Paternal Exercise Improves the Metabolic Health of Offspring via Epigenetic Modulation of the Germline

Background/Aims: Epigenetic regulation is considered the main molecular mechanism underlying the developmental origin of health and disease’s (DOHAD) hypothesis. Previous studies that have investigated the role of paternal exercise on the metabolic health of the offspring did not control for the amount and intensity of the training or possible effects of adaptation to exercise and produced conflicting results regarding the benefits of parental exercise to the next generation. We employed a precisely regulated exercise regimen to study the transgenerational inheritance of improved metabolic health. Methods: We subjected male mice to a well-controlled exercise -training program to investigate the effects of paternal exercise on glucose tolerance and insulin sensitivity in their adult progeny. To investigate the molecular mechanisms of epigenetic inheritance, we determined chromatin markers in the skeletal muscle of the offspring and the paternal sperm. Results: Offspring of trained male mice exhibited improved glucose homeostasis and insulin sensitivity. Paternal exercise modulated the DNA methylation profile of PI3Kca and the imprinted H19/Igf2 locus at specific differentially methylated regions (DMRs) in the skeletal muscle of the offspring, which affected their gene expression. Remarkably, a similar DNA methylation profile at the PI3Kca, H19, and Igf2 genes was present in the progenitor sperm indicating that exercise-induced epigenetic changes that occurred during germ cell development contributed to transgenerational transmission. Conclusion: Paternal exercise might be considered as a strategy that could promote metabolic health in the offspring as the benefits can be inherited transgenerationally.

Aqueous Extract of Alstonia boonei Bark Reduces Chronic Hyperglycemia and Prevents its Complications through Increase of Hepatic Global Dna Methylation in Diabetic Wistar Rats

Aim: DNA methylation profile is involved in several physiological processes. Its alterations in the liver of diabetic patients characterized by global hypomethylation are associated with the pathophysiology of type 2 diabetes and its complications. The present study has evaluated the effect of the aqueous extract of Alstonia boonei barks on the global methylation of hepatic DNA in association with hyperglycemia and diabetes complications induced by high-fat diet (HFD) feeding and administering of streptozotocin (STZ) which mimics the metabolic abnormalities very similar to those seen in human Type 2 diabetes. Methods: A. boonei barks were harvested, processed, dried, ground and an aqueous extraction was prepared (ratio 1/10 w/v). An in vivo study was conducted in an animal model of high-fat-streptozotocin (HF-STZ) induced diabetes. Rats were divided into five groups of five rats each: a normoglycemic group, an untreated hyperglycemic group, three hyperglycemic groups including two test groups receiving aqueous extract of A. boonei barks (AEAB) by esophageal gavage at the doses of 200 and 400 mg/kg body weight once daily and a reference group receiving metformin at 10 mg/kg body weight. After 28 days of experimentation during which fasting blood glucose levels were taken every 14 days under fasting conditions, the animals were sacrificed. Plasma and liver homogenate samples from the sacrificed rats were used for biochemical assays (markers of oxidative stress such as malondialdehyde level, superoxide dismutase (SOD) and catalase activity, and markers of lipid profile such as total cholesterol, and triglycerides, HDL-c, LDL-c and VLDL-c). The analysis of the global DNA methylation profile was performed by the immunoprecipitation. Pearson's correlation was used to evaluate the relationship between the values. Results: The aqueous extract increased the hepatic DNA methylation by 0.41% and 0.63% at 200 and 400 mg/kg body weight, respectively, compared to metformin (0.47%±0, 05). This effect was significantly associated with the hypoglycemic effect obtained at 400 mg/kg body weight with a decrease in initial blood glucose level of -29.87%. Conclusion: AEAB reduces chronic hyperglycemia and prevents its complications by increasing global hepatic DNA methylation.

Methylation of BDNF and SLC6A4 Gene Promoters in Brazilian Patients With Temporal Lobe Epilepsy Presenting or Not Psychiatric Comorbidities

The relationship between epilepsy and psychiatric comorbidities has been recognized for centuries, but its pathophysiological mechanisms are still misunderstood. It is biologically plausible that genetic or epigenetic variations in genes that codify important neurotransmitters involved in epilepsy as well as in psychiatric disorders may influence the development of the latter in patients with epilepsy. However, this possibility remains poorly investigated. The aim of this study was to evaluate the methylation profile of the BDNF and SLC6A4, two genes importantly involved in neuroplasticity, in patients with temporal lobe epilepsy (TLE) regarding the development or not of psychiatric comorbidities. One hundred and thirty-nine patients with TLE, 90 females and 45 males, were included in the study. The mean age of patients was 44.0 (+12.0) years, and mean duration of epilepsy was 25.7 (+13.3) years. The Structured Clinical Interview for DSM-IV shows that 83 patients (59.7%) had neuropsychiatric disorders and 56 (40.3%) showed no psychiatric comorbidity. Mood disorders were the most common psychiatric disorder observed, being present in 64 (46.0%) of all 139 patients. Thirty-three (23.7%) patients showed anxiety disorders, 10 (7.2%) patients showed history of psychosis and 8 (5.8%) patients showed history of alcohol//drug abuse. Considering all 139 patients, 18 (12.9%) demonstrated methylation of the promoter region of both BDNF and SLC6A4 genes. A significant decreased methylation profile was observed only in TLE patients with mood disorders when compared with TLE patients without a history of mood disorders (O.R. = 3.45; 95% C.I. = 1.08–11.11; p = 0.04). A sub-analysis showed that TLE patients with major depressive disorder mostly account for this result (O.R. = 7.20; 95% C.I. = 1.01–56.16; p = 0.042). A logistic regression analysis showed that the independent factors associated with a history of depression in our TLE patients was female sex (O.R. = 2.30; 95% C.I. = 1.02–5.18; p = 0.044), not controlled seizures (O.R. = 2.51; 95% C.I. = 1.16–5.41; p = 0.019) and decreased methylation in BDNF and SLC6A4 genes (O.R. = 5.32; 95% C.I. = 1.14–25.00; p = 0.033). Our results suggest that BDNF or SLC6A4 genes profile methylation is independently associated with depressive disorders in patients with epilepsy. Further studies are necessary to clarify these matters.

Methylome and transcriptome analyses of soybean response to bean pyralid larvae

Background Bean pyralid is one of the major leaf-feeding insects that affect soybean crops. DNA methylation can control the networks of gene expressions, and it plays an important role in responses to biotic stress. However, at present the genome-wide DNA methylation profile of the soybean resistance to bean pyralid has not been reported so far. Results Using whole-genome bisulfite sequencing (WGBS) and RNA-sequencing (RNA-seq), we analyzed the highly resistant material (Gantai-2-2, HRK) and highly susceptible material (Wan82–178, HSK), under bean pyralid larvae feeding 0 h and 48 h, to clarify the molecular mechanism of the soybean resistance and explore its insect-resistant genes. We identified 2194, 6872, 39,704 and 40,018 differentially methylated regions (DMRs), as well as 497, 1594, 9596 and 9554 differentially methylated genes (DMGs) in the HRK0/HRK48, HSK0/HSK48, HSK0/HRK0 and HSK48/HRK48 comparisons, respectively. Through the analysis of global methylation and transcription, 265 differentially expressed genes (DEGs) were negatively correlated with DMGs, there were 34, 49, 141 and 116 negatively correlated genes in the HRK0/HRK48, HSK0/HSK48, HSK0/HRK0 and HSK48/HRK48, respectively. The MapMan cluster analysis showed that 114 negatively correlated genes were clustered in 24 pathways, such as protein biosynthesis and modification; primary metabolism; secondary metabolism; cell cycle, cell structure and component; RNA biosynthesis and processing, and so on. Moreover, CRK40; CRK62; STK; MAPK9; L-type lectin-domain containing receptor kinase VIII.2; CesA; CSI1; fimbrin-1; KIN-14B; KIN-14 N; KIN-4A; cytochrome P450 81E8; BEE1; ERF; bHLH25; bHLH79; GATA26, were likely regulatory genes involved in the soybean responses to bean pyralid larvae. Finally, 5 DMRs were further validated that the genome-wide DNA data were reliable through PS-PCR and 5 DEGs were confirmed the relationship between DNA methylation and gene expression by qRT-PCR. The results showed an excellent agreement with deep sequencing. Conclusions Genome-wide DNA methylation profile of soybean response to bean pyralid was obtained for the first time. Several specific DMGs which participated in protein kinase, cell and organelle, flavonoid biosynthesis and transcription factor were further identified to be likely associated with soybean response to bean pyralid. Our data will provide better understanding of DNA methylation alteration and their potential role in soybean insect resistance.

Development of a Machine Learning Classifier for Brain Tumors Diagnosis Based on DNA Methylation Profile

Background: More than 150 types of brain tumors have been documented. Accurate diagnosis is important for making appropriate therapeutic decisions in treating the diseases. The goal of this study is to develop a DNA methylation profile-based classifier to accurately identify various kinds of brain tumors.Methods: Thirteen datasets of DNA methylation profiles were downloaded from the Gene Expression Omnibus (GEO) database, of which GSE90496 and GSE109379 were used as the training set and the validation set, respectively, and the remaining 11 sets were used as the independent test set. The random forest algorithm was used to select the CpG sites based on the importance of the features and a multilayer perceptron (MLP) model was trained to classify the samples. Deconvolution with the debCAM package was used to explore the cellular composition difference among tumors.Results: From training datasets with 2,801 samples, 396,568 CpG sites were retained after preprocessing, of which 767 were selected as the modeling features. A three-layer MLP model was developed, which consists of 1,320 nodes in the hidden layer, to predict the histological types of brain tumors. The prediction accuracy is 99.2, 87.0, and 96.58%, respectively, on the training, validation and test sets. The results of deconvolution analysis showed that the cell proportions of different tumor subtypes were different, and it is approximately enough to distinguish different tumor entities.Conclusion: We developed a classifier that is robust for the classification of central nervous system tumors, and tried to analyze the reasons for the classification performance.

Restriction-modification systems and DNA methylation profile of Pectobacterium carotovorum 2A

The methylation profile of Pectobacterium carotovorum 2A genome was studied using the Oxford Nanopore sequencing technology. The specificity of the methylase subunits of the three restriction-modification systems of this strain was determined. Analysis of homologous systems showed the uniqueness of the type I restriction-modification system and the type IV restriction system specific to methylated DNA of this strain. The work confirms the applicability of Oxford Nanopore technology to the analysis of bacterial DNA modifications and is also the first example of such an analysis for Pectobacterium spp.

907 Modulation of tumor immunogenicity by DNA methylation of immune synapse genes in cancers

BackgroundCancer immunotherapy represents a major paradigm shift in cancer care. Despite such breakthrough, majority of cancer patients remains refractory to existing immunotherapeutic modalities highlighting the inherent capacity of tumors to escape immunosurveillance mechanisms. Frequently, cancer cells utilize the epigenetic machinery to silence tumor suppressors or activate oncogenes for survival and proliferation. Likewise, tumor cells might employ the epigenetic reprogramming of immune-related pathways to evade the immune system. Methylation is one of the major epigenetic mechanisms modulating gene transcription. Thus, we investigated the methylation profile of both co-stimulatory and immune checkpoint genes in cancer.MethodsData from The Cancer Genome Atlas (TCGA) were used for methylation profiling and RNA-sequencing analysis. Twenty-six epithelial cancer cell lines with more than 3 mock and three 5-azacitidine–treated samples were selected for analysis from the GSE57342 dataset. t-distributed stochastic neighbor embedding (t-SNE) was calculated using 247 probes for the selected 20 genes across all TCGA samples. t-SNE analysis was performed on 8,186 solid tumors and 745 normal adjacent tissues for methylation levels for all probes. For principal component analysis, first and second principal components were used to represent the overall methylation status for 8,931 tumor and normal samples in the TCGA database. Survival analyses were retrieved from a prior publication.1ResultsWe found that methylation profile of immune synapse genes is distinct in tumor versus normal adjacent tissue. Interestingly, our results demonstrate hypermethylation of co-stimulatory genes such as CD40 and hypo-methylation of immune checkpoint genes such as HHLA2 and PDL1 across multiple tumor types in comparison with the normal adjacent tissue. In addition, an inverse correlation between methylation and gene expression was manifest among tumor and normal adjacent tissue, confirming the epigenetic mechanism of gene suppression by gene methylation. Furthermore, we observed a reversal of hypermethylation of the co-stimulatory genes including CD40 by the demethylating agent 5-azacytidine in the data set of 26 epithelial cancer cell lines. Finally, we found that that hypomethylation of co-stimulatory genes within the immune synapse correlates with functional T cell recruitment to the tumor microenvironment and is followed by a favorable clinical outcome in melanoma patients.ConclusionsOur finding unveils methylation of immune synapse genes as a crucial driver of the immune evasive phenotype of cancer cells. Notably, identification of actionable targets to restore tumor immunogenicity is an attractive strategy in combination with immune checkpoint blockade.AcknowledgementsThis work was supported by NIH grant K08 CA194273, the Immunology Innovation Fund, an NCI Cancer Center Support grant, (P30-CA076292), the Miriam and Sheldon G. Adelson Foundation, and the Moffitt Foundation.ReferenceLiu J, Lichtenberg T, Hoadley KA, Poisson LM, Lazar AJ, Cherniack AD, Kovatich AJ, Benz CC, Levine DA, Lee AV, Omberg L, Wolf DM, Shriver CD, Thorsson V, Cancer Genome Atlas Research N, Hu H. An integrated TCGA pan-cancer clinical data resource to drive high-quality survival outcome analytics. Cell 2018;173(2):400–16 e11.