Hypermethylation Of Genes Research Articles

DNA hypermethylation of tumor suppressor genes among oral squamous cell carcinoma patients: a prominent diagnostic biomarker.

Oral Squamous Cell Carcinoma is a globally revealing form of oral malignancy. Epigenetics, which studies genetic modifications in gene expression without altering the sequence of DNA, is crucial for understanding OSCC. Key epigenetic modifications such as histone modifications, DNA methylation, and microRNA regulation play significant roles in Oral carcinoma. Aberrant methylation of DNA of tumor suppressor genes which leads to their inactivation, promoting cancer development, and specific methylation patterns are emerging as biomarkers for early OSCC detection.Current treatments like surgery, radiotherapy, and chemotherapy often fall short, prompting research into epigenetic therapies. Agents like DNMT and HDAC inhibitors demonstrate the potential for reversing aberrant epigenetic patterns, perhaps reactivating silenced TSGs, and suppressing oncogenes. Despite early promise, the development of effective combination medicines and the identification of reliable biomarkers continue to present challenges.In OSCC, resistance to therapy is also influenced by epigenetic processes. Aberrant DNA methylation and changes in histone modifications impact genes involved in medication metabolism and the survival of cells. Enhancing treatment efficacy and overcoming medication resistance may be possible by recognizing and focusing on these processes. This review explores the interplay between epigenetic changes and OSCC, their role in the disease's initiation and progression, and their impact on diagnosis and treatment. It also discusses the potential of epigenetic drugs (epi-drugs) to improve diagnostic precision and treatment outcomes.

Transcriptome-wide identification and analysis reveals m6A regulation of metabolic reprogramming in shrimp (Marsupenaeus japonicus) under virus infection.

It has been reported that the most common post-transcriptional modification of eukaryotic RNA is N6-methyladenosine (m6A). Previous studies show m6A is a key regulator for viral infection and immune response. However, whether there is a pathogen stimulus-dependent m6A regulation in invertebrate shrimp has not been studied. In this study, we performed a transcriptome-wide profiling of mRNA m6A methylation in shrimp (Marsupenaeus japonicus) after white spot syndrome virus (WSSV) infection by methylated RNA immunoprecipitation sequencing (MeRIP-seq). A total of 15,436 m6A peaks were identified in the shrimp, distributed in 8,108 genes, mainly enriched in the CDS, 3' UTR region and near the stop codon. After WSSV infection, we identified 2,260 m6A peaks with significantly changes, of which 1,973 peaks were significantly up-regulated and 287 peaks were significantly down-regulated. 1,795 genes were identified as differentially methylated genes. GO and KEGG analysis showed that hyper-methylated genes or hypo-methylated genes were highly associated with innate immune process and related to metabolic pathways including HIF-1 signaling pathway, lysine degradation and Wnt signaling pathway. Combined analysis showed a positive correlation between m6A methylation levels and mRNA expression levels. In addition, computational predictions of protein-protein interaction indicated that genes with altered levels of m6A methylation and mRNA expression clustered in metabolism, DNA replication, and protein ubiquitination. ZC3H12A and HIF-1 were two hub genes in protein-protein interaction (PPI) network that involved in immune and metabolism processes, respectively. Our study explored the m6A methylation pattern of mRNA in shrimp after WSSV infection, exhibited the first m6A map of shrimp at the stage of WSSV induced metabolic reprogramming. These findings may reveal the possible mechanisms of m6A-mediated innate immune response in invertebrates.

DNA methylation analysis of multiple genes in thymic epithelial tumors.

Aim: To investigate DNA methylation levels of a panel of genes in thymic epithelial tumors (TETs).Materials & methods: We selected 15 genes among the most promising epigenetic biomarkers of TETs and evaluated their methylation levels in 71 TET samples.Results: thymic carcinomas (TCs) showed hypermethylation of GHSR and ELF3 genes and reduced IL1RN methylation levels compared with thymomas (TMs) and healthy thymic tissues. RAG1 was hypomethylated in TMs compared with healthy thymic tissues. No difference in the methylation levels of the investigated genes was seen among TM stages and subtypes. No changes in blood methylation levels of the investigated genes were seen among TET subtypes.Conclusion: The present study confirms GHSR, ELF3, IL1RN and RAG1 as TET epigenetic biomarkers.

Discovery of KW0113 as a First and Effective PROTAC Degrader of DNMT1 Protein.

DNA methyltransferase 1 (DNMT1) is an attractive therapeutic target for acute myelocytic leukemia (AML) and other malignancies. It has been reported that the genetic depletion of DNMT1 inhibited AML cell proliferation through reversing DNA methylation abnormalities. However, no DNMT1-targeted PROTAC degraders have been reported yet. Herein, a series of proteolysis-targeting chimera (PROTAC) degrader of DNMT1 based on dicyanopyridine scaffold and VHL E3 ubiquitin ligase ligand was developed. Among them, KW0113 (DC50=643/899 nM in MV4-11/MOLM-13 cells) exhibited optimal DNMT1 degradation. KW0113 induced DNMT1-selective degradation in a dose- and time-dependent manner through VHL engagement. Moreover, KW0113 inhibited AML cell growth by reversing promoter DNA hypermethylation and tumor-suppressor genes silencing. In conclusion, these findings proved the capability of PROTAC strategy for inducing DNMT1 degradation, demonstrated the therapeutic potential of DNMT1-targeted PROTACs. This work also provided a convenient chemical knockdown tool for DNMT1-related studies.

Ten Hypermethylated lncRNA Genes Are Specifically Involved in the Initiation, Progression, and Lymphatic and Peritoneal Metastasis of Epithelial Ovarian Cancer.

Our work aimed to evaluate and differentiate the role of ten lncRNA genes (GAS5, HAND2-AS1, KCNK15-AS1, MAGI2-AS3, MEG3, SEMA3B-AS1, SNHG6, SSTR5-AS1, ZEB1-AS1, and ZNF667-AS1) in the development and progression of epithelial ovarian cancer (EOC). A representative set of clinical samples was used: 140 primary tumors from patients without and with metastases and 59 peritoneal metastases. Using MS-qPCR, we demonstrated an increase in methylation levels of all ten lncRNA genes in tumors compared to normal tissues (p < 0.001). Using RT-qPCR, we showed downregulation and an inverse relationship between methylation and expression levels for ten lncRNAs (rs < -0.5). We further identified lncRNA genes that were specifically hypermethylated in tumors from patients with metastases to lymph nodes (HAND2-AS1), peritoneum (KCNK15-AS1, MEG3, and SEMA3B-AS1), and greater omentum (MEG3, SEMA3B-AS1, and ZNF667-AS1). The same four lncRNA genes involved in peritoneal spread were associated with clinical stage and tumor extent (p < 0.001). Interestingly, we found a reversion from increase to decrease in the hypermethylation level of five metastasis-related lncRNA genes (MEG3, SEMA3B-AS1, SSTR5-AS1, ZEB1-AS1, and ZNF667-AS1) in 59 peritoneal metastases. This reversion may be associated with partial epithelial-mesenchymal transition (EMT) in metastatic cells, as indicated by a decrease in the level of the EMT marker, CDH1 mRNA (p < 0.01). Furthermore, novel mRNA targets and regulated miRNAs were predicted for a number of the studied lncRNAs using the NCBI GEO datasets and analyzed by RT-qPCR and transfection of SKOV3 and OVCAR3 cells. In addition, hypermethylation of SEMA3B-AS1, SSTR5-AS1, and ZNF667-AS1 genes was proposed as a marker for overall survival in patients with EOC.

Genome-wide DNA methylation profiling reveals a novel hypermethylated biomarker PRKCB in gastric cancer

Globally, gastric cancer (GC) ranks among the most prevalent forms of malignancy, posing a significant health burden. Epigenetic modifications, predominantly characterized by alterations in DNA methylation patterns, have been linked to a diverse array of neoplastic processes. Here, we undertake a comprehensive analysis of the DNA methylation signature in GC, with the aim to discover the potential diagnostic epigenetic biomarkers. Utilizing the Illumina 935 K BeadChip, we conducted a genome-wide exploration of DNA methylation patterns in four paired samples of GC tissues and adjacent non-cancerous counterparts. The bisulfite-pyrosequencing (n = 7) was employed to the quantification for methylated gene. The pubic databases including GWAS Catalog, TCGA and GEO were used. The immunohistochemistry and qRT-PCR analysis were performed. In contrast to adjacent tissues, GC tissues manifested pronounced hypermethylation patterns specifically within the promoter cytosine-phosphate-guanine (CpG) islands, indicating localized epigenetic alterations. DNA methylome analysis further revealed 4432 differentially-methylated probes (DMPs), with the gene PRKCB exhibited the most prominent average DNA methylation disparity (mean Δβ = 0.353). Pyrosequencing validation confirmed three DMPs within the PRKCB promoter (cg08406370, cg00735962, and cg18526361). Notably, the mean methylation levels of PRKCB were inversely correlated with mRNA expression levels in the GWAS Catalog. Furthermore, both mRNA and protein expression levels of PRKCB were significantly reduced in GCs when compared to their adjacent non-cancerous counterparts, verified by TCGA and GEO database. Our study reveals significant DNA methylation alterations in GC and emphasizes the pivotal role of PRKCB gene hypermethylation in conferring GC risk, which offers fresh perspectives for advancing diagnostic approaches and therapeutic strategies for GC.

DNA Methylation Signatures Characterize Gene Expression Modulation in Lung Cancer Patients Affected by Anorexia.

The pathophysiology of cancer anorexia is multifactorial and unclear. Transcriptomic analysis from PBMCs RNA showed diverse patterns of gene expression pathways in anorexic cancer patients. We assessed whether the different transcriptomic signatures are modulated by DNA methylation in lung cancer patients presenting with poor appetite. Lung cancer patients and controls were enrolled, and anorexia was assessed by the FAACT-score questionnaire. Genome-wide DNA methylation was determined by Human Infinium MethylationEPIC BeadChip Kit. Data from genome-wide methylation analysis were merged with those from gene expression analysis, previously obtained by RNA sequencing (NGS). Four groups of genes were identified for each comparison: hypermethylated repressed, hypermethylated induced, hypomethylated repressed, and hypomethylated induced. Cancer patients (n = 16) showed 382 differentially methylated genes when compared with controls (n = 8). Anorexic patients (n = 8) presented 586 hypomethylated and 174 hypermethylated genes compared with controls. In anorexic patients vs. non-anorexic (n = 8), 211 genes were identified as hypomethylated and 90 hypermethylated. When microarray methylation data were merged with transcriptomic data by RNA sequencing, we observed significant differences in anorexic patients vs. controls; a total of 42 genes resulted as hypomethylated and induced, 5 hypermethylated repressed, 10 hypermethylated induced, and 15 hypomethylated repressed. The CG sites analyzed by targeted bisulfite NGS in four genes of interest (FLNA, PGRMC1, GNL3L, and FHL1) resulting as hypomethylated in anorexic vs. controls allowed the validation of the data obtained from DNA methylation. Interestingly, the four genes resulted as hypomethylated in anorexic patients vs. non-anorexic patients and vs. controls (p < 0.0001). Our data support that methylation is implicated in cancer-associated anorexia and nutritional derangements among lung cancer patients.

5hmC-profiles in Puerto Rican Hispanic/Latino men with aggressive prostate cancer.

Puerto Rican (PR) Hispanic/Latino (H/L) men are an understudied population that has the highest prostate cancer (PCa) specific mortality among other Hispanic populations. Little information is known about the higher mortality in PR H/L men. It is thought that epigenetic changes in key genes may play a critical role in aggressive tumors. We aimed to identify key 5-hydroxymethylcytosine (5hmC) changes in PR H/L men with aggressive PCa. We performed sequencing analysis using the 5hmC-enriched DNA from 22 prostate tumors and 24 adjacent normal FFPE samples. We identified 808 differentially methylated genes (DMGs) in tumors compared to adjacent normal tissues (FDR<0.05, log2FC>|0.4|). Pathway analysis of DMGs demonstrated that DNA repair pathway was most upregulated in tumors. Since 5hmC abundance positively correlates with gene expression levels, we further investigated 808 DMGs in TCGA PCa gene expression data. Further, we identified 59 DMGs (80.1%, FDR<0.05, ΔGE (gene expression) >|1|) with significant gene expression changes in the same direction. Additionally, we identified 111 aggressiveness-related DMGs, of which, two hypomethylated genes ( CCDC122 , NUDT15 ) and four hypermethylated genes ( PVT1 , RPL30 , TRMT12 , UBR5 ) were found to be altered at transcriptomic level in a concordant manner in PR H/L PCa patients (N=86). The aberrant 5hmC (N=55) and GE (N=497) changes in these six genes were also associated with progression-free survival in the mixed PCa population. In conclusion, our study identified 59 DMGs showing concordant epigenetic and transcriptomic changes in tumor tissues and 111 DMGs showing association with aggressive PCa among PR H/L men.

GLUT1 and cerebral glucose hypometabolism in human focal cortical dysplasia is associated with hypermethylation of key glucose regulatory genes.

Focal cortical dysplasia (FCD) is recognized as a significant etiological factor in pharmacoresistant intractable epilepsy, linked with disturbances in neurovascular metabolism. Our study investigated regulation of glucose-transporter1 (GLUT1) and cerebral hypometabolism within FCD subtypes. Surgically excised human brain specimens underwent histopathological categorization. A subset of samples (paired with matching blood) was assessed for DNA methylation changes of glucose metabolism-related genes. We evaluated GLUT1, VEGFα, MCT2, and mTOR expression by western blot analysis, measured glucose-lactate concentrations, and established correlations with patients' demographic and clinical profiles. Furthermore, we investigated the impact of DNA methylation inhibitor decitabine and hypometabolic condition on the uptake of [3H]-2-deoxyglucose and ATPase in epileptic brain endothelial cells (EPI-EC). We observed hypermethylation of GLUT1 and glucose metabolic genes in FCD brain/blood samples and could distinguish FCDIIa/b from mMCD, MOGHE and non-lesional types in brain. Low GLUT1 and glucose-lactate ratios corresponded to elevated VEGFα and MCT2 in FCDIIa/b vs non-lesional tissues, independent of age, gender, seizure-onset, or duration of epilepsy. Increased mTOR signaling in FCDIIa/b tissues was evident. Decitabine stimulation increased GLUT1, decreased VEGFα expression, restored glucose uptake and ATPase activity in EPI-ECs and reduced mTOR and MCT2 levels in HEK cells. We demonstrated: 1) hypermethylation of glucose regulatory genes distinguish FCDIIa/b from mMCD, MOGHE and non-lesional types, 2) glucose uptake reduction is due to GLUT1 suppression mediated possibly by a GLUT1-mTOR mechanism; and 3) DNA methylation regulates cellular glucose update and metabolism. Together, these studies may lead to GLUT1-mediated biomarkers, glucose metabolism and identify early intervention strategies in FCD.

North India Cancer Risk: A Detailed Review with Focus on Jammu and Kashmir Demographics.

Cancer is a global medical challenge, and research is at its peak to understand the unique mechanisms of cancer cells. The expanding field of epidemiology, including molecular and environmental studies, helps us better understand the distribution of molecular changes and environmental risk factors in the population. In the present review, we aimed to find out the different genes and environmental factors that are associated with different cancers in the Jammu & Kashmir (J&K) region of the North Indian population. A Systematic approach of literature survey was used to curate research data based on genetic and environmental epidemiology specifying the J&K region. Of 640 articles found initially and screening of 490 records, 97 studies were included for the final review. It was observed that numerous genes that are strongly linked to various cancer types have been discovered as a result of the rising genotyping trend, which has grown in the demography exponentially over the last few decades. The majority of these genes are related to cell cycle regulation, cell growth signaling, and apoptosis regulation. Additionally, high promoter hypermethylation of various genes which were found to be attributed to the presence of distinct dietary patterns. The most important environmental risk attributes were salt tea consumption and dried pickles. In conclusion, the J&K population possesses many common polymorphisms in various genes with a small effect size that makes individuals more prone to different forms of cancers interacting with different environmental factors. What we can't do is, change the gene sequence or molecular changes which are the main changes for determining the susceptibility of any altered condition but what we can do is lower/ limit the exposure to the environmental factors which is a key element playing with the susceptibility's threshold. Therefore, limiting exposure to environmental factors could be a major step in lowering the risk of disease.

Exploring the effects of Merkel cell polyomavirus T antigens expression in REH and MCC13 cells by methylome and transcriptome profiling.

Merkel cell carcinoma (MCC) is a rare, aggressive skin cancer with a tripled incidence in the US and Europe over the past decade. Around 80% of MCC is linked to Merkel cell polyomavirus, but the cell of origin remains unknown. We stably introduced Merkel cell polyomavirus (MCPyV)-sT) and LT antigens to MCC13 and REH cell lines, analyzing DNA methylation and gene transcriptional regulation. Gene ontology analysis assessed MCPyV effects, and integrative analysis correlated gene expression and methylation. Expression patterns were compared with 15 previously sequenced primary MCCs. We found that MCPyV-LT induces DNA methylation changes in both cell lines, while MCPyV-sT only affected REH cells. Greater gene expression changes are observed in MCC13 cells, with upregulated genes associated with cellular components and downregulated genes related to biological processes. Integrative analysis of differentially expressed genes (DEG) and differentially methylated regions (DMR) of REH cell lines revealed that no genes were commonly methylated and differentially expressed. The study compared DEGs and DMG in MCC13 and REH cells to overlapping genes in MCPyV-positive cell lines (MKL1, MKL2, and WaGa), identifying hypomethylated genes in the gene body and hypermethylated genes at TSS1500. GO analysis of the two cell lines showed that MCPyV-TAs can downregulate genes in MHC-I pathways; this downregulation offers a target that can be used to create novel and efficient MCC immunotherapy approaches. Finally, it was confirmed that MCPyV-LT controls gene expression in MCC tissues using an integrative investigation of DNA methylation and gene expression.

Diagnostic Study Of Bladder Cancer Using Urine Methylation Biomarkers Of Iraqi Patients

Background: Bladder cancer (BC) is the 4th most frequently occurring malignancy and the 9th most common cause of death worldwide in men. The invasive and metastatic form of the cancer is the main cause of death or unfavorable prognosis for BC patients. The presence of a bladder tumor is often discovered after episodes of painless macroscopic hematuria. At initial diagnosis, the disease is non-muscle-invasive in approximately 75% of patients. In recent years, detecting aberrantly methylated DNA in urine has emerged as a promising and noninvasive approach to aid BC diagnosis. Objective: To detect bladder tumors in Iraqi patients. Methods: In this cross-sectional study, sixty cases were suspected of having bladder tumors according to the opinion of the physician and some signs and symptoms, the best sign was hematuria in urine (per diagnosis). An additional 30 healthy controls with similar age and sex were recruited as control groups, with an average age of 37_85 years. from Baghdad Medical City / Ghazi al–Hariri Teaching Hospital and AlKindi Specialized Hospital during the period from September / 2023 to April / 2024. Using molecular polymerase chain reaction (high-resolution melting-PCR). Results: Out of 60 bladder inflammations, the result of hypermethylation genes ZNF154, EOMES, and POU4F2, Sensitivity were (71.7, 85, and 75) %, and Specificity (60, 33.3, and 50) % respectively with high significant differences (p &lt; 0.01). In addition. Conclusions: Detecting DNA hypermethylation is considered a good diagnosis method for bladder tumors.

204 Changes in DNA methylation 5 days after exposure to acute heat stress in beef cattle skeletal muscle

Abstract The objective of this study was to investigate the effects of acute heat stress on the skeletal muscle epigenome using reduced representation bisulfite sequencing (RRBS). March-born crossbred (5/8 Red Angus, 3/8 Simmental) steers (n = 14; average body weight = 308.7 kg) were exposed to heat stress (HS) conditions of 35°C + 35% relative humidity (Temperature Humidity Index = 81) for 12h/d for 2 d (48 h). Longissimus dorsi samples were collected via biopsy and flash-frozen in liquid nitrogen immediately following the HS period at 48 h and from the contralateral longissimus dorsi 5 d later. DNA was isolated and underwent RRBS using 150 bp paired-end reads at an achieved average depth of 15.3 million reads/sample. Reads were trimmed and mapped to a bisulfite-converted bovine genome (ARSUCD1.2). Methylation calls were extracted and used to identify differentially methylated regions (DMRs) with methylkit in R comparing samples collected 5 d post-HS to those taken immediately post-48 h of HS. Differentially methylated regions (1,000 bp) had q &amp;lt; 0.05 and at least a 15% difference in methylation. There were 1,015 DMRs in genes. Hypomethylated DMRs 5 d post-HS included IGF1R, IGF2, IGF2BP1, IGFBP4, IGF2R, IKBKB, and IL34. Hypermethylated DMRs 5 d post-HS included FGF5, FGFR1, and FGFR3. There were 81 DMRs in putative promoter regions (2000 bp 5’ of a gene). MYLK2, SIGIRR, and 25 other genes had hypomethylated promoter regions. Genes that contained DMRs were used as input for DAVID to explore KEGG pathways. Enriched pathways (P &amp;lt; 0.1) included multiple hormone pathways, MAPK, PI3K-Akt, HIF-1, and VEGF. Hypomethylation of IGF-related genes and hypermethylation of FGF-related genes suggests impaired growth capacity post-HS, as does the enrichment of MAPK and PI3K-Akt pathways. Hypomethylation of SIGIRR’s promoter region, IL34 and IKBKB, indicates enhanced anti-inflammatory activity given their known regulation of inflammatory cytokine production. This, coupled with enrichment of the HIF-1 and VEGF pathways, suggests regulatory dampening of HS-induced oxidative stress. These findings reveal key regulatory changes in muscle growth, inflammatory response, and oxidative stress activity for cattle recovering from acute HS. Previous cattle skeletal muscle RNA studies predicted that HS altered metabolism, inflammation, and oxidative stress pathways, similar to the methylation results of this study. Overall, evidence suggests growth dysregulation, which may help explain observed HS-induced poor growth due to HS, as well as subsequent anti-inflammatory activity during recovery from HS.

