Epigenetic Changes In DNA Methylation Research Articles

Epigenetic DNA methylation changes in episodic and chronic migraine.

Epigenetic DNA methylation changes in episodic and chronic migraine.

Sjögren's syndrome X-chromosome dose effect: An epigenetic perspective.

Sjögren's syndrome (SS) is a chronic autoimmune disease affecting exocrine glands leading to mouth and eyes dryness. The extent to which epigenetic DNA methylation changes are responsible for an X-chromosome dose effect has yet to be determined. Our objectives were to (i) describe how epigenetic DNA methylation changes could explain an X-chromosome dose effect in SS for women with normal 46,XX genotype and (ii) determine the relevant relationships to this dose effect, between X-linked genes, genes controlling X-chromosome inactivation (XCI) and genes encoding associated transcription factors, all of which are differentially expressed and/or differentially methylated in the salivary glands of patients with SS. We identified 58 upregulated X-chromosome genes, including 22 genes previously shown to escape XCI, based on the analysis of SS patient salivary gland GEO2R gene expression datasets. Moreover, we found XIST and its cis regulators RLIM, FTX, and CHIC1, and polycomb repressor genes of the PRC1/2 complexes to be upregulated. Many of the X-chromosome genes implicated in SS pathogenesis can be regulated by transcription factors which we found to be overexpressed and/or differentially methylated in patients with SS. Determination of the mechanisms underlying methylation-dependent gene expression and impaired XCI is needed to further elucidate the etiopathogenesis of SS.

GLI2 promoter hypermethylation in saliva of children with a respiratory allergy

BackgroundThe prevalence of respiratory allergy in children is increasing. Epigenetic DNA methylation changes are plausible underlying molecular mechanisms.ResultsSaliva samples collected in substudies of two longitudinal birth cohorts in Belgium (FLEHS1 & FLEHS2) were used to discover and confirm DNA methylation signatures that can differentiate individuals with respiratory allergy from healthy subjects. Genome-wide analysis with Illumina Methylation 450K BeadChips revealed 23 differentially methylated gene regions (DMRs) in saliva from 11y old allergic children (N=26) vs. controls (N=20) in FLEHS1. A subset of 7 DMRs was selected for confirmation by iPLEX MassArray analysis. First, iPLEX analysis was performed in the same 46 FLEHS1 samples for analytical confirmation of the findings obtained during the discovery phase. iPLEX results correlated significantly with the 450K array data (P <0.0001) and confirmed 4 out of the 7 DMRs. Aiming for additional biological confirmation, the 7 DMRs were analyzed using iPLEX in a substudy of an independent birth cohort (FLEHS2; N=19 cases vs. 20 controls, aged 5 years). One DMR in the GLI2 promoter region showed a consistent statistically significant hypermethylation in individuals with respiratory allergy across the two birth cohorts and technologies. In addition to its involvement in TGF-β signaling and T-helper differentiation, GLI2 has a regulating role in lung development.ConclusionGLI2 is considered an interesting candidate DNA methylation marker for respiratory allergy.

Soil Selenium (Se) Biofortification Changes the Physiological, Biochemical and Epigenetic Responses to Water Stress in Zea mays L. by Inducing a Higher Drought Tolerance.

Requiring water and minerals to grow and to develop its organs, Maize (Zea mays L.) production and distribution is highly rainfall-dependent. Current global climatic changes reveal irregular rainfall patterns and this could represent for maize a stressing condition resulting in yield and productivity loss around the world. It is well known that low water availability leads the plant to adopt a number of metabolic alterations to overcome stress or reduce its effects. In this regard, selenium (Se), a trace element, can help reduce water damage caused by the overproduction of reactive oxygen species (ROS). Here we report the effects of exogenous Se supply on physiological and biochemical processes that may influence yield and quality of maize under drought stress conditions. Plants were grown in soil fertilized by adding 150 mg of Se (sodium selenite). We verified the effects of drought stress and Se treatment. Selenium biofortification proved more beneficial for maize plants when supplied at higher Se concentrations. The increase in proline, K concentrations and nitrogen metabolism in aerial parts of plants grown in Se-rich substrates, seems to prove that Se-biofortification increased plant resistance to water shortage conditions. Moreover, the increase of SeMeSeCys and SeCys2 forms in roots and aerial parts of Se-treated plants suggest resistance strategies to Se similar to those existing in Se-hyperaccumulator species. In addition, epigenetic changes in DNA methylation due to water stress and Se treatment were also investigated using methylation sensitive amplified polymorphism (MSAP). Results suggest that Se may be an activator of particular classes of genes that are involved in tolerance to abiotic stresses. In particular, PSY (phytoene synthase) gene, essential for maintaining leaf carotenoid contents, SDH (sorbitol dehydrogenase), whose activity regulates the level of important osmolytes during drought stress and ADH (alcohol dehydrogenase), whose activity plays a central role in biochemical adaptation to environmental stress. In conclusion, Se-biofortification could help maize plants to cope with drought stress conditions, by inducing a higher drought tolerance.

Epigenetics of inflammatory arthritis.

Aberrant epigenetic changes in DNA methylation, histone marks, and noncoding RNA expression regulate the pathogenesis of many rheumatic diseases. The present article will review the recent advances in the epigenetic profile of inflammatory arthritis and discuss diagnostic biomarkers and potential therapeutic targets. Methylation signatures of fibroblast-like synoviocytes not only distinguish rheumatoid arthritis (RA) and osteoarthritis (OA), but also early RA from late RA or juvenile idiopathic arthritis. Methylation patterns are also specific to individual joint locations, which might explain the distribution of joint involvement in some rheumatic diseases. Hypomethylation in systemic lupus erythematosus (SLE) T cells is, in part, because of active demethylation and 5-hydroxymethylation. The methylation status of some genes in SLE is associated with disease severity and has potential as a diagnostic marker. An integrative analysis of OA methylome, transcriptome, and proteome in chondrocytes has identified multiple-evidence genes that might be evaluated for therapeutic potential. Class-specific histone deacetylase inhibitors are being evaluated for therapy in inflammatory arthritis. Disease pathogenesis is regulated by the interplay of genetics, environment, and epigenetics. Understanding how these mechanisms regulate cell function in health and disease has implications for individualized therapy.

Research Advances in the Epigenetics of Diffuse Large B-Cell Lymphoma-Review

The pathogenesis of diffuse large B-cell lymphoma(DLBCL) has not been fully elucidated, but people realized that the occurrence of DLBCL is caused by the mutual change of genetics and epigenetics with the rapid development of modern biological technology. Epigenetics is to explore the expression changes of genes and proteins without the changes of gene sequences, and this change is reversible and can be inherited. In this review, the latest advances of epigenetic changes in DNA methylation, microRNA regulation, and histone modification for DLBCL are summarized, so as to provide a new strategy to study the early diagnosis, treatment and prognosis of DLBCL.

Ascorbic acid-mediated enhanced cardiomyocyte differentiation of mouse ES-cells involves interplay of DNA methylation and multiple-signals

Embryonic stem cells (ES-cells) provide a good model system to study lineage-specific differentiation. Though, the differentiation of ES-cells to cardiomyocytes is documented, a clear understanding of the molecular mechanism of differentiation and improved functional-differentiation efficiency are yet to be achieved. In this regard, ascorbic acid (Aa) is shown to be one of the effective cardiac inducers in ES-cells. But, its mechanism is poorly understood. We therefore, investigated the mechanism of Aa-mediated cardiomyocyte differentiation of ES-cells. Here, we describe the potential involvement of epigenetic (DNA methylation) as well as integrin- and Erk- signaling systems during cardiomyocyte differentiation. Transgenic GS-2 ES-cells and wild-type D3 ES-cells were differentiated to cardiomyocytes, in the presence or absence of Aa and with or without inhibitors of Erk-, collagen- and integrin- pathways. At specific time points, differentiated states of ES-cells were scored by gene expression analyses and the proportion of functional cTnI+ cardiomyocytes. DNA methylation changes of Isl-1, BMP-2, GATA-4 and α-MHC in cardiogenic cells, following stimulation with Aa, were analyzed by using methylation specific PCR (MSP). We observed that Aa, when applied in initial phase of ES-cell differentiation, consistently enhanced cardiac differentiation (99%) over that observed during spontaneous differentiation (70%). This was associated with enhanced expressions of cardiogenesis-associated genes. A two-fold increase in cTnI+ cells was observed, with appropriate myofibril arrangement. The observed effect of Aa was due to enhanced collagen and integrin signaling, coupled with a high p-ERK1/2 expression, downstream. Besides, the involvement of DNA methylation in regulating the expression of cardiac genes i.e., Isl-1 and α-MHC was also observed. Overall, this study, for the first time, demonstrates that Aa-mediated cardiac enhancement is brought about, mechanistically, through the interplay of epigenetic changes in DNA methylation of cardiac genes (Isl-1 and α-MHC) and integrin signaling system.

Epigenetic DNA methylation changes associated with headache chronification: A retrospective case-control study.

Background The biological mechanisms of headache chronification are poorly understood. We aimed to identify changes in DNA methylation associated with the transformation from episodic to chronic headache. Methods Participants were recruited from the population-based Norwegian HUNT Study. Thirty-six female headache patients who transformed from episodic to chronic headache between baseline and follow-up 11 years later were matched against 35 controls with episodic headache. DNA methylation was quantified at 485,000 CpG sites, and changes in methylation level at these sites were compared between cases and controls by linear regression analysis. Data were analyzed in two stages (Stages 1 and 2) and in a combined meta-analysis. Results None of the top 20 CpG sites identified in Stage 1 replicated in Stage 2 after multiple testing correction. In the combined meta-analysis the strongest associated CpG sites were related to SH2D5 and NPTX2, two brain-expressed genes involved in the regulation of synaptic plasticity. Functional enrichment analysis pointed to processes including calcium ion binding and estrogen receptor pathways. Conclusion In this first genome-wide study of DNA methylation in headache chronification several potentially implicated loci and processes were identified. The study exemplifies the use of prospectively collected population cohorts to search for epigenetic mechanisms of disease.

DNA damage, repair monitoring and epigenetic DNA methylation changes in seedlings of Chernobyl soybeans

This pilot study was carried out to assess the effect of radio-contaminated Chernobyl environment on plant genome integrity 27 years after the accident. For this purpose, nuclei were isolated from root tips of the soybean seedlings harvested from plants grown in the Chernobyl area for seven generations. Neutral, neutral-alkaline, and methylation-sensitive comet assays were performed to evaluate the induction and repair of primary DNA damage and the epigenetic contribution to stress adaptation mechanisms. An increased level of single and double strand breaks in the radio-contaminated Chernobyl seedlings at the stage of primary root development was detected in comparison to the controls. However, the kinetics of the recovery of DNA breaks of radio-contaminated Chernobyl samples revealed that lesions were efficiently repaired at the stage of cotyledon. Methylation-sensitive comet assay revealed comparable levels in the CCGG methylation pattern between control and radio-contaminated samples with a slight increase of approximately 10% in the latter ones. The obtained preliminary data allow us to speculate about the onset of mechanisms providing an adaptation potential to the accumulated internal irradiation after the Chernobyl accident.Despite the limitations of this study, we showed that comet assay is a sensitive and flexible technique which can be efficiently used for genotoxic screening of plant specimens in natural and human-made radio-contaminated areas, as well as for safety monitoring of agricultural products.

Gilles de la Tourette syndrome is associated with hypermethylation of the dopamine D2 receptor gene

Several lines of evidence support a “dopaminergic hypothesis” in the pathophysiology of Gilles de la Tourette syndrome (TS). The aim of this study was to investigate for the first time epigenetic changes in DNA methylation in different dopamine genes in adult patients with TS. We included 51 well characterized adult patients with TS (41 males, 10 females, mean age = 35 ± 12.6 years, range, 18–71 years) and compared results with data from a group of 51 sex- and age-matched healthy controls. Bisulfite sequencing was used to measure peripheral DNA methylation of the dopamine transporter (DAT), the dopamine D2 receptor (DRD2), and the catechol-O-methyltransferase (COMT) genes. Compared to healthy controls, patients with TS showed significantly elevated methylation level of the DRD2 gene that positively correlated with tic severity. In contrast, DAT methylation was lower in more severely affected patients. Our results provide evidence for a role of altered epigenetic regulation of dopaminergic genes in the pathophysiology of TS. While DRD2 hypermethylation seems to be directly related to the neurobiology of TS that may lead to dopaminergic dysfunction resulting in enhanced thalamo-cortical movement-stimulating activity, DAT hypomethylation might reflect a secondary mechanism in order to compensate for increased dopaminergic signal transduction due to DRD2 hypermethylation. In addition, it can be speculated that spontaneous fluctuations of tics may be caused by short-term alterations of methylation levels of dopaminergic genes resulting in dynamic changes of tonic/phasic dopaminergic signaling in the striatum and thalamo-cortical output pathways.

A child’s spit epigenome can reveal its respiratory allergy risk

Environmental exposures like parental lifestyle and diet, smoking, obesity and exposure to toxin can modulate disease risk. Epigenetic DNA methylation changes can be part of the underlying molecular mechanisms. Early life exposures can alter DNA methylation patterns, and thereby predispose the child to develop respiratory allergy (RA) later in life. Longitudinal birth cohorts are instrumental to study disease development, but DNA biomarker research is hampered because blood sampling is kept to a minimum for practical and ethical reasons. Saliva is a non-invasive and convenient source of DNA that can be used for biomarker research. In this study, we aimed at discovery and confirmation of differential methylation regions (DMR) in saliva of children with RA when comparing to controls. Saliva samples collected in two independent longitudinal birth cohorts in Belgium were analyzed using Illumina Methylation 450K BeadChips.A statistical analysis pipeline was developed in R to identify genome-wide differential methylation. The Illumina Methylation 450K BeadChips revealed 13 DMR in saliva from 11y old allergic children (N=26) vs. controls (N=20). 5 DMR were located in genes involved in IL4 signalling and Th2-response, showing a link with wheezing and other RA phenotypes. The 13 DMR were selected for further biological and technical validation by iPLEX MassArray analysis in the same birth cohort as well as in a second cohort (N=78). This project is providing novel insights in the molecular mechanisms that may predispose children to RA development. We are among the first to show the utility of saliva to identify DNA methylation marks in children that are relevant for RA.

Sex steroid-induced DNA methylation changes and inflammation response in prostate cancer

BackgroundSex steroid hormones have been reported to induce inflammation causing dysregulation of cytokines in prostate cancer cells. However, the underlying epigenetic mechanism has not well been studied. The objective of this study was to evaluate the effect of sex steroid hormones on epigenetic DNA methylation changes in prostate cancer cells using a signature PCR methylation array panel that correspond to 96 genes with biological function in the human inflammatory and autoimmune signals in prostate cancer. Of the 96-gene panel, 32 genes showed at least 10% differentially methylation level in response to hormonal treatment when compared to untreated cells. Genes that were hypomethylated included CXCL12, CXCL5, CCL25, IL1F8, IL13RAI, STAT5A, CXCR4 and TLR5; and genes that were hypermethylated included ELA2, TOLLIP, LAG3, CD276 and MALT1. Quantitative RT-PCR analysis of select genes represented in a cytokine expression array panel showed inverse association between DNA methylation and gene expression for TOLLIP, CXCL5, CCL18 and IL5 genes and treatment of prostate cancer cells with 5′-aza-2′-deoxycytidine with or without trichostatin A induced up-regulation of TOLLIP expression. Further analysis of relative gene expression of matched prostate cancer tissues when compared to benign tissues from individual patients with prostate cancer showed increased and significant expression for CCL18 (2.6-fold; p<0.001), a modest yet significant increase in IL5 expression (1.17-fold; p=0.015), and a modest increase in CXCL5 expression (1.4-fold; p=0.25). In conclusion, our studies demonstrate that sex steroid hormones can induce aberrant gene expression via differential methylation changes in prostate carcinogenesis.

Mitochondrial polymerase gamma dysfunction and aging cause cardiac nuclear DNA methylation changes.

Cardiomyopathy (CM) is an intrinsic weakening of myocardium with contractile dysfunction and congestive heart failure (CHF). CHF has been postulated to result from decreased mitochondrial energy production and oxidative stress. Effects of decreased mitochondrial oxygen consumption also can accelerate with aging. We previously showed DNA methylation changes in human hearts with CM. This was associated with mitochondrial DNA depletion, being another molecular marker of CM. We examined the relationship between mitochondrial dysfunction and cardiac epigenetic DNA methylation changes in both young and old mice. We used genetically engineered C57Bl/6 mice transgenic for a cardiac-specific mutant of the mitochondrial polymerase-γ (termed Y955C). Y955C mice undergo left ventricular hypertrophy (LVH) at a young age (∼ 94 days old), and LVH decompensated to CHF at old age (∼ 255 days old). Results found 95 genes differentially expressed as a result of Y955C expression, while 4,452 genes were differentially expressed as a result of aging hearts. Moreover, cardiac DNA methylation patterns differed between Y955C (4,506 peaks with 68.5% hypomethylation) and aged hearts (73,286 peaks with 80.2% hypomethylated). Correlatively, of the 95 Y955C-dependent differentially expressed genes, 30 genes (31.6%) also displayed differential DNA methylation; in the 4,452 age-dependent differentially expressed genes, 342 genes (7.7%) displayed associated DNA methylation changes. Both Y955C and aging demonstrated significant enrichment of CACGTG-associated E-box motifs in differentially methylated regions. Cardiac mitochondrial polymerase dysfunction alters nuclear DNA methylation. Furthermore, aging causes a robust change in cardiac DNA methylation that is partially associated with mitochondrial polymerase dysfunction.

Epigenetic Studies Point to DNA Replication/Repair Genes as a Basis for the Heritable Nature of Long Term Complications in Diabetes.

Metabolic memory (MM) is defined as the persistence of diabetic (DM) complications even after glycemic control is pharmacologically achieved. Using a zebrafish diabetic model that induces a MM state, we previously reported that, in this model, tissue dysfunction was of a heritable nature based on cell proliferation studies in limb tissue and this correlated with epigenetic DNA methylation changes that paralleled alterations in gene expression. In the current study, control, DM, and MM excised fin tissues were further analyzed by MeDIP sequencing and microarray techniques. Bioinformatics analysis of the data found that genes of the DNA replication/DNA metabolism process group (with upregulation of the apex1, mcm2, mcm4, orc3, lig1, and dnmt1 genes) were altered in the DM state and these molecular changes continued into MM. Interestingly, DNA methylation changes could be found as far as 6–13 kb upstream of the transcription start site for these genes suggesting potential higher levels of epigenetic control. In conclusion, DNA methylation changes in members of the DNA replication/repair process group best explain the heritable nature of cell proliferation impairment found in the zebrafish DM/MM model. These results are consistent with human diabetic epigenetic studies and provide one explanation for the persistence of long term tissue complications as seen in diabetes.

Particulate matter-induced epigenetic changes and lung cancer.

Lung cancer is the leading cause of cancer death worldwide. Cigarette smoking is the well-known risk factor for lung cancer. Epidemiological studies suggest that air pollution, especially particulate matter (PM) exposure, is associated with increased lung cancer risk and mortality independent of cigarette smoking. English-language publications focusing on PM, epigenetic changes, and lung cancer were reviewed. The epigenome serves as an interface between the environment and the genome. PM is one of the environmental factors that can cause epigenetic changes. The epigenome serves as an interface between the environment and the genome. Some of the epigenetic changes lead to increased disease susceptibility and progression. In cardiovascular disease and asthma, the association between PM exposure and the disease specific epigenetic changes has been identified. In lung cancer, the epigenetic changes in DNA methylation, histone modification and microRNA expression are commonly found, but the specific link between PM exposure and lung cancer remains incompletely understood. The results of epidemiological studies indicate the important effects of PM exposure on lung cancer. PM2.5 is consistently associated with the increased lung cancer risk and mortality. Based on the epidemiological associations between PM exposure and lung cancer, PM-induced epigenetic changes may play important roles in the pathogenesis of lung cancer. In this review, we focus on the current knowledge of epigenetic changes associated with PM exposure and lung cancer. Better understanding of the link between PM exposure and lung cancer at the epigenomic level by comprehensive comparison approach may identify lung cancer early detection biomarkers and novel therapeutic targets.

Ecstasy (MDMA) Alters Cardiac Gene Expression and DNA Methylation: Implications for Circadian Rhythm Dysfunction in the Heart.

MDMA (ecstasy) is an illicit drug that stimulates monoamine neurotransmitter release and inhibits reuptake. MDMA's acute cardiotoxicity includes tachycardia and arrhythmia which are associated with cardiomyopathy. MDMA acute cardiotoxicity has been explored, but neither long-term MDMA cardiac pathological changes nor epigenetic changes have been evaluated. Microarray analyses were employed to identify cardiac gene expression changes and epigenetic DNA methylation changes. To identify permanent MDMA-induced pathogenetic changes, mice received daily 10- or 35-day MDMA, or daily 10-day MDMA followed by 25-day saline washout (10 + 25 days). MDMA treatment caused differential gene expression (p < .05, fold change >1.5) in 752 genes following 10 days, 558 genes following 35 days, and 113 genes following 10-day MDMA + 25-day saline washout. Changes in MAPK and circadian rhythm gene expression were identified as early as 10 days. After 35 days, circadian rhythm genes (Per3, CLOCK, ARNTL, and NPAS2) persisted to be differentially expressed. MDMA caused DNA hypermethylation and hypomethylation that was independent of gene expression; hypermethylation of genes was found to be 71% at 10 days, 68% at 35 days, and 91% at 10 + 25 days washout. Differential gene expression paralleled DNA methylation in 22% of genes at 10-day treatment, 17% at 35 days, and 48% at 10 + 25 days washout. We show here that MDMA induced cardiac epigenetic changes in DNA methylation where hypermethylation predominated. Moreover, MDMA induced gene expression of key elements of circadian rhythm regulatory genes. This suggests a fundamental organism-level event to explain some of the etiologies of MDMA dysfunction in the heart.

Abstract 32: Improper Histone Modification Of Gene Expression In Kawasaki Disease May Influence Both Its Development And Outcome

Kawasaki disease (KD) is characterized by development of an autoimmune vasculitis and potential coronary artery lesions (CAL). Patients present with significantly marked expression of various immune cells and the inflammatory cells that they produce, recruit, or secrete, particularly cytokines. Concurrently, regulatory immune cells that suppress immune and inflammatory cells are significantly reduced in KD, particularly regulatory T cells (Tregs) such as CD4+CD25+FOXP3+ Tregs. Therefore, the clinical picture results from an unchecked, marked increase of immune and inflammatory cells infiltrating passed activated endothelial cells and into the vascular wall. This appears to largely be a consequence of amplified cell signaling via the Ca2+/NFAT pathway in KD, which is involved in targeted gene expression. As a result, we hypothesized that epigenetic control of gene expression through histone modifications may play a leading role in development of KD, its complications, and recovery. A previous study found significant DNA hypomethylation of CpG sites for the SOCS1 gene in KD patients (n=32), which was higher in patients without CAL. We conducted a preliminary study to widely examine epigenetic changes in DNA methylation of CpG sites before and after IVIG treatment in KD patients (n=9) using age-matched febrile controls (n=8). Our results demonstrated ≥10% differences in DNA methylation rates for 67 genes in KD patients, in addition to 10 genes with ≥15% methylation difference. Larger studies are required to accurately determine the role of epigenomic changes in KD development, in addition to investigations of the methylation reactions which regulate them (see Figure 1).

Methylene tetra hydrofolate Reductase C677T Gene Polymorphism in Heroin Dependence

Dependence to opiates is a chronic relapsing brain disease that can cause major health, social and economic problems [1]. Dependence vulnerability is affected by environmental factors, drug-induced factors and hereditary factors, which represent approximately 40–60% of the risk of developing dependence [2]. Illicit drug use could lead to epigenetic changes in DNA methylation which may play an important role in the development of chronic substance abuse by enhancement of drug reward, craving and relapse use [3,4]. Epigenetic factors could be also responsible for vulnerability to drug response, dependence and addiction treatment response [5].

Evidence for Premature Aging of (Clonal but also) Non-Clonal Hematopoietic Stem Cells in Patients with Acute Myeloid Leukemia (AML)

Introduction:AML is an aggressive clonal myeloid neoplasia. Incidence increases with age and age is also the most dominant prognostic indicator for adult patients with AML. Although numerous genetic events have recently been identified in good and intermediate risk AML, understanding of the pathophysiology behind poor risk AML, which is most predominant in elderly patients and characterized by complex karyotypic abnormalities remains limited. Changes in the epigenetic signature and telomere attrition are two well recognized hallmarks of aging and genetic instability. Here, we investigate the hypothesis that poor risk AML arises from a prematurely aged, genetically instable hematopoietic stem cell population.Methods:We analyzed 45 newly diagnosed AML patients who were followed from diagnosis, through achievement of complete cytogenetic remission (CCR) and/or until refractory disease or relapse. Overall, 95 samples (n=88 bone marrow (BM), n=7 peripheral blood, (PB)) were available. Analysis were performed at first diagnosis (Dx, n=20), after two cycles of induction chemotherapy (post-IC, n=42), after three additional cycles of consolidation chemotherapy (post-CC, n=28) and after one year follow up (n=5). Mean age of the cohort was 50.4 years (range 21-75y). A recently developed “epigenetic aging signature (EAS)” capable to predicted to the chronological age of healthy individuals with a mean absolute deviation (MAD) of 5 years (Weidner et al., Genome Biol, 2014) was used to express the epigenetic age of the AML samples. Epigenetic changes in DNA methylation were analyzed by pyrosequencing and monochrome multiplex quantitative-PCR was used for telomere length (TL) analysis. TL was age-adapted based on a control cohort of 87 healthy donors as described previously.Results:A cross-sectional analysis of TL reveals substantial telomere shortening in AML patients at the time of first diagnosis compared to healthy controls (mean±SE: -0.65 ± 0.2 T/S ratio, p=0.001). When studied sequentially, the TL deficit was less pronounced in remission after IC (-0.33 ± 0.1 T/S ratio, n=34) however, still significantly shortened compared to age-adapted healthy subjects (p= 0.001). This tendency continued in patