Impaired DNA Methylation Research Articles

Experimental Varicocele Impairs DNA Methylation and Demethylation in Germ Cells, TESE, and Epididymal Spermatozoa, Impacting Active DNA Demethylation in Zygotes.

Varicocele causes infertility. The current study has investigated the impact of experimental varicocele on DNA methylation, demethylation, and damage in the germ cells, TESE-derived and epididymal spermatozoa. Moreover, the results were compared between epidydimal and TESE-derived spermatozoa. Finally, the varicocele-induced effect on active DNA demethylation (ADD) of male pronucleus and pre-implantation embryo development was assessed. The mature male rats were divided into control, control-sham (undergone simple laparotomy), and experimental varicocele-induced groups (n = 6/each group). The left renal vein semi-ligation was considered to induce varicocele. The expression levels of DNA methyltransferase 1 (DNMT1) and ten-eleven-translocation proteins (TET1, 2, 3), and global DNA methylation in testicular tissue, TESE, and epididymis-derived spermatozoa, and the ADD in zygotes male pronucleus as well as pre-implantation embryo development were assessed. The expression levels of DNMT1 and TET1, 2, 3 in testicles, TESE, and epididymis-derived spermatozoa were decreased in the varicocele group compared to the control and control-sham groups. The TESE-derived spermatozoa exhibited higher DNMT1, higher DNMT1, and TET 1, 2, and no change in TET3 expression compared to epididymis-derived spermatozoa. The varicocele group represented lower DNA methylation in the testicles, TESE-derived and epididymal spermatozoa, higher 5mC+ signal in male pronucleus, and a lower pre-implantation embryo development compared to control and control-sham rats. The TESE-derived spermatozoa exhibited higher 5mC protein expression compared to epididymal spermatozoa. In conclusion, varicocele can negatively impact the DNA methylation/demethylation processes impairing spermatogenesis and leading to fertilization failure, which may ultimately result in a decrease in embryo development by increasing susceptibility to DNA damage.

Exploring the connection: endocrine disruptors and polycystic ovarian syndrome

Polycystic ovarian syndrome (PCOS) is heterogeneous endocrine disorder in females manifesting reproductive dysfunction and metabolic abnormalities. Endocrinopathy in the form of hyperandrogenism leading to alteration in clinical phenotype and fertility seen. Atherogenic dyslipidaemia and insulin resistance as a result of metabolic disturbance also encountered. Recent years, endocrine disrupting chemicals (EDCs) are widely studied and linked for their alleged role in the development of PCOS. EDCs like bisphenol A (BPA), Phthalate, methoxychlor and chlorpyrifos which are present in many industrial as well as daily use products poses risk of development of various diseases. This review discusses the role of EDCS specially BPA in the pathogenesis of PCOS with study of interest identified and extracted from databases like Pub Med and Google scholars using MeSH keywords. BPA has estrogenic property and binds to oestrogen receptors α and β. Stimulation of ovarian theca cells and dysregulation of steroid biosynthesis leads to androgen overproduction. It stimulates GnRH Pulse generator, decreasing the level of LH hence fertility is affected. BPA also interact with adipose tissue receptors and causes differentiation, lipid deposition and inhibition of adiponectin. Its serum and urinary levels are found to be elevated in PCOS patient. In animal studies, it is found that BPA exposure causes impaired folliculogenesis, insulin resistance and DNA methylation. EDC exposure, especially BPA which is an integral constituent of many industrial and daily use items may cause PCOS possibly by altering androgen synthesis, adipocyte stimulation and epigenetic modification.

Gestational Caloric Restriction Alters Adipose Tissue Methylome and Offspring's Metabolic Profile in a Swine Model.

Limited nutrient supply to the fetus results in physiologic and metabolic adaptations that have unfavorable consequences in the offspring. In a swine animal model, we aimed to study the effects of gestational caloric restriction and early postnatal metformin administration on offspring's adipose tissue epigenetics and their association with morphometric and metabolic variables. Sows were either underfed (30% restriction of total food) or kept under standard diet during gestation, and piglets were randomly assigned at birth to receive metformin (n = 16 per group) or vehicle treatment (n = 16 per group) throughout lactation. DNA methylation and gene expression were assessed in the retroperitoneal adipose tissue of piglets at weaning. Results showed that gestational caloric restriction had a negative effect on the metabolic profile of the piglets, increased the expression of inflammatory markers in the adipose tissue, and changed the methylation of several genes related to metabolism. Metformin treatment resulted in positive changes in the adipocyte morphology and regulated the methylation of several genes related to atherosclerosis, insulin, and fatty acids signaling pathways. The methylation and gene expression of the differentially methylated FASN, SLC5A10, COL5A1, and PRKCZ genes in adipose tissue associated with the metabolic profile in the piglets born to underfed sows. In conclusion, our swine model showed that caloric restriction during pregnancy was associated with impaired inflammatory and DNA methylation markers in the offspring's adipose tissue that could predispose the offspring to later metabolic abnormalities. Early metformin administration could modulate the size of adipocytes and the DNA methylation changes.

