Rosa canina extract relieves methylation alterations of pancreatic genes in STZ-induced diabetic rats : Gene methylation in diabetic rats treated with an extract.

BACKGROUND: Diabetes is a chronic metabolic disease that affects many parts of the body. Considering diabetes as a beta cells' defect and loss, the focus is on finding mechanisms and compounds involved in stimulating the function and regeneration of pancreatic ß-cells. DNA methylation as an epigenetic mechanism plays a pivotal role in the ß-cells' function and development. Considering the regenerative and anti-diabetic effects of Rosa canina extract, this study aimed to assess the methylation levels of Pdx-1, Pax-4, and Ins-1 genes in diabetic rats treated with Rosa Canina extract. METHODS AND RESULTS: Streptozotocin-induced diabetic rats were used to evaluate the frequency of Pdx-1, Pax-4, and Ins-1 gene methylation. Treatment groups were exposed to Rosa canina as spray-dried and decoction extracts. Following blood glucose measurement, pancreatic DNA was extracted and bisulfited. Genes' methylation was measured using MSP-PCR and qRT-PCR techniques. Oral administration of Rosa canina extracts significantly reduced blood sugar levels in diabetic rats compared to the control group. The methylation levels of the Pdx-1, Pax-4, and Ins-1 genes promoter in streptozotocin-induced diabetic rats increased compared to the control rats while, the treatment of diabetic rats with Rosa canina extracts, spray-dried samples especially, led to a decreased methylation in these genes. CONCLUSION: The results of this study showed that Rosa canina extract as a spray-dried sample could be effective in treating diabetes by regulating the methylation of genes including Pdx-1, Pax-4, and Ins-1 involved in the activity and regeneration of pancreatic islet cells.

Dapagliflozin suppresses diabetes-induced oxidative DNA damage and hypermethylation in mouse somatic cells.

Diabetes mellitus is a complex metabolic disorder resulting from the interplay of environmental, genetic, and epigenetic factors that increase the risk of cancer development. However, it is unclear whether the increased cancer risk is due to poor glycemic control or the use of some antidiabetic medications. Therefore, we investigated the genetic and epigenetic changes in somatic cells in a mouse model of diabetes and studied whether multiple exposures to the antidiabetic medication dapagliflozin influence these changes. We also elucidated the mechanism(s) of these ameliorations. The micronucleus test and modified comet assay were used to investigate bone marrow DNA damage and methylation changes. These assays revealed that dapagliflozin is non-genotoxic in the tested regimen, and oxidative DNA damage and hypermethylation were significantly higher in diabetic mice. Spectrophotometry also evaluated oxidative DNA damage and global DNA methylation, revealing similar significant alterations induced by diabetes. Conversely, the dapagliflozin-treated diabetic animals significantly reduced these changes. The expression of some genes involved in DNA repair and DNA methylation was disrupted considerably in the somatic cells of diabetic animals. In contrast, dapagliflozin treatment significantly restored these disruptions and enhanced DNA repair. The simultaneous effects of decreased oxidative DNA damage and hypermethylation levels suggest that dapagliflozin can be used as a safe antidiabetic drug to reduce DNA damage and hypermethylation in diabetes, demonstrating its usefulness in patients with diabetes to control hyperglycemia and decrease the development of its subsequent complications.

Signatures of epigenetic, biological and mitotic age acceleration and telomere shortening are associated with arsenic-induced skin lesions.

Chronic arsenic exposure is a global health hazard significantly associated with the development of deleterious cutaneous changes and increased keratinocyte cancer risk. Although arsenic exposure is associated with broad-scale cellular and molecular changes, gaps exist in understanding how these changes impact the skin and facilitate malignant transformation. Recently developed epigenetic "clocks" can accurately predict chronological, biological and mitotic age, as well as telomere length, on the basis of tissue DNA methylation state. Deviations of predicted from expected age (epigenetic age dysregulation) have been associated with numerous complex diseases, increased all-cause mortality and higher cancer risk. We investigated the ability of these algorithms to detect molecular changes associated with chronic arsenic exposure in the context of associated skin lesions. To accomplish this, we utilized a multi-algorithmic approach incorporating seven "clocks" (Horvath, Skin&Blood, PhenoAge, PCPhenoAge, GrimAge, DNAmTL and epiTOC2) to analyze peripheral blood of pediatric and adult cohorts of arsenic-exposed (n = 84) and arsenic-naïve (n = 33) individuals, among whom n = 18 were affected by skin lesions. Arsenic-exposed adults with skin lesions exhibited accelerated epigenetic (Skin&Blood: + 7.0 years [95% CI 3.7; 10.2], q = 6.8 × 10-4), biological (PhenoAge: + 5.8 years [95% CI 0.7; 11.0], q = 7.4 × 10-2, p = 2.8 × 10-2) and mitotic age (epiTOC2: + 19.7 annual cell divisions [95% CI 1.8; 37.7], q = 7.4 × 10-2, p = 3.2 × 10-2) compared to healthy arsenic-naïve individuals; and accelerated epigenetic age (Skin&Blood: + 2.8 years [95% CI 0.2; 5.3], q = 2.4 × 10-1, p = 3.4 × 10-2) compared to lesion-free arsenic-exposed individuals. Moreover, lesion-free exposed adults exhibited accelerated Skin&Blood age (+ 4.2 [95% CI 1.3; 7.1], q = 3.8 × 10-2) compared to their arsenic-naïve counterparts. Compared to the pediatric group, arsenic-exposed adults exhibited accelerated epigenetic (+ 3.1 to 4.4 years (95% CI 1.2; 6.4], q = 2.4 × 10-4-3.1 × 10-3), biological (+ 7.4 to 7.8 years [95% CI 3.0; 12.1] q = 1.6 × 10-3-2.8 × 10-3) and mitotic age (+ 50.0 annual cell divisions [95% CI 15.6; 84.5], q = 7.8 × 10-3), as well as shortened telomere length (- 0.23 kilobases [95% CI - 0.13; - 0.33], q = 2.4 × 10-4), across all seven algorithms. We demonstrate that lifetime arsenic exposure and presence of arsenic-associated skin lesions are associated with accelerated epigenetic, biological and mitotic age, and shortened telomere length, reflecting altered immune signaling and genomic regulation. Our findings highlight the usefulness of DNA methylation-based algorithms in identifying deleterious molecular changes associated with chronic exposure to the heavy metal, serving as potential prognosticators of arsenic-induced cutaneous malignancy.

Sulfonamide-induced DNA hypomethylation disturbed sugar metabolism in rice (Oryza sativa L.).

DNA methylation is well-accepted as a bridge to unravel the complex interplay between genome and environmental exposures, and its alteration regulated the cellular metabolic responses towards pollutants. However, the mechanism underlying site-specific aberrant DNA methylation and metabolic disorders under pollutant stresses remained elusive. Herein, the multilevel omics interferences of sulfonamides (i.e., sulfadiazine and sulfamerazine), a group of antibiotics pervasive in farmland soils, towards rice in 14 days of 1 mg/L hydroponic exposure were systematically evaluated. Metabolome and transcriptome analyses showed that 57.1-71.4 % of mono- and disaccharides were accumulated, and the differentially expressed genes were involved in the promotion of sugar hydrolysis, as well as the detoxification of sulfonamides. Most differentially methylated regions (DMRs) were hypomethylated ones (accounting for 87-95 %), and 92 % of which were located in the CHH context (H = A, C, or T base). KEGG enrichment analysis revealed that CHH-DMRs in the promoter regions were enriched in sugar metabolism. To reveal the significant hypomethylation of CHH, multi-spectroscopic and thermodynamic approaches, combined with molecular simulation were conducted to investigate the molecular interaction between sulfonamides and DNA in different sequence contexts, and the result demonstrated that sulfonamides would insert into the minor grooves of DNA, and exhibited a stronger affinity with the CHH contexts of DNA compared to CG or CHG contexts. Computational modeling of DNA 3D structures further confirmed that the binding led to a pitch increase of 0.1 Å and a 3.8° decrease in the twist angle of DNA in the CHH context. This specific interaction and the downregulation of methyltransferase CMT2 (log2FC = -4.04) inhibited the DNA methylation. These results indicated that DNA methylation-based assessment was useful for metabolic toxicity prediction and health risk assessment.

A pharmacodynamic assay to monitor treatment with the hypomethylating cytosine analogs, decitabine and azacitidine.

Hypomethylating therapies using decitabine or azacitidine are actively investigated to treat acute myeloid leukemia, myelodysplastic syndromes, as maintenance therapy after allogenic stem cell transplant and hemoglobinopathies. The therapeutic mechanism is to de-repress genes that have been turned off through oncogenesis or development via methylation. The therapy can be non-cytotoxic at low dosage, sparing healthy stem cells and operating on committed precursors. Because the methods of determining maximum tolerated dose are not well suited to this paradigm, and because the mechanism of action, which is depletion of DNA methylase 1 (DNMT1), is complex and dependent on passing through a cell cycle, a pharmacodynamic assay that measures DNMT1 can inform clinical trials aimed at establishing and improving therapy. Herein, we provide an assay that measures DNMT1 relative levels in circulating T cells of peripheral blood.

Distinct epigenetic modulation of differentially expressed genes in the adult mouse brain following prenatal exposure to low-dose bisphenol A.

Bisphenol A (BPA) is a common component in the manufacture of daily plastic consumer goods. Recent studies have suggested that prenatal exposure to BPA can increase the susceptibility of offspring to mental illness, although the underlying mechanisms remain unclear. In this study, we performed transcriptomic and epigenomic profiling in the adult mouse brain following prenatal exposure to low-dose BPA. We observed a sex-specific transcriptional dysregulation in the cortex, with more significant differentially expressed genes was observed in adult cortex from male offspring. Moreover, the upregulated genes primarily influenced neuronal functions, while the downregulated genes were significantly associated with energy metabolism pathways. More evidence supporting impaired mitochondrial function included a decreased ATP level and a reduced number of mitochondria in the cortical neuron of the BPA group. We further investigated the higher-order chromatin regulatory patterns of DEGs by incorporating published Hi-C data. Interestingly, we found that upregulated genes exhibited more distal interactions with multiple enhancers, while downregulated genes displayed relatively short-range interactions among adjacent genes. Our data further revealed decreased H3K9me3 signal on the distal enhancers of upregulated genes, whereas increased DNA methylation and H3K27me3 signals on the promoters of downregulated genes. In summary, our study provides compelling evidence for the potential health risks associated with prenatal exposure to BPA, and uncovers sex-specific transcriptional changes with a complex interplay of multiple epigenetic mechanisms.

Prenatal exposure to low-dose bisphenol A disrupts hippocampal DNA methylation and demethylation in male rat offspring.

Earlier research has demonstrated that developmental exposure to bisphenol A (BPA) has persistent impacts on both adult brain growth and actions. It has been suggested that BPA might obstruct the methylation coding of the genes in the brain. In this study, the methylation changes in the hippocampus tissue of male rat pups were examined following prenatal BPA exposure. Pregnant Sprague-Dawley rats were treated with either vehicle (tocopherol-stripped corn oil) or BPA (4, 40, or 400 µg/kg·body weight/day) throughout the entire duration of gestation and lactation. At 3 weeks of age, the male rat offspring were euthanized, and the hippocampus were dissected out for analysis. The expression levels of DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B) and DNA demethylases (TET1, Gadd45a, Gadd45b, and Apobec1) were analyzed in the hippocampus by means of quantitative real-time polymerase chain reaction and Western blotting, respectively. The results showed that prenatal exposure to BPA upregulated the expression of enzymes associated with DNA methylation and demethylation processes in the hippocampus of male rat offspring. These findings suggest that prenatal exposure to a low dose of BPA could potentially disrupt the balance of methylation and demethylation in the hippocampus, thereby perturbing epigenetic modifications. This may represent a neurotoxicity mechanism of BPA.

Prenatal nicotine exposure leads to epigenetic alterations in peripheral nervous system signaling genes in the testis of the rat.

BACKGROUND: Prenatal nicotine exposure (PNE) has been documented to cause numerous deleterious effects on fetal development. However, the epigenetic changes promoted by nicotine exposure on germ cells are still not well understood. OBJECTIVES: In this study, we focused on elucidating the impact of prenatal nicotine exposure on regulatory epigenetic mechanisms important for germ cell development. METHODS: Sprague-Dawley rats were exposed to nicotine during pregnancy and male progeny was analyzed at 11 weeks of age. Testis morphology was analyzed using frozen testis sections and expression of germ cell markers was examined by RT-qPCR; histone modifications were assessed by Western Blot (WB). DNA methylation analysis was performed by methylation-specific PCR of bisulfite converted DNA. Genome-wide DNA methylation was analyzed using Methylated DNA immunoprecipitation (MeDIP)-seq. We also carried out transcriptomics analysis of pituitary glands by RNA-seq. RESULTS: We show that gestational exposure to nicotine reduces germ cell numbers, perturbs meiosis, affects the expression of germ line reprogramming responsive genes, and impacts the DNA methylation of nervous system genes in the testis. PNE also causes perturbation of gene expression in the pituitary gland of the brain. CONCLUSIONS: Our data demonstrate that PNE leads to perturbation of male spermatogenesis, and the observed effects are associated with changes of peripheral nervous system signaling pathways. Alterations in the expression of genes associated with diverse biological activities such as cell migration, cell adhesion and GABA signaling in the pituitary gland underscore the complexity of the effects of nicotine exposure during pregnancy.

Targeting Aging and Longevity with Exogenous Nucleotides (TALENTs): Rationale, Design, and Baseline Characteristics from a Randomized Controlled Trial in Older Adults.

Nucleotides (NTs), important biomolecules involved in numerous cellular processes, have been proposed as potential candidates for anti-aging interventions. However, whether nucleotides can act as an anti-aging supplement in older adults remains unclear. TALENTs is a randomized, double-blinded, placebo-controlled trial that evaluates the efficacy and safety of NTs as an anti-aging supplement in older adults by exploring the effects of NTs on multiple dimensions of aging in a rigorous scientific setting. Eligible community-dwelling adults aged 60-70 years were randomly assigned equally to two groups: nucleotides intervention group and placebo control group. Comprehensive geriatric health assessments were performed at baseline, 2-months, and 4-months of the intervention. Biological specimens were collected and stored for age-related biomarker testing and multi-omics sequencing. The primary outcome was the change from baseline to 4 months on leukocyte telomere length and DNA methylation age. The secondary aims were the changes in possible mechanisms underlying aging processes (immunity, inflammatory profile, oxidative stress, gene stability, endocrine, metabolism, and cardiovascular function). Other outcomes were changes in physical function, body composition and geriatric health assessment (including sleep quality, cognitive function, fatigue, frailty, and psychology). In the RCT, 301 participants were assessed for eligibility and 122 were enrolled. Participants averaged 65.65 years of age, and were predominately female (67.21%). All baseline characteristics were well-balanced between groups, as expected due to randomization. The majority of participants were pre-frailty and had at least one chronic condition. The mean scores for physical activity, psychological, fatigue and quality of life were within the normal range. However, nearly half of the participants still had room for improvement in cognitive level and sleep quality. This TALENTs trial will represent one of the most comprehensive experimental clinical trials in which supplements are administered to elderly participants. The findings of this study will contribute to our understanding of the anti-aging effects of NTs and provide insights into their potential applications in geriatric healthcare.

Effects of different extenders on epigenetic patterns and functional parameters of bull sperm during cryopreservation process.

The cryopreservation process induces alterations in cellular parameters and epigenetic patterns in bull sperm, which can be prevented by adding cryoprotectants in the freezing extenders. The purpose of this study was to compare the protective effects of two extenders based on soybean lecithin (SLE) and egg yolk (EYE) on epigenetic patterns and quality parameters of sperm such as motility parameters, mitochondrial membrane integrity, DNA fragmentation, viability, and apoptotic-like changes of bull sperm after cryopreservation. Results demonstrated that cryopreservation significantly (p < .05) reduced the level of DNA global methylation, H3K9 histone acetylation, and H3K4 histone methylation in both frozen groups compared to the fresh sperm. Also, the level of H3K9 acetylation was lower in the frozen SLE group (21.2 ± 1.86) compared to EYE group (15.2 ± 1.86). In addition, the SLE frozen group had a higher percentage of viability, progressive motility, and linearity (LIN) in SLE frozen group compared to EYE frozen group. However, no difference was observed in mitochondrial membrane integrity and DNA fragmentation between SLE and EYE frozen groups. While soybean-lecithin-based extender showed some initial positive impacts of epigenetics and semen parameters, further investigations can provide useful information for better freezing.

Analysis of the nischarin expression across human tumor types reveals its context-dependent role and a potential as a target for drug repurposing in oncology.

Nischarin was reported to be a tumor suppressor that plays a critical role in breast cancer initiation and progression, and a positive prognostic marker in breast, ovarian and lung cancers. Our group has found that nischarin had positive prognostic value in female melanoma patients, but negative in males. This opened up a question whether nischarin has tumor type-specific and sex-dependent roles in cancer progression. In this study, we systematically examined in the public databases the prognostic value of nischarin in solid tumors, regulation of its expression and associated signaling pathways. We also tested the effects of a nischarin agonist rilmenidine on cancer cell viability in vitro. Nischarin expression was decreased in tumors compared to the respective healthy tissues, most commonly due to the deletions of the nischarin gene and promoter methylation. Unlike in healthy tissues where it was located in the cytoplasm and at the membrane, in tumor tissues nischarin could also be observed in the nuclei, implying that nuclear translocation may also account for its cancer-specific role. Surprisingly, in several cancer types high nischarin expression was a negative prognostic marker. Gene set enrichment analysis showed that in tumors in which high nischarin expression was a negative prognostic marker, signaling pathways that regulate stemness were enriched. In concordance with the findings that nischarin expression was negatively associated with pathways that control cancer growth and progression, nischarin agonist rilmenidine decreased the viability of cancer cells in vitro. Taken together, our study lays a ground for functional studies of nischarin in a context-dependent manner and, given that nischarin has several clinically approved agonists, provides rationale for their repurposing, at least in tumors in which nischarin is predicted to be a positive prognostic marker.

Betaine alleviates spermatogenic cells apoptosis of oligoasthenozoospermia rat model by up-regulating methyltransferases and affecting DNA methylation.

BACKGROUND: Oligoasthenozoospermia is the most common type of semen abnormality in male infertile patients. Betaine (BET) has been proved to have pharmacological effects on improving semen quality. BET also belongs to endogenous physiological active substances in the testis. However, the physiological function of BET in rat testis and its pharmacological mechanism against oligoasthenozoospermia remain unclear. PURPOSE: This research aims to prove the therapeutic effect and potential mechanism of BET on oligoasthenozoospermia rat model induced by Tripterygium wilfordii glycosides (TWGs). METHODS: The oligoasthenozoospermia rat model was established by a continuous gavage of TWGs (60 mg/kg) for 28 days. Negative control group, oligoasthenozoospermia group, positive drug group (levocarnitine, 300 mg/kg), and 200 mg/kg, 400 mg/kg, and 800 mg/kg BET groups were created for exploring the therapeutic effect of BET on the oligoasthenozoospermia rat model. The therapeutic effect was evaluated by HE and TUNEL staining. Immunofluorescence assay of DNMT3A, PIWIL1, PRMT5, SETDB1, BHMT2, and METTL3, methylation capture sequencing, Pi-RNA sequencing, and molecular docking were used to elucidate potential pharmacological mechanisms. RESULTS: It is proved that BET can significantly restore testicular pathological damage induced by TWGs, which also can significantly reverse the apoptosis of spermatogenic cells. The spermatogenic cell protein expression levels of DNMT3A, PIWIL1, PRMT5, SETDB1, BHMT2, and METTL3 significantly decreased in oligoasthenozoospermia group. 400 mg/kg and 800 mg/kg BET groups can significantly increase expression level of the above-mentioned proteins. Methylation capture sequencing showed that BET can significantly increase the 5mC methylation level of Spata, Spag, and Specc spermatogenesis-related genes. Pi-RNA sequencing proved that the above-mentioned genes produce a large number of Pi-RNA under BET intervention. Pi-RNA can form complexes with PIWI proteins to participate in DNA methylation of target genes. Molecular docking indicated that BET may not directly act as substrate for methyltransferase and instead participates in DNA methylation by promoting the methionine cycle and increasing S-adenosylmethionine synthesis. CONCLUSION: BET has a significant therapeutic effect on oligoasthenozoospermia rat model induced by TWPs. The mechanism mainly involves that BET can increase the methylation level of Spata, Specc, and Spag target genes through the PIWI/Pi-RNA pathway and up-regulation of methyltransferases (including DNA methyltransferases and histone methyltransferases).

Arsenic disulfide promoted the demethylation of PTPL1 in diffuse large B cell lymphoma cells.

Background: Promoter hypermethylation of the tumor suppressor gene is one of the well-studied causes of cancer development. The drugs that reverse the process by driving demethylation could be a candidate for anticancer therapy. This study was designed to investigate the effects of arsenic disulfide on PTPL1 methylation in diffuse large B cell lymphoma (DLBCL). Methods: We knocked down the expression of PTPL1 in two DLBCL cell lines (i.e., DB and SU-DHL-4 cells) using siRNA. Then the DLBCL proliferation was determined in the presence of PTPL1 knockdown. The methylation of PTPL1 in DLBCL cells was analyzed by methylation specific PCR (MSPCR). The effect of arsenic disulfide on the PTPL1 methylation was determined in DLBCL cell lines in the presence of different concentrations of arsenic disulfide (5 µM, 10 µM and 20 µM), respectively. To investigate the potential mechanism on the arsenic disulfide-mediated methylation, the mRNA expression of DNMT1, DNMT3B and MBD2 was determined. Results: PTPL1 functioned as a tumor suppressor gene in DLBCL cells, which was featured by the fact that PTPL1 knockdown promoted the proliferation of DLBCL cells. PTPL1 was found hypermethylated in DLBCL cells. Arsenic disulfide promoted the PTPL1 demethylation in a dose-dependent manner, which was related to the inhibition of DNMTs and the increase of MBD2. Conclusion: Experimental evidence shows that PTPL1 functions as a tumor suppressor gene in DLBCL progression. PTPL1 hyper-methylation could be reversed by arsenic disulfide in a dose-dependent manner.

Long-Term Epigenetic Regulation of Foxo3 Expression in Neonatal Valproate-Exposed Rat Hippocampus with Sex-Related Differences.

Perinatal exposure to valproic acid is commonly used for autism spectrum disorder (ASD) animal model development. The inhibition of histone deacetylases by VPA has been proposed to induce epigenetic changes during neurodevelopment, but the specific alterations in genetic expression underlying ASD-like behavioral changes remain unclear. We used qPCR-based gene expression and epigenetics tools and Western blotting in the hippocampi of neonatal valproic acid-exposed animals at 4 weeks of age and conducted the social interaction test to detect behavioral changes. Significant alterations in gene expression were observed in males, particularly concerning mRNA expression of Foxo3, which was significantly associated with behavioral changes. Moreover, notable differences were observed in H3K27ac chromatin immunoprecipitation, quantitative PCR (ChIP-qPCR), and methylation-sensitive restriction enzyme-based qPCR targeting the Foxo3 gene promoter region. These findings provide evidence that epigenetically regulated hippocampal Foxo3 expression may influence social interaction-related behavioral changes. Furthermore, identifying sex-specific gene expression and epigenetic changes in this model may elucidate the sex disparity observed in autism spectrum disorder prevalence.

Enhanced ALOX12 Gene Expression Predicts Therapeutic Susceptibility to 5-Azacytidine in Patients with Myelodysplastic Syndromes.

5-azacytidine (AZA), a representative DNA-demethylating drug, has been widely used to treat myelodysplastic syndromes (MDS). However, it remains unclear whether AZA's DNA demethylation of any specific gene is correlated with clinical responses to AZA. In this study, we investigated genes that could contribute to the development of evidence-based epigenetic therapeutics with AZA. A DNA microarray identified that AZA specifically upregulated the expression of 438 genes in AZA-sensitive MDS-L cells but not in AZA-resistant counterpart MDS-L/CDA cells. Of these 438 genes, the ALOX12 gene was hypermethylated in MDS-L cells but not in MDS-L/CDA cells. In addition, we further found that (1) the ALOX12 gene was hypermethylated in patients with MDS compared to healthy controls; (2) MDS classes with excess blasts showed a relatively lower expression of ALOX12 than other classes; (3) a lower expression of ALOX12 correlated with higher bone marrow blasts and a shorter survival in patients with MDS; and (4) an increased ALOX12 expression after AZA treatment was associated with a favorable response to AZA treatment. Taking these factors together, an enhanced expression of the ALOX12 gene may predict favorable therapeutic responses to AZA therapy in MDS.

Gene Expression and DNA Methylation Profiling Suggest Potential Biomarkers for Azacitidine Resistance in Myelodysplastic Syndrome.

Myelodysplastic syndrome/neoplasm (MDS) comprises a group of heterogeneous hematopoietic disorders that present with genetic mutations and/or cytogenetic changes and, in the advanced stage, exhibit wide-ranging gene hypermethylation. Patients with higher-risk MDS are typically treated with repeated cycles of hypomethylating agents, such as azacitidine. However, some patients fail to respond to this therapy, and fewer than 50% show hematologic improvement. In this context, we focused on the potential use of epigenetic data in clinical management to aid in diagnostic and therapeutic decision-making. First, we used the F-36P MDS cell line to establish an azacitidine-resistant F-36P cell line. We performed expression profiling of azacitidine-resistant and parental F-36P cells and used biological and bioinformatics approaches to analyze candidate azacitidine-resistance-related genes and pathways. Eighty candidate genes were identified and found to encode proteins previously linked to cancer, chronic myeloid leukemia, and transcriptional misregulation in cancer. Interestingly, 24 of the candidate genes had promoter methylation patterns that were inversely correlated with azacitidine resistance, suggesting that DNA methylation status may contribute to azacitidine resistance. In particular, the DNA methylation status and/or mRNA expression levels of the four genes (AMER1, HSPA2, NCX1, and TNFRSF10C) may contribute to the clinical effects of azacitidine in MDS. Our study provides information on azacitidine resistance diagnostic genes in MDS patients, which can be of great help in monitoring the effectiveness of treatment in progressing azacitidine treatment for newly diagnosed MDS patients.

Epigenetic Activation of Cytochrome P450 1A2 Sensitizes Hepatocellular Carcinoma Cells to Sorafenib.

Cytochrome P450 1A2 (CYP1A2) is a known tumor suppressor in hepatocellular carcinoma (HCC), but its expression is repressed in HCC and the underlying mechanism is unclear. In this study, we investigated the epigenetic mechanisms of CYP1A2 repression and potential therapeutic implications. In HCC tumor tissues, the methylation rates of CYP1A2 CpG island (CGI) and DNA methyltransferase (DNMT) 3A protein levels were significantly higher, and there was a clear negative correlation between DNMT3A and CYP1A2 protein expression. Knockdown of DNMT3A by siRNA significantly increased CYP1A2 expression in HCC cells. Additionally, treating HCC cells with decitabine (DAC) resulted in a dose-dependent upregulation of CYP1A2 expression by reducing the methylation level of CYP1A2 CGI. Furthermore, we observed a decreased enrichment of H3K27Ac in the promoter region of CYP1A2 in HCC tissues. Treatment with the trichostatin A (TSA) restored CYP1A2 expression in HCC cells by increasing H3K27Ac levels in the CYP1A2 promoter region. Importantly, combination treatment of sorafenib with DAC or TSA resulted in a leftward shift of the dose-response curve, lower IC50 values, and reduced colony numbers in HCC cells. Our findings suggest that hypermethylation of the CGI at the promoter, mediated by the high expression of DNMT3A, and hypoacetylation of H3K27 in the CYP1A2 promoter region, leads to CYP1A2 repression in HCC. Epigenetic drugs DAC and TSA increase HCC cell sensitivity to sorafenib by restoring CYP1A2 expression. Our study provides new insights into the epigenetic regulation of CYP1A2 in HCC and highlights the potential of epigenetic drugs as a therapeutic approach for HCC. SIGNIFICANCE STATEMENT: This study marks the first exploration of the epigenetic mechanisms underlying cytochrome P450 (CYP) 1A2 suppression in hepatocellular carcinoma (HCC). Our findings reveal that heightened DNA methyltransferase expression induces hypermethylation of the CpG island at the promoter, coupled with diminished H3K27Ac levels, resulting in the repression of CYP1A2 in HCC. The use of epigenetic drugs such as decitabine and trichostatin A emerges as a novel therapeutic avenue, demonstrating their potential to restore CYP1A2 expression and enhance sorafenib sensitivity in HCC cells.

Decreased Treg cells induced by bisphenol A is associated with up-regulation of PI3K/Akt/mTOR signaling pathway and Foxp3 DNA methylation in spleen of adolescent mice.

The current study aimed to explore whether bisphenol A (BPA) exposure aggravated the decrease in Tregs induced by ovalbumin (OVA) in adolescent female mouse models of asthma, and whether the process was associated with mTOR-mediated signaling pathways and DNA methylation levels. A total of 40 female C57BL/6 mice at the age of four weeks were used and divided into five groups after 1 week of domestication. Each group consisted of eight mice: the control group, OVA group, OVA + BPA (0.1 µg mL-1) group, OVA + BPA (0.2 µg mL-1) group, and OVA + BPA (0.4 µg mL-1) group. Results revealed that Foxp3 protein levels decreased in the spleens of mice exposed to BPA compared to those in the OVA group. After an elevation in BPA dose, the mRNAs of methyltransferases (Dnmt1, Dnmt3a, and Dnmt3b) were gradually upregulated. The mechanism was related to the activity of TLR4/NF-κB and PI3K/Akt/mTOR signaling pathways and the enhancement of Foxp3 DNA methylation. Our results, collectively, provided a new view for studying the mechanisms underlying BPA exposure-induced immune dysfunction. Investigation of the regulatory mechanisms of DNA methylation in the abnormal Th immune response caused by BPA exposure could help reveal the causes and molecular mechanisms underlying the high incidence of allergic diseases in children in recent years.

Specific CpG sites methylation is associated with hematotoxicity in low-dose benzene-exposed workers.

Benzene is a broadly used industrial chemicals which causes various hematologic abnormalities in human. Altered DNA methylation has been proposed as epigenetic biomarkers in health risk evaluation of benzene exposure, yet the role of methylation at specific CpG sites in predicting hematological effects remains unclear. In this study, we recruited 120 low-level benzene-exposed and 101 control male workers from a petrochemical factory in Maoming City, Guangdong Province, China. Urinary S-phenylmercapturic acid (SPMA) in benzene-exposed workers was 3.40-fold higher than that in control workers (P < 0.001). Benzene-induced hematotoxicity was characterized by reduced white blood cells counts and nuclear division index (NDI), along with an increased DNA damage and urinary 8-hydroxy-2'-deoxyguanosine (all P < 0.05). Methylation levels of TRIM36, MGMT and RASSF1a genes in peripheral blood lymphocytes (PBLCs) were quantified by pyrosequencing. CpG site 6 of TRIM36, CpG site 2, 4, 6 of RASSF1a and CpG site 1, 3 of MGMT methylation were recognized as hot CpG sites due to a strong correlation with both internal exposure and hematological effects. Notably, integrating hot CpG sites methylation of multiple genes reveal a higher efficiency in prediction of integrative damage compared to individual genes at hot CpG sites. The negative dose-response relationship between the combined methylation of hot CpG sites in three genes and integrative damage enabled the classification of benzene-exposed individuals into high-risk or low-risk groups using the median cut-off value of the integrative index. Subsequently, a prediction model for integrative damage in benzene-exposed populations was built based on the methylation status of the identified hot CpG sites in the three genes. Taken together, these findings provide a novel insight into application prospect of specific CpG site methylation as epi-biomarkers for health risk assessment of environmental pollutants.

Genome-wide profiling of DNA methylome and transcriptome reveals epigenetic regulation of Urechis unicinctus response to sulfide stress.

Sulfide is a well-known environmental pollutant that can have detrimental effects on most organisms. However, few metazoans living in sulfide-rich environments have developed mechanisms to tolerate and adapt to sulfide stress. Epigenetic mechanisms, including DNA methylation, have been shown to play a vital role in environmental stress adaptation. Nevertheless, the precise function of DNA methylation in biological sulfide adaptation remains unclear. Urechis unicinctus, a benthic organism inhabiting sulfide-rich intertidal environments, is an ideal model organism for studying adaptation to sulfide environments. In this study, we conducted a comprehensive analysis of the DNA methylome and transcriptome of U. unicinctus after exposure to 50 µM sulfide. The results revealed dynamic changes in the DNA methylation (5-methylcytosine) landscape in response to sulfide stress, with U. unicinctus exhibiting elevated DNA methylation levels following stress exposure. Integrating differentially expressed genes (DEGs) and differentially methylated regions (DMRs), we identified a crucial role of gene body methylation in predicting gene expression. Furthermore, using a DNA methyltransferase inhibitor, we validated the involvement of DNA methylation in the sulfide stress response and the gene regulatory network influenced by DNA methylation. The results indicated that by modulating DNA methylation levels during sulfide stress, the expression of glutathione S-transferase, glutamyl aminopeptidase, and cytochrome c oxidase could be up-regulated, thereby facilitating the metabolism and detoxification of exogenous sulfides. Moreover, DNA methylation was found to regulate and enhance the oxidative phosphorylation pathway, including NADH dehydrogenase, isocitrate dehydrogenase, and ATP synthase. Additionally, DNA methylation influenced the regulation of Cytochrome P450 and macrophage migration inhibitory factor, both of which are closely associated with oxidative stress and stress resistance. Our findings not only emphasize the role of DNA methylation in sulfide adaptation but also provide novel insights into the potential mechanisms through which marine organisms adapt to environmental changes.