Epigenetics of Dietary Phytochemicals in Cancer Prevention: Fact or Fiction.

Cancer development takes 10 to 50 years, and epigenetics plays an important role. Recent evidence suggests that ~80% of human cancers are linked to environmental factors impinging upon genetics/epigenetics. Because advanced metastasized cancers are resistant to radiation/chemotherapeutic drugs, cancer prevention by relatively nontoxic "epigenetic modifiers" will be logical. Many dietary phytochemicals possess powerful antioxidant and anti-inflammatory properties that are hallmarks of cancer prevention. Dietary phytochemicals can regulate gene expression of the cellular genome via epigenetic mechanisms. In this review, we will summarize preclinical studies that demonstrate epigenetic mechanisms of dietary phytochemicals in skin, colorectal, and prostate cancer prevention. Key examples of the importance of epigenetic regulation in carcinogenesis include hypermethylation of the NRF2 promoter region in cancer cells, resulting in inhibition of NRF2-ARE signaling. Many dietary phytochemicals demethylate NRF2 promoter region and restore NRF2 signaling. Phytochemicals can also inhibit inflammatory responses via hypermethylation of inflammation-relevant genes to block gene expression. Altogether, dietary phytochemicals are excellent candidates for cancer prevention due to their low toxicity, potent antioxidant and anti-inflammatory properties, and powerful epigenetic effects in reversing procarcinogenic events.

Hypermethylation of the BRCA2 gene promoter and its co-hypermethylation with the BRCA1 gene promoter in patients with breast cancer.

The BRCA2 gene is an important tumour suppressor in breast cancer, and alterations in BRCA2 may lead to cancer progression. The aim of the study was to investigate the association of hypermethylation of the BRCA2 gene promoter and its co-hypermethylation with the BRCA1 gene promoter with the development and course of breast cancer in women. This study included 74women with breast cancer (tumour tissue samples and peripheral blood) and 62women without oncological pathology (peripheral blood)- control group. Hypermethylation of the BRCA2 gene was significantly more frequently detected in the tumour tissue of women with breast cancer compared to their peripheral blood and peripheral blood of control subjects (p= 0.0006 and p= 0.00001, respectively). Hypermethylation of BRCA2 was more frequently detected in patients with breast cancer over the age of 50 and in patients with higher Ki67 expression levels (p= 0.045 and p= 0.045, respectively). There was a high frequency of unmethylated BRCA1 and BRCA2 gene combination in women of the control group compared to women with breast cancer, both in blood samples and tumour tissue samples (p= 0.014 and p= 0.00001, respectively). Our study confirms the hypothesis that BRCA2 hypermethylation plays an important role in the pathogenesis of breast cancer and the importance of assessing its co-hypermethylation with BRCA1 in predicting the course of the disease.

Maternal Undernutrition Affects Fetal Thymus DNA Methylation, Gene Expression, and, Thereby, Metabolism and Immunopoiesis in Wagyu (Japanese Black) Cattle.

We aimed to determine the effects of maternal nutrient restriction (MNR) on the DNA methylation and gene expression patterns associated with metabolism and immunopoiesis in the thymuses of fetal Wagyu cattle. Pregnant cows were allocated to two groups: a low-nutrition (LN; 60% nutritional requirement; n = 5) and a high-nutrition (HN; 120% nutritional requirement, n = 6) group, until 8.5 months of gestation. Whole-genome bisulfite sequencing (WGBS) and RNA sequencing were used to analyze DNA methylation and gene expression, while capillary electrophoresis-Fourier transform mass spectrometry assessed the metabolome. WGBS identified 4566 hypomethylated and 4303 hypermethylated genes in the LN group, with the intergenic regions most frequently being methylated. Pathway analysis linked hypoDMGs to Ras signaling, while hyperDMGs were associated with Hippo signaling. RNA sequencing found 94 differentially expressed genes (66 upregulated, 28 downregulated) in the LN group. The upregulated genes were tied to metabolic pathways and oxidative phosphorylation; the downregulated genes were linked to natural killer cell cytotoxicity. Key overlapping genes (GRIA1, CACNA1D, SCL25A4) were involved in cAMP signaling. The metabolomic analysis indicated an altered amino acid metabolism in the MNR fetuses. These findings suggest that MNR affects DNA methylation, gene expression, and the amino acid metabolism, impacting immune system regulation during fetal thymus development in Wagyu cattle.

Clinical value of sequential circulating tumor DNA analysis using next-generation sequencing and epigenetic modifications for guiding thermal ablation for colorectal cancer metastases: a prospective study.

While thermal ablation is now a standard treatment option for oligometastatic colorectal cancer patients, selecting those who will benefit most from locoregional therapies remains challenging. This proof-of-concept study is the first to assess the feasibility of routine testing of ctDNA before and after thermal ablation with curative intent, analyzed by next-generation sequencing (NGS) and methylation specific digital droplet PCR (ddPCR). Our prospective study primary objective was to assess the prognostic value of ctDNA before thermal ablation. This single-center prospective study from November 2021 to June 2022 included colorectal cancer patients referred for curative-intent thermal ablation. Cell-free DNA was tested at different time points by next-generation sequencing and detection of WIF1 and NPY genes hypermethylation using ddPCR. The ctDNA was considered positive if either a tumor mutation or hypermethylation was detected; recurrence-free survival was used as the primary endpoint. The study enrolled 15 patients, and a total of 60 samples were analyzed. The median follow-up after ablation was 316days, and median recurrence-free survival was 250days. CtDNA was positive for 33% of the samples collected during the first 24h. The hazard ratio for progression according to the presence of baseline circulating tumor DNA was estimated at 0.14 (CI 95%: 0.03-0.65, p = 0.019). The dynamics are provided, and patients with no recurrence were all negative at H24 for ctDNA. This study shows the feasibility of routine testing of ctDNA before and after thermal ablation with curative intent. We report that circulating tumor DNA is detectable in patients with low tumor burden using 2 techniques. This study emphasizes the potential of ctDNA for discerning patients who are likely to benefit from thermal ablation from those who may not, which could shape future referrals. The dynamics of ctDNA before and after ablation shed light on the need for further research and larger studies.

Exploring the impact of diabetes on aging: insights from TERT and COL1A1 methylation.

Aging, a multifaceted biological process, leads to diminished physical performance, especially in older adults with diabetes, where a mismatch between biological and chronological age is noticeable. Numerous studies have demonstrated that diabetes accelerates aging at the cellular and organ levels. Notable aging markers are telomerase reverse transcriptase (TERT), related to telomere length, and type 1 chain collagen (COL1A1), a key component of skin collagen. Additionally, age-related methylation increases, as revealed through methylation analysis, augmenting aspects of aging. However, the detailed interplay between aging and diabetes, particularly regarding methylation, remains underexplored and warrants further study to elucidate the biological links between the two. In this study, we elucidate the modulatory influence of diabetes on the aging process, focusing specifically on the modifications in TERT in the kidney and COL1A1 in the skin using mice of Swiss Webster strain as the diabetes model. Specimens were categorized into three distinct chronological cohorts: chronologically young (16 weeks; n = 5), chronologically old (40 weeks; n = 5), and a periodically assessed group (16 weeks; n = 30), from which five mice were systematically sacrificed on a weekly basis. Our findings reveal a marked impact of diabetes on the methylation statuses of TERT and COL1A1, characterized by an elevation in methylation levels within the periodic group (1st-6th week) and a simultaneous, progressive attenuation in the expression of TERT and COL1A1 genes. The observed alterations in the methylation levels of TERT and COL1A1 propound the hypothesis that diabetes potentially expedites the aging process, concomitantly impinging on the production of TERT and COL1A, ostensibly through the mechanism of promoter gene hypermethylation.

Abstract We145: δ-catenin mRNA m 6 A Methylation Promotes Cardiac Fibrosis following Acute Myocardial Infarction

Background: Cardiac fibrosis is pivotal in heart failure progression, where excessive extracellular matrix (ECM) secretion by activated fibroblasts leads to adverse remodeling and dysfunction. While Wnt/β-catenin signaling influences fibroblast activation and cardiac fibrosis post-MI, its precise regulation remains unclear. Emerging evidence suggests N6-methyladenosine (m 6 A) mRNA methylation's role in disease pathology, yet its specific contribution to post-MI cardiac fibrosis is not well understood. Thus, we hypothesized that “ MI-induced METTL3 (a Key m6A mRNA methyltransferase) activation stabilizes δ-catenin mRNA, facilitating cardiac fibrosis and adverse remodeling ”. Methods: The mice underwent sham/MI surgeries for 4 weeks, after which heart tissues were collected for biochemical and histological analysis following heart function measurements. Additionally, methyl-immunoprecipitation followed by RNA sequencing (MeRIP-sequencing) was performed on the heart tissue post-surgery, and the data were analyzed to identify fibrosis-associated targets. Results: Ischemic injury significant increase in m6A mRNA methylation in heart tissues. This increase in m 6 A RNA was also observed in adult cardiac fibroblasts (AMFs) following TGFb treatment. Interestingly, METTL3 inhibition (METTL3 siRNA) resulted in a significant reduction in TGFb-induced periostin and fibronectin gene expression, while METTL3 overexpression enhanced the expression of fibrotic genes in AMFs. To identify differentially regulated m6A target genes, MeRIP-seq was performed on RNA isolated from mice hearts post-MI. The sequencing data suggested hypermethylation of fibrosis-associated genes, including δ-catenin, post-MI. Notably, TGFb-induced increased δ-catenin mRNA methylation led to δ-catenin mRNA stabilization. In contrast, METTL3 inhibition using siMETTL3 in AMFs and in METTL3 KO mice significantly reduced fibroblast activation and cardiac fibrosis. Conclusion: Our data suggests that hypermethylation of δ-catenin mRNA plays a significant role in the progression of cardiac fibrosis following AMI. Therefore, regulating METTL3 could be a potential therapeutic target for attenuating cardiac fibrosis.

Hypermethylation of genes on chromosome 3p as a biomarker for nasopharyngeal carcinoma diagnosis: A Vietnamese case-control study.

The crucial event driving nasopharyngeal tumorigenesis is the hypermethylation of chromosome 3p-located tumor suppressor genes. This case-control study aims to investigate the methylation characteristics of RASSF1A, Blu, ADAMTS9, and DLEC1 to potentially develop effective diagnostic biomarkers for nasopharyngeal carcinoma, either individually or in combination. The methylation of RASSF1A, Blu, ADAMTS9, and DLEC1 in the collection of 93 biopsy samples and 100 healthy swab specimens were evaluated by Nested methylation-specific polymerase chain reaction. The strength of the correlation between candidate genes and nasopharyngeal carcinoma was estimated by the evaluation of odds ratios (ORs). Promoter hypermethylation of RASSF1A, Blu, ADAMTS9, and DLEC1 were found in 60.22%, 80.65%, 62.37%, and 74.19%, respectively, in nasopharyngeal carcinoma tumors. A significant association between the methylation status of candidate genes with nasopharyngeal carcinoma was reported. The methylation of candidate genes significantly increased the risk of nasopharyngeal carcinoma in cancerous samples compared with control samples (OR > 1). Based on the value of the methylation index, methylation of at least one gene was found in 95.70% of nasopharyngeal tumors. Additionally, the methylation index among 93 tumors significantly correlated with advanced stage nasopharyngeal tumors. The study explored a higher frequency of hypermethylation at least one candidate gene. Methylation of a panel of potential genes can be utilized to discriminate between nasopharyngeal carcinoma and non-cancer cells, particularly in the advanced stages of nasopharyngeal carcinoma. Thus, it could serve as a valuable marker for the diagnosis and monitoring of nasopharyngeal carcinoma.