; mean age of 22.0, SD=4.13). All the participants were assessed for cognitive (by NIH toolbox—executive function, memory, and processing speed), psychological (by PROMIS—depression, anxiety, and sleep disturbances), visual function (by King-Devick and binocular accommodative tests), postural balance performance (by a force plate), and blood 5-mC% (global methylation) levels. Students with mTBI reported significantly poorer episodic memory, severe anxiety, and more sleep disturbance problems. They also had higher blood 5-mC% level (all p’s<.05). No significant differences were found in visual function and postural balance. These findings validate changes in cognitive, psychosocial, and global DNA methylation long after mTBI.

scholarly journals Porous nanozymes: the peroxidase-mimetic activity of mesoporous iron oxide for the colorimetric and electrochemical detection of global DNA methylation

2018 ◽  
Vol 6 (29) ◽  
pp. 4783-4791 ◽  
Author(s):  
Ripon Bhattacharjee ◽  
Shunsuke Tanaka ◽  
Sofia Moriam ◽  
Mostafa Kamal Masud ◽  
Jianjian Lin ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Iron Oxide ◽  
Electrochemical Detection ◽  
Global Dna Methylation ◽  
Naked Eye ◽  
Peroxidase Mimetic

Peroxidase-mimetic activity of mesoporous Fe2O3 nanomaterials in global DNA methylation detection using naked eye and electrochemical readout.

scholarly journals Global DNA methylation and cellular 5-methylcytosine and H4 acetylated patterns in primary and secondary dormant seeds of Capsella bursa-pastoris (L.) Medik. (shepherd’s purse)

PROTOPLASMA ◽  
2021 ◽  
Author(s):  
Sara Gomez-Cabellos ◽  
Peter E. Toorop ◽  
María Jesús Cañal ◽  
Pietro P. M. Iannetta ◽  
Eduardo Fernández-Pascual ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Seed Dormancy ◽  
Temporal Distribution ◽  
Global Dna Methylation ◽  
Histone H4 ◽  
Global Methylation ◽  
Secondary Dormancy ◽  
Spatio Temporal ◽  
Epigenetic Patterns

AbstractDespite the importance of dormancy and dormancy cycling for plants’ fitness and life cycle phenology, a comprehensive characterization of the global and cellular epigenetic patterns across space and time in different seed dormancy states is lacking. Using Capsella bursa-pastoris (L.) Medik. (shepherd’s purse) seeds with primary and secondary dormancy, we investigated the dynamics of global genomic DNA methylation and explored the spatio-temporal distribution of 5-methylcytosine (5-mC) and histone H4 acetylated (H4Ac) epigenetic marks. Seeds were imbibed at 30 °C in a light regime to maintain primary dormancy, or in darkness to induce secondary dormancy. An ELISA-based method was used to quantify DNA methylation, in relation to total genomic cytosines. Immunolocalization of 5-mC and H4Ac within whole seeds (i.e., including testa) was assessed with reference to embryo anatomy. Global DNA methylation levels were highest in prolonged (14 days) imbibed primary dormant seeds, with more 5-mC marked nuclei present only in specific parts of the seed (e.g., SAM and cotyledons). In secondary dormant seeds, global methylation levels and 5-mC signal where higher at 3 and 7 days than 1 or 14 days. With respect to acetylation, seeds had fewer H4Ac marked nuclei (e.g., SAM) in deeper dormant states, for both types of dormancy. However, the RAM still showed signal after 14 days of imbibition under dormancy-inducing conditions, suggesting a central role for the radicle/RAM in the response to perceived ambient changes and the adjustment of the seed dormancy state. Thus, we show that seed dormancy involves extensive cellular remodeling of DNA methylation and H4 acetylation.

scholarly journals Essential roles of Hdac1 and 2 in lineage development and genome-wide DNA methylation during mouse preimplantation development

2019 ◽  
Author(s):  
Panpan Zhao ◽  
Huanan Wang ◽  
Han Wang ◽  
Yanna Dang ◽  
Lei Luo ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Histone Modifications ◽  
Hippo Pathway ◽  
Epigenetic Modifications ◽  
Preimplantation Development ◽  
Global Dna Methylation ◽  
Lineage Specification ◽  
Pluripotent Cells ◽  
Global Methylation

AbstractEpigenetic modifications, including DNA methylation and histone modifications, are reprogrammed considerably following fertilization during mammalian early embryonic development. Incomplete epigenetic reprogramming is a major factor leading to poor developmental outcome in embryos generated by assisted reproductive technologies, such as somatic cell nuclear transfer. However, the role of histone modifications in preimplantation development is poorly understood. Here, we show that co-knockdown (cKD) of Hdac1 and 2 (but not individually) resulted in developmental failure during the morula to blastocyst transition. This outcome was also confirmed with the use of small-molecule Hdac1/2-specific inhibitor FK228. We observed reduced cell proliferation and increased incidence of apoptosis in cKD embryos, which were likely caused by increased acetylation of Trp53. Importantly, both RNA-seq and immunostaining analysis revealed a failure of lineage specification to generate trophectoderm and pluripotent cells. Among many gene expression changes, a substantial decrease of Cdx2 may be partly accounted for by the aberrant Hippo pathway occurring in cKD embryos. In addition, we observed an increase in global DNA methylation, consistent with increased DNA methyltransferases and Uhrf1. Interestingly, deficiency of Rbbp4 and 7 (both are core components of several Hdac1/2-containing epigenetic complexes) results in similar phenotypes as those of cKD embryos. Overall, Hdac1 and 2 play redundant functions required for lineage specification, cell viability and accurate global DNA methylation, each contributing to critical developmental programs safeguarding a successful preimplantation development.SignificanceSubstantial changes to epigenetic modifications occur during preimplantation development and can be detrimental when reprogrammed incompletely. However, little is known about the role of histone modifications in early development. Co-knockdown of Hdac1 and 2, but not individually, resulted in developmental arrest during morula to blastocyst transition, which was accompanied by reduced cell number per embryo and increased incidence of apoptosis. Additionally, we observed a failure of first lineage specification to generate trophectoderm and pluripotent cells, which were associated with reduced expression of key lineage-specific genes and aberrant Hippo pathway. Moreover, an increase in global DNA methylation was found with upregulated Dnmts and Uhrf1. Thus, Hdac1 and 2 play overlapping roles in lineage development, apoptosis, and global methylation during preimplantation development.

scholarly journals Global DNA methylation pattern and correlation with complement system proteins in Brazilian patients with systemic lupus erythematosus

2015 ◽  
Vol 13 (4) ◽  
pp. 516-523
Author(s):  
Paolo Ruggero Errante ◽  
Sandro Félix Perazzio ◽  
Francisco Sandro Menezes Rodrigues ◽  
Renato Ribeiro Nogueira Ferraz ◽  
Afonso Caricati-Neto
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Dna Methylation ◽  
Lupus Erythematosus ◽  
Genomic Dna ◽  
Expression Patterns ◽  
Global Dna Methylation ◽  
Healthy Individuals ◽  
Systemic Lupus ◽  
Global Methylation ◽  
Laboratory Parameters

Introduction: Nucleic acid methylation may have major effects on gene expression patterns and, by consequence, on the development of autoimmunity, like Systemic Lupus Erythematosus (SLE). Objective: To investigate the pattern of global DNA methylation in SLE patients and compare this pattern with laboratory parameters. Methods: Genomic DNA was isolated from SLE patients with non-active disease (SLEDAI6), and healthy individuals. Global DNA methylation was evaluated by digestion of genomic DNA with HpaII and MspI and compared with laboratory parameters. Results and conclusion: A statistical difference in DNA global methylation was observed when SLE patients were compared to healthy individuals. A positive correlation was observed between the frequency of global methylation and C3 and C4 serum levels for SLE patients with SLEDAI

Peripheral blood mononuclear cells are hypomethylated in active rheumatoid arthritis and methylation correlates with disease activity

Rheumatology ◽  
2020 ◽  
Author(s):  
Ilka Liebold ◽  
Andreas Grützkau ◽  
Anika Göckeritz ◽  
Velia Gerl ◽  
Randall Lindquist ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Disease Activity ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Global Dna Methylation ◽  
Peripheral Blood Mononuclear ◽  
Global Methylation ◽  
Blood Mononuclear Cells ◽  
Signal Intensities

Abstract Objective Epigenetic modifications are dynamic and influence cellular disease activity. The aim of this study was to investigate global DNA methylation in peripheral blood mononuclear cells (PBMCs) of RA patients to clarify whether global DNA methylation pattern testing might be useful in monitoring disease activity as well as the response to therapeutics. Methods Flow cytometric measurement of 5-methyl-cytosine (5′-mC) was established using the cell line U937. In the subsequent prospective study, 62 blood samples were investigated, including 17 healthy donors and 45 RA patients at baseline and after 3 months of treatment with methotrexate, the IL-6 receptor inhibitor sarilumab, and Janus kinase inhibitors. Methylation status was assessed with an anti-5′-mC antibody and analysed in PBMCs and CD4+, CD8+, CD14+ and CD19+ subsets. Signal intensities of 5′-mC were correlated with 28-joint DASs with ESR and CRP (DAS28-ESR and DAS28-CRP). Results Compared with healthy individuals, PBMCs of RA patients showed a significant global DNA hypomethylation. Signal intensities of 5′-mC correlated with transcription levels of DNMT1, DNMT3B and MTR genes involved in methylation processes. Using flow cytometry, significant good correlations and linear regression values were achieved in RA patients between global methylation levels and DAS28-ESR values for PBMCs (r = −0.55, P = 0.002), lymphocytes (r = −0.57, P = 0.001), CD4+ (r = −0.57, P = 0.001), CD8+ (r = −0.54, P = 0.001), CD14+ (r = −0.49, P = 0.008) and CD19+ (r = −0.52, P = 0.004) cells. Conclusions The degree of global DNA methylation was found to be associated with disease activity. Based on this novel approach, the degree of global methylation is a promising biomarker for therapy monitoring and the prediction of therapy outcome in inflammatory diseases.

scholarly journals Association of air pollution and homocysteine with global DNA methylation: A population-based study from North India

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0260860
Author(s):  
Suniti Yadav ◽  
Imnameren Longkumer ◽  
Priyanka Rani Garg ◽  
Shipra Joshi ◽  
Sunanda Rajkumari ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Air Pollution ◽  
Methylation Status ◽  
Population Based ◽  
North India ◽  
Nutritional Deficiencies ◽  
Global Dna Methylation ◽  
Vitamin B ◽  
Global Methylation ◽  
B Vitamin

Background Anthropogenic air pollution has been implicated in aberrant changes of DNA methylation and homocysteine increase (>15μM/L). Folate (<3 ng/mL) and vitamin B12 (<220 pg/mL) deficiencies also reduce global DNA methylation via homocysteine increase. Although B-vitamin supplements can attenuate epigenetic effects of air pollution but such understanding in population-specific studies are lacking. Hence, the present study aims to understand the role of air pollution, homocysteine, and nutritional deficiencies on methylation. Methods We examined cross-sectionally, homocysteine, folate, vitamin B12 (chemiluminescence) and global DNA methylation (colorimetric ELISA Assay) among 274 and 270 individuals from low- and high- polluted areas, respectively, from a single Mendelian population. Global DNA methylation results were obtained on 254 and 258 samples from low- and high- polluted areas, respectively. Results Significant decline in median global DNA methylation was seen as a result of air pollution [high-0.84 (0.37–1.97) vs. low-0.96 (0.45–2.75), p = 0.01]. High homocysteine in combination with air pollution significantly reduced global DNA methylation [high-0.71 (0.34–1.90) vs. low-0.93 (0.45–3.00), p = 0.003]. Folate deficient individuals in high polluted areas [high-0.70 (0.37–1.29) vs. low-1.21 (0.45–3.65)] showed significantly reduced global methylation levels (p = 0.007). In low polluted areas, despite folate deficiency, if normal vitamin B12 levels were maintained, global DNA methylation levels improved significantly [2.03 (0.60–5.24), p = 0.007]. Conversely, in high polluted areas despite vitamin B12 deficiency, if normal folate status was maintained, global DNA methylation status improved significantly [0.91 (0.36–1.63)] compared to vitamin B12 normal individuals [0.54 (0.26–1.13), p = 0.04]. Conclusions High homocysteine may aggravate the effects of air pollution on DNA methylation. Vitamin B12 in low-polluted and folate in high-polluted areas may be strong determinants for changes in DNA methylation levels. The effect of air pollution on methylation levels may be reduced through inclusion of dietary or supplemented B-vitamins. This may serve as public level approach in natural settings to prevent metabolic adversities at community level.

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