TET2 modulates spatial relocalization of heterochromatin in aged hematopoietic stem and progenitor cells.

DNA methylation deregulation at partially methylated domains (PMDs) represents an epigenetic signature of aging and cancer, yet the underlying molecular basis and resulting biological consequences remain unresolved. We report herein a mechanistic link between disrupted DNA methylation at PMDs and the spatial relocalization of H3K9me3-marked heterochromatin in aged hematopoietic stem and progenitor cells (HSPCs) or those with impaired DNA methylation. We uncover that TET2 modulates the spatial redistribution of H3K9me3-marked heterochromatin to mediate the upregulation of endogenous retroviruses (ERVs) and interferon-stimulated genes (ISGs), hence contributing to functional decline of aged HSPCs. TET2-deficient HSPCs retain perinuclear distribution of heterochromatin and exhibit age-related clonal expansion. Reverse transcriptase inhibitors suppress ERVs and ISGs expression, thereby restoring age-related defects in aged HSPCs. Collectively, our findings deepen the understanding of the functional interplay between DNA methylation and histone modifications, which is vital for maintaining heterochromatin function and safeguarding genome stability in stem cells.

A transgenic mice model of retinopathy of cblG-type inherited disorder of one-carbon metabolism highlights epigenome-wide alterations related to cone photoreceptor cells development and retinal metabolism

BackgroundMTR gene encodes the cytoplasmic enzyme methionine synthase, which plays a pivotal role in the methionine cycle of one-carbon metabolism. This cycle holds a significant importance in generating S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), the respective universal methyl donor and end-product of epigenetic transmethylation reactions. cblG type of inherited disorders of vitamin B12 metabolism due to mutations in MTR gene exhibits a wide spectrum of symptoms, including a retinopathy unresponsive to conventional therapies.MethodsTo unveil the underlying epigenetic pathological mechanisms, we conducted a comprehensive study of epigenomic-wide alterations of DNA methylation by NGS of bisulfited retinal DNA in an original murine model with conditional Mtr deletion in retinal tissue. Our focus was on postnatal day 21, a critical developmental juncture for ocular structure refinement and functional maturation.ResultsWe observed delayed eye opening and impaired visual acuity and alterations in the one-carbon metabolomic profile, with a notable dramatic decline in SAM/SAH ratio predicted to impair DNA methylation. This metabolic disruption led to epigenome-wide changes in genes involved in eye development, synaptic plasticity, and retinoid metabolism, including promoter hypermethylation of Rarα, a regulator of Lrat expression. Consistently, we observed a decline in cone photoreceptor cells and reduced expression of Lrat, Rpe65, and Rdh5, three pivotal genes of eye retinoid metabolism.ConclusionWe introduced an original in vivo model for studying cblG retinopathy, which highlighted the pivotal role of altered DNA methylation in eye development, cone differentiation, and retinoid metabolism. This model can be used for preclinical studies of novel therapeutic targets.

THE CONTRIBUTION OF EPIGENETIC MECHANISMS TO THE FORMATION, MAINTENANCE AND RECONSOLIDATION OF THE LONG-TERM FOOD AVERSIVE MEMORY IN TERRESTRIAL SNAIL

This work was devoted to the analysis of the role of epigenetic mechanisms (histone acetylation, DNA methylation, histone serotonylation) in the formation, storage, and reconsolidation of long-term food aversive memory in the snail. In the first part of the work, we investigated the effect of systemic administration of sodium butyrate, an inhibitor of histone deacetylase, in a model of conditioned food aversion reflex of the snail. We have shown that the administration of sodium butyrate to animals with poor memory resulted in memory enhancement. Further, in experiments using the DNA methyltransferase blocker RG108, it was found that impaired DNA methylation impairs long-term food aversive memory in the snail. However, memory reactivation neutralizes the effect of inhibiting the activity of DNA methyltransferases. In the final section of the work, the hypothesis of the involvement of transglutaminase-mediated serotonylation in the processes of reconsolidation of food aversive memory in the snail was tested. Behavioral analysis showed that administration of the transglutaminase blocker monodansylcadaverine (hence, blockade of serotonylation) after a reminder disrupted the reconsolidation process and led to memory suppression/erasure.

Young plasma transfer recovers decreased sperm counts and restores epigenetics in aged testis

Background and aimAging is one of the causes of male infertility, and abnormal global DNA methylation and imprinting defects have been characterized in testis during biological aging. One of the important emerging approaches aims to take advantage of the healing properties of young blood plasma to limit the progression of aging in various organs in the body. We aimed to show whether blood plasma transfer has an effect on DNA methylation and spermatogenetic cell development. In addition, we aimed to show whether the young plasma transfer to old mice has an effect on the rejuvenation of the old and whether the impaired DNA methylation and PCNA expression in old age can be restored. MethodsGroups were (i) young control, (ii) young plasma transfer to aged, (iii) aged control, (iv) aged plasma transfer to young. We utilized IHC and WB in protein level of Dnmts. For the global DNA methylation level, we used 5-methylcytosine staining. We also analyzed PCNA protein expressions in all groups by IHC. ResultsWe found that transfusion of young plasma into the old animal restored DNA methylation and PCNA expression as it did in the young animal. Most importantly, we observed an increase in spermatogonia and spermatid counts in older animals after young blood plasma transfer. ConclusionsOur findings show that young plasma transfer can restore epigenetic disorders that occur with aging and solve infertility problems by increasing the sperm count that decreases. It needs to be supported by different studies, especially human studies.

Prospects for the use of chondroitin sulfate and glucosamine sulfate in the treatment of patients with obesity-associated osteoarthritis (metabolic syndrome)

The relationship between the pathophysiology of osteoarthritis (OA) and metabolic disorders (metabolic syndrome, obesity) is provided not only by mechanical causes (increased body weight pressure on the joints). A complex of molecular mechanisms, which mediates OA effect on the development of obesity, was established. Excessive activity of toll receptors, the NF-κB cascade, and metabolic disorders of endogenous chondroitin sulfates (CS) lead to chronic inflammation and the development of a complex of comorbid pathologies, including OA, atherosclerosis, and obesity. The relationship between insulin resistance and CS metabolism is also mediated by impaired genomic DNA methylation. Exogenous CS and glucosamine sulfate (GS) used in the long-term treatment of OA also contribute to the inhibition of the pathophysiology of obesity. By inhibiting O-glucosamination of intranuclear proteins (i.e., p53), GS can accelerate lipolysis of visceral fat. Anti-inflammatory effects of CS and GS is associated with inhibition of toll receptors and NF-κB, increased levels of antioxidant enzymes, regulation of expression of fibroblast growth factor 21, activation of adenosine monophosphate-activated protein kinase, and inhibition of secretion of chemoattractant protein MCP-1 and pancreatic lipase. Positive effect of CS and its oligosaccharides exposure on the pathophysiology of metabolic disorders is associated not only with a decrease in inflammation and normalization of fat metabolism but also with an improvement in the state of the intestinal microbiota. Experimental and clinical studies confirm the effects of CS and GS on body mass control. CS and GS are effective and safe when used in patients with OA associated with metabolic syndrome and/or obesity.

Epigenetic Factors in Eutopic Endometrium in Women with Endometriosis and Infertility.

Eutopic endometrium in patients with endometriosis is characterized by aberrant expression of essential genes during the implantation window. It predisposes to disturbance of endometrial receptivity. The pathomechanism of implantation failures in women with endometriosis remains unclear. This paper aims to summarize the knowledge on epigenetic mechanisms in eutopic endometrium in the group of patients with both endometriosis and infertility. The impaired DNA methylation patterns of gene promoter regions in eutopic tissue was established. The global profile of histone acetylation and methylation and the analysis of selected histone modifications showed significant differences in the endometrium of women with endometriosis. Aberrant expression of the proposed candidate genes may promote an unfavorable embryonic implantation environment of the endometrium due to an immunological dysfunction, inflammatory reaction, and apoptotic response in women with endometriosis. The role of the newly discovered proteins regulating gene expression, i.e., TET proteins, in endometrial pathology is not yet completely known. The cells of the eutopic endometrium in women with endometriosis contain a stable, impaired methylation pattern and a histone code. Medication targeting critical genes responsible for the aberrant gene expression pattern in eutopic endometrium may help treat infertility in women with endometriosis.

Novel anticancer drug curaxin CBL0137 impairs DNA methylation by eukaryotic DNA methyltransferase Dnmt3a

Novel DNA intercalating anticancer drug curaxin CBL0137 significantly inhibited in vitro DNA methylation by eukaryotic DNA methyltransferase Dnmt3a catalytic domain (Dnmt3a-CD) at low micromolar concentrations (IC50 3–9 µM). CBL0137 reduced the binding affinity of Dnmt3a-CD to its DNA target, causing up to four-fold increase in the Kd of the enzyme/DNA complex. Binding of CBL0137 to Dnmt3a-CD was not observed. The observed decrease in methylation activity of Dnmt3a-CD in the presence of CBL0137 can be explained by curaxin’s ability to intercalate into DNA.

Loss of Apc Rapidly Impairs DNA Methylation Programs and Cell Fate Decisions in Lgr5+ Intestinal Stem Cells.

Colorectal cancer initiation and progression result from the accumulation of genetic and epigenetic alterations. Although aberrant gene expression and DNA methylation profiles are considered hallmarks of colorectal cancer development, the precise timing at which these are produced during tumor establishment remains elusive. Here we investigated the early transcriptional and epigenetic changes induced by adenomatous polyposis coli (Apc) inactivation in intestinal crypts. Hyperactivation of the Wnt pathway via Apc inactivation in crypt base columnar intestinal stem cells (ISC) led to their rapid accumulation driven by an impaired molecular commitment to differentiation, which was associated with discrete alterations in DNA methylation. Importantly, inhibiting the enzymes responsible for de novo DNA methylation restored the responsiveness of Apc-deficient intestinal organoids to stimuli regulating the proliferation-to-differentiation transition in ISC. This work reveals that early DNA methylation changes play critical roles in the establishment of the impaired fate decision program consecutive to Apc loss of function. SIGNIFICANCE: This study demonstrates the functional impact of changes in DNA methylation to determine the colorectal cancer cell phenotype following loss of Apc function.

Folic acid supplementation alleviates reduced ureteric branching, nephrogenesis, and global DNA methylation induced by maternal nutrient restriction in rat embryonic kidney.

We previously reported that maternal nutrient restriction (NR) inhibited ureteric branching, metanephric growth, and nephrogenesis in the rat. Here we examined whether folic acid, a methyl-group donor, rescues the inhibition of kidney development induced by NR and whether DNA methylation is involved in it. The offspring of dams given food ad libitum (CON) and those subjected to 50% food restriction (NR) were examined. NR significantly reduced ureteric tip number at embryonic day 14, which was attenuated by folic acid supplementation to nutrient restricted dams. At embryonic day 18, glomerular number, kidney weight, and global DNA methylation were reduced by NR, and maternal folic acid supplementation again alleviated them. Among DNA methyltransferases (DNMTs), DNMT1 was strongly expressed at embryonic day 15 in CON but was reduced in NR. In organ culture, an inhibitor of DNA methylation 5-aza-2 '-deoxycytidine as well as medium lacking methyl donors folic acid, choline, and methionine, significantly decreased ureteric tip number and kidney size mimicking the effect of NR. In conclusion, global DNA methylation is necessary for normal kidney development. Folic acid supplementation to nutrient restricted dams alleviated the impaired kidney development and DNA methylation in the offspring.

Oocyte IVM or vitrification significantly impairs DNA methylation patterns in blastocysts as analysed by single-cell whole-genome methylation sequencing.

To explore the mechanisms leading to the poor quality of IVF blastocysts, the single-cell whole-genome methylation sequencing technique was used in this study to analyse the methylation patterns of bovine blastocysts derived from invivo, fresh (IVF) or vitrified (V_IVF) oocytes. Genome methylation levels of blastocysts in the IVF and V_IVF groups were significantly lower than those of the invivo group (P<0.05). In all, 1149 differentially methylated regions (DMRs) were identified between the IVF and invivo groups, 1578 DMRs were identified between the V_IVF and invivo groups and 151 DMRs were identified between the V_IVF and IVF groups. For imprinted genes, methylation levels of insulin-like growth factor 2 receptor (IGF2R) and protein phosphatase 1 regulatory subunit 9A (PPP1R9A) were lower in the IVF and V_IVF groups than in the invivo group, and the methylation level of paternally expressed 3 (PEG3) was lower in the V_IVF group than in the IVF and invivo groups. Genes with DMRs between the IVF and invivo and the V_IVF and IVF groups were primarily enriched in oocyte maturation pathways, whereas DMRs between the V_IVF and invivo groups were enriched in fertilisation and vitrification-vulnerable pathways. The results of this study indicate that differences in the methylation of critical DMRs may contribute to the differences in quality between invitro- and invivo-derived embryos.

DNA Methylation of the Insulin-Like Growth Factor 2-Imprinted Gene in Trophoblast Cells of Elongated Bovine Embryo: Effects of the In Vitro Culture.

DNA methylation is an essential epigenetic mark for embryo development and can be susceptible to environment factors such as in vitro conditions. The aim of this study was to verify the effect of in vitro culture until Day (D) 14 of the development on the embryo size and DNA methylation pattern of the insulin-like growth factor 2 (IGF2)-imprinted gene. To achieve this, we produced bovine embryos completely in vivo, completely in vitro, and in vitro until D7 and then in vivo up to D14. The embryos produced in in vitro were smaller than those in other two groups (p = 0.024); no differences in embryo size were observed between genders. The in vitro embryos showed a higher level of DNA methylation in the IGF2 as compared with that in the completely in vivo-produced (IVV) embryos (p = 0.009). Furthermore, totally in vitro-produced male embryos showed higher levels of DNA methylation as compared with those observed for the totally IVV male embryos (p = 0.034). No differences were observed among genders for IGF2 DNA methylation. These results showed that the window between D7 and D14 is critical for embryo development and alterations in the environmental conditions during this period can impair DNA methylation establishment of important developmental imprinted genes. This study brings unprecedented data for bovine embryos regarding the impact of the environmental conditions during the posthatching development.

Association of Polymorphisms in Genes Involved in One-Carbon Metabolism with MTHFR Methylation Levels.

Methylenetetrahydrofolate reductase (MTHFR) is a pivotal enzyme in the one-carbon metabolism, a metabolic pathway required for DNA synthesis and methylation reactions. MTHFR hypermethylation, resulting in reduced gene expression, can contribute to several human disorders, but little is still known about the factors that regulate MTHFR methylation levels. We performed the present study to investigate if common polymorphisms in one-carbon metabolism genes contribute to MTHFR methylation levels. MTHFR methylation was assessed in peripheral blood DNA samples from 206 healthy subjects with methylation-sensitive high-resolution melting (MS-HRM); genotyping was performed for MTHFR 677C>T (rs1801133) and 1298A>C (rs1801131), MTRR 66A>G (rs1801394), MTR 2756A>G (rs1805087), SLC19A1 (RFC1) 80G>A (rs1051266), TYMS 28-bp tandem repeats (rs34743033) and 1494 6-bp ins/del (rs34489327), DNMT3A -448A>G (rs1550117), and DNMT3B -149C>T (rs2424913) polymorphisms. We observed a statistically significant effect of the DNMT3B -149C>T polymorphism on mean MTHFR methylation levels, and particularly CT and TT carriers showed increased methylation levels than CC carriers. The present study revealed an association between a functional polymorphism of DNMT3B and MTHFR methylation levels that could be of relevance in those disorders, such as inborn defects, metabolic disorders and cancer, that have been linked to impaired DNA methylation.

PS1060 CO‐OCCURRENCE OF DNMT3A‐R882, FLT3 AND NPM1 MUTATIONS IDENTIFIES A SUBSET OF AML PATIENTS WITH ADVERSE PROGNOSIS

Background:DNMT3A mutations are found in approximately 22% of adult acute myeloid leukemia (AML) cases, occurring more commonly in a hotspot at codon 882. These mutations are an early event in leukemogenesis, causing impaired DNA methylation and aberrant gene expression. Previous studies have shown that isolated DNMT3A mutations may cause limited clonal hematopoiesis in healthy individuals, however, additional mutations in genes such as FLT3 or NPM1 are required for leukemic transformation. Despite its well‐recognized role in AML development, little is known about the prognostic repercussions of the interaction between these three genes.Aims:Here, we aimed to investigate i) the clinical and biological differences between #R882 (hotspot) and non‐R882 mutations in AML and ii) the impact of the synergism between DNMT3a, FLT3 and NPM1 mutations on patient survival.Methods:218 non‐selected adult AML patients (non‐AML M3) followed in a reference center in northeast Brazil were enrolled. Additionally, 180 patients from The Cancer Genome Atlas (TCGA) databank were evaluated as an independent cohort. Brazilian patients were treated according to the 7+3 chemotherapy protocol followed by consolidation with high doses of cytarabine, while patients from the TCGA cohort were treated according to the National Comprehensive Cancer Network recommendations. Screening for DNMT3a mutations was performed by direct Sanger Sequencing.Results:DNMT3A R882 mutations were associated with FAB monocytic AML subtypes (M4 and M5), higher leukocyte counts, normal karyotype and FLT3 and NPM1 mutations (more evident for FLT3/NPM1 double‐positivity), while non‐R882 mutations were associated with NRAS and IDH2 mutations. Analysis of TCGA patient's transcriptome data revealed that patients harboring concomitant DNMT3a, FLT3 and NPM1 mutations had different patterns of gene expression compared to their counterparts with isolated mutations. Next, to address whether the association between DNMT3a R882, FLT3 and NPM1 mutations is clinically relevant, we created a composite variable grouping patients harboring DNMT3a R882 mutations with at least one mutation in FLT3 or NPM1 (here referred to as R882/FLT3/NPM1). Strikingly, patients assigned to the R882/FLT3/NPM1 group had significantly poorer overall (Brazilian cohort: HR: 2.44, 95% CI: 1.35–4.44; P = 0.002 and TCGA cohort: HR: 1.83, 95% CI: 1.09–3.06; P = 0.022) and disease‐free survival rates (Brazilian cohort: HR: 1.94, 95% CI: 1.08–5.92; P = 0.027 and TCGA cohort: HR: 2.06, 95% CI: 1.11–3.81; P = 0.021) in both cohorts. Similar results were observed when analysis was restricted to the normal‐karyotype group. On the other hand, individual analysis of DNMT3a (or #R882 only) mutations showed only a tendency for shorter survival that did not reached statistical significance in both cohorts. Proportional hazards modeling showed that R882/FLT3/NPM1, but not single mutational status, remained independently predictive of poor outcomes (Brazilian cohort: HR: 1.97, 95% CI: 1.06–3.84; P = 0.048 and TCGA cohort: HR: 2.14, 95% CI: 1.24–3.68; P = 0.006).Summary/Conclusion:AML patients with R882 mutations have distinct clinical and molecular signatures from those with non‐R882. At the transcriptome level, triple positive‐mutated patients (DNMT3a‐R882, FLT3 and NPM1) cluster differently from the single positives ones, suggesting functional interaction between these mutations. By transposing these findings into a composite variable, we show that the synergism between DNMT3a, FLT3 and NPM1 mutations identifies a subset of AML with a particularly adverse prognosis.

Oncogenic Roles and Inhibitors of DNMT1, DNMT3A, and DNMT3B in Acute Myeloid Leukaemia.

Epigenetic alteration has been proposed to give rise to numerous classic hallmarks of cancer. Impaired DNA methylation plays a central role in the onset and progression of several types of malignancies, and DNA methylation is mediated by DNA methyltransferases (DNMTs) consisting of DNMT1, DNMT3A, and DNMT3B. DNMTs are frequently implicated in the pathogenesis and aggressiveness of acute myeloid leukaemia (AML) patients. In this review, we describe and discuss the oncogenic roles of DNMT1, DNMT3A, and DNMT3B in AML. The clinical response predictive roles of DNMTs in clinical trials utilising hypomethylating agents (azacitidine and decitabine) in AML patients are presented. Novel hypomethylating agent (guadecitabine) and experimental DNMT inhibitors in AML are also discussed. In summary, hypermethylation of tumour suppressors mediated by DNMT1 or DNMT3B contributes to the progression and severity of AML (except MLL-AF9 and inv(16)(p13;q22) AML for DNMT3B), while mutation affecting DNMT3A represents an early genetic lesion in the pathogenesis of AML. In clinical trials of AML patients, expression of DNMTs is downregulated by hypomethylating agents while the clinical response predictive roles of DNMT biomarkers remain unresolved. Finally, nucleoside hypomethylating agents have continued to show enhanced responses in clinical trials of AML patients, and novel non-nucleoside DNMT inhibitors have demonstrated cytotoxicity against AML cells in pre-clinical settings.

DNA methylation, environmental exposures and early embryo development.

The first crucial step in the developmental program occurs during pre-implantation, the time after the oocyte has been fertilized and before the embryo implants in the uterus. This period represents a vulnerable window as the epigenome undergoes dynamic changes in DNA methylation profiles. Alterations in the early embryonic reprogramming wave can impair DNA methylation patterns and induce permanent changes to the developmental program, leading to the onset of adverse health outcomes in offspring. Although there is an increasing body of evidence indicating that harmful exposures during pre-implantation embryo development can trigger lasting epigenetic alterations in offspring, the mechanisms are still not fully understood. Since physiological or pathological changes in DNA methylation can occur as a response to environmental cues, proper environmental milieu plays a critical role in the success of embryonic development. In this review, we depict the mechanisms behind the embryonic epigenetic reprogramming of DNA methylation and highlight how maternal environmental stressors (e.g., alcohol, heat stress, nutrient availability) during pre-implantation and assisted reproductive technology procedures affect development and DNA methylation marks.

Brain iron loading impairs DNA methylation and alters GABAergic function in mice.

Iron deficiency is closely associated with altered GABA metabolism and affective behavior. While mutation in the hemochromatosis ( HFE) gene disrupts iron homeostasis and promotes oxidative stress that increases the risk of neurodegeneration, it is largely unknown whether HFE mutation modifies GABAergic homeostasis and emotional behavior. The goal of our study was to investigate the impact of HFE on GABAergic neurochemistry and redox-epigenetic regulation in the brain using H67D HFE-mutant mice that recapitulates the H63D-HFE mutation in humans. H67D mice displayed elevated redox-active iron levels in the brain by 32% compared to age-matched wild-type mice. Moreover, the H67D brain had increased isoprostane and decreased glutathione, indicating elevated oxidative stress. Additionally, the H67D brain had decreased global methylation and attenuated DNA methyltransferase (DNMT) activity. Direct addition of iron to purified DNMT in vitro decreased enzyme activity in a concentration-dependent manner. Last, H67D mice exhibited decreased anxiety-like behavior, which was associated with increased expression of the GABAA receptor α2 subunits by 93%, and these changes were also observed in H67D mice fed a low-iron diet. Taken together, our results suggest a putative role of HFE in regulating labile iron status in the brain, and mutation in H67D perturbs redox-methylation status, contributing to GABAergic dysfunction.-Ye, Q., Trivedi, M., Zhang, Y., Böhlke, M., Alsulimani, H., Chang, J., Maher, T., Deth, R., Kim, J. Brain iron loading impairs DNA methylation and alters GABAergic function in mice.

Superovulation alters DNA methyltransferase protein expression in mouse oocytes and early embryos.

DNA methylation is an epigenetic mechanism that plays critical roles during mammalian oocyte and preimplantation embryo development. It is achieved by adding a methyl group to the fifth carbon atom of cytosine residues within cytosine-phosphate-guanine (CpG) and non-CpG dinucleotide sites using DNA methyltransferase (DNMT) enzymes for de novo and maintenance methylation processes. DNMT1, DNMT3A, and DNMT3B play important roles in establishing methylation of developmentally related genes in oocytes and early embryos. The purpose ofthis study is to identify the effect of superovulation on the expression and subcellular localizations of these three DNMT enzymes in the mouse oocytes and early embryos. Three groups composed of control, normal dose [5IU pregnant mare serum gonadotropin (PMSG) and 5IU human chorionic gonadotropin (hCG)], and high dose [7.5IU PMSG and 7.5IU hCG] were created from 4-5-week-old female BALB/c mice. The relative expression and subcellular localizations of the DNMT proteins in the control and experiment groups have been characterized by using immunofluorescence staining subsequently analyzed in detailed. DNMT1, DNMT3A, and DNMT3B protein expression in the germinal vesicle and metaphase II oocytes and in one-cell and two-cell embryos differed significantly when some of the normal- and high-dose groups were compared with the control counterparts. This study has demonstrated for the first time that superovulation alters expression levels of the DNMT proteins, a finding that indicates that certain developmental defects in superovulated oocytes and early embryos may result from impaired DNA methylation processes.