scholarly journals Epigenetic Changes in Equine Embryos After Short-term Storage at Different Temperatures

2021 ◽  
Author(s):  
Gustavo Desire Antunes Gastal ◽  
Dragos Scarlet ◽  
Maria Melchert ◽  
Reinhard Ertl ◽  
Christine Aurich
Keyword(s):  
Dna Methylation ◽  
Growth And Development ◽  
Methylation Status ◽  
Embryo Morphology ◽  
Global Dna Methylation ◽  
Short Term ◽  
Global Methylation ◽  
Short Term Storage ◽  
Maternal Recognition Of Pregnancy ◽  
Term Storage

Abstract Background: In embryos subjected to assisted reproductive techniques, epigenetic modifications may occur that can influence embryonic development and establishment of pregnancy. In horses, the storage temperature during transport of fresh embryos before transfer is a major concern. The aim of this study was to determine the effects of two storage temperatures (5˚C and 20˚C) on equine embryos, collected at day 7 after ovulation and stored for 24 hours, on the following characteristics: (i) morphological and histological development; (ii) expression of candidate genes associated with embryo growth and development, maternal recognition of pregnancy, methylation, and apoptosis; and (iii) gene-specific and global-DNA methylation. Embryos (n=80) were collected from Haflinger mares (n=15) on day 7 (n=60) or day 8 (n=20) after ovulation and assigned to 4 groups: day 7 control (E7F, fresh); day 7, stored for 24h at 5°C (E5C); day 7, stored for 24h at 20°C (E20C); and day 8 control (E8F, fresh 24h time control). The embryos and the storage medium from all groups were analyzed for: (i) medium temperature, pH, and lipid peroxidation (malondialdehyde; MDA), (ii), embryo morphology, mRNA expression, and DNA methylation (immunohistochemistry and gene-specific DNA methylation). Results: Temperature during storage (E5C and E20C) did not affect embryo size (382±47 and 553±99 µm, respectively). There were no changes in pH and MDA accumulation irrespective of group. The relative mRNA abundance of specific genes related to growth and development (POU5F1, SOX2, NANOG), maternal recognition of pregnancy (CYP19A1, PTGES2), DNA methylation (DNMT1, DNMT3A, DNMT3B), and apoptosis (BAX) in the E5C and E20C were either up or downregulated (P<0.05) when compared to controls (E7F and E8F). The global methylation status, even as 5mC and 5hmC immune expression were similar among treatment groups. The specific genes ESR1, NANOG, and DNMT1 were hypomethylated (P<0.001) in E20C embryos when compared to E8F (advanced stage). Conclusions: Therefore, our study demonstrates for the first time the gene-specific and global-DNA methylation status of fresh equine embryos collected on days 7 and 8 after ovulation. In addition, short-term storage, regardless of temperature, modified gene expression and methylation of genes involved in embryo development and may therefore compromise embryo viability and development after transfer.

85 Influence of Short-Term Storage on Gene Expression of Equine Embryos

2018 ◽  
Vol 30 (1) ◽  
pp. 182
Author(s):  
G. D. A. Gastal ◽  
D. Scarlet ◽  
R. Ertl ◽  
C. Aurich
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Embryo Development ◽  
Geometric Mean ◽  
Rna Extraction ◽  
Short Term ◽  
Expression Of Genes ◽  
Short Term Storage ◽  
Term Storage ◽  
Fresh Embryos

Short-term storage for embryo transportation is a routine procedure in equine embryo transfer programs. The success rate after transfer of stored embryos varies among different protocols when embryos are transported overnight. To the best of our knowledge, there is no study evaluating the effect of different temperatures during storage for 24 h on gene expression of equine embryos. Therefore, this study aimed to evaluate the effects of storage of equine embryos for 24 h at 2 temperatures (20°C or 5°C) on the expression of genes related to embryo development (IGF2, H19, POU5F1, SOX2), and regulation of DNA methylation (DNMT1, DNMT3a, DNMT3b). Embryos (n = 24) were collected on Day 7 (n = 18) or Day 8 (n = 6) after ovulation and assigned to 4 groups: Day 7 control (D7, fresh); Day 7, 24 h at 5°C (E5C); Day 7, 24 h at 20°C (E20C); and Day 8 control (D8, fresh 24-h time control). After flushing, embryos were washed and kept in holding medium (Minitube, Tiefenbach, Germany) for morphological classification and measurements. Fresh and stored embryos were treated with pronase (10 mg mL−1), washed with PBS solution and placed in RLT Lysis buffer (Qiagen, Hilden, Germany) for RNA extraction. Total RNA was extracted from each individual embryo using the RNeasy mini kit (Qiagen) following the recommended protocol for animal tissues. After RNA purification, RNA quality was assessed and quantified. Subsequently, cDNA synthesis was performed for RT-qPCR analysis. Two replicates were performed and the relative gene expression was calculated using the 2(–delta delta CT) method, with the target gene expression levels normalized to the geometric mean of PSMB4/SNRPD3. The software SPSS v.24 (IBM/SPSS, Armonk, NY, USA) was used for statistical analyses using the nonparametric tests Kruskal-Wallis and Mann-Whitney U-test to compare differences among groups. Embryos sizes differed (P < 0.05) between D7 (431 ± 48 mm) and D8 (1114 ± 205 mm). Storage temperature did not affect (P > 0.05) embryo size. The mRNA expression of H19 and IGF2 was similar (P > 0.05) among all groups. Expression of POU5F1 and SOX2 was higher (P < 0.05) in D7 and E5C embryos compared with D8 embryos. In addition, E20C had similar (P > 0.05) expression of POU5F1 with D7, E5C, and D8, but lower (P < 0.05) expression of SOX2 when compared with D7. Expression of DNMT1 and DNMT3a were similar (P > 0.05) among D7, D8, and E5C, but lower (P < 0.05) in E20C. Furthermore, expression of DNMT3b was lower (P < 0.05) in D8 and E20C embryos compared with D7 and E5C. In conclusion, temperature during short-term storage seems not to affect the expression of IGF2 and H19 but influences the expression of POU5F1, SOX2, DNMT1, DNMT3a, and DNMT3b. Therefore, these findings suggest that embryos stored at 20°C sustain the pattern of gene expression similar to that of fresh embryos at Day 8, whereas embryos stored at 5°C maintain the gene expression similar to that of fresh embryos at Day 7. Thus, alterations caused by temperature during short-term storage on the expression of genes related to embryo development and DNA methylation may modify the pattern of equine embryonic tissue development, requiring further investigation.

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.

scholarly journals Analysis at the single-cell level indicates an important role of heterogeneous global DNA methylation status on the progression of lung adenocarcinoma

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Quan-Fang Chen ◽  
Han Gao ◽  
Qing-Yun Pan ◽  
Ying-Ju Wang ◽  
Xiao-Ning Zhong
Keyword(s):  
Dna Methylation ◽  
Lung Adenocarcinoma ◽  
Single Cell ◽  
Tumor Cells ◽  
Methylation Status ◽  
Cell Junctions ◽  
Global Dna Methylation ◽  
Single Cell Level ◽  
Cell Level ◽  
Global Methylation

AbstractAberrant DNA modifications affect the tumorigenesis and progression of lung cancer. However, the global methylation status of tumor cells and the heterogeneous methylation status of cells within the same tumor need further study. We used publicly available single-cell RNAseq data to investigate the impact and diversity of global methylation status on lung adenocarcinoma. Clustering cells into subgroups and cell differentiation pseudotime analysis, based on expression profile, demonstrated that the global methylation status was crucial to lung adenocarcinoma function and progression. Hypermethylated tumor cells had increased activity related to the hypoxia response. Hyper- and hypomethylated cells indicated upregulation in pathways involving focal adhesion and cell junctions. Pseudotime analysis showed that cell clusters with unique methylation activities were located at the ends of the putative trajectories, suggesting that DNA methylation and demethylation activities were essential to tumor cell progression. Expression of SPP1 was associated with the global methylation status of tumor cells and with patient prognosis. Our study identified the importance and diversity of global DNA methylation status by analysis at the single-cell level. Our findings provide new information about the global DNA methylation status of tumor cells and suggest new approaches for precision medical treatments for lung adenocarcinoma.

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 The effect of short-term storage temperature on the key headspace volatile compounds observed in Canadian faba bean flour

2021 ◽  
pp. 108201322199884
Author(s):  
Rami Akkad ◽  
Ereddad Kharraz ◽  
Jay Han ◽  
James D House ◽  
Jonathan M Curtis
Keyword(s):  
Fatty Acids ◽  
Volatile Compounds ◽  
Room Temperature ◽  
Unsaturated Fatty Acids ◽  
Solid Phase ◽  
Short Term ◽  
Bean Flour ◽  
Odour Activity Value ◽  
Short Term Storage ◽  
Term Storage

The odour emitted from the high-tannin fab bean flour ( Vicia faba var. minor), was characterized by headspace solid-phase microextraction/gas chromatography-mass spectrometry (HS-SPME/GC–MS). The relative odour activity value (ROAV) was used to monitor the changes in key volatile compounds in the flour during short-term storage at different temperature conditions. The key flavour compounds of freshly milled flour included hexanal, octanal, nonanal, decanal, 3-methylbutanal, phenyl acetaldehyde, (E)-2-nonenal, 1-hexanol, phenyl ethyl alcohol, 1-octen-3-ol, β-linalool, acetic acid, octanoic acid, and 3-methylbutyric acid; these are oxidative degradation products of unsaturated fatty acids and amino acids. Despite the low lipid content of faba beans, the abundances of aldehydes arising during room temperature storage greatly contributed to the flavour of the flour due to their very low odour thresholds. Two of the key volatiles responsible for beany flavour in flour (hexanal, nonanal) increased greatly after 2 weeks of storage at room temperature or under refrigerated conditions. These volatile oxidation products may arise as a result of enzymatic activity on unsaturated fatty acids, and was seen to be arrested by freezing the flour.

Effect of short-term storage on phenolic content,o-diphenolase and peroxidase activities of cut yam tubers (Dioscorea sp)

1992 ◽  
Vol 60 (3) ◽  
pp. 309-312 ◽  
Author(s):  
Helen N. Asemota ◽  
Max A. Wellington ◽  
Adewale A. Odutuga ◽  
Mohammed H. Ahmad
Keyword(s):  
Phenolic Content ◽  
Short Term ◽  
Short Term Storage ◽  
Term Storage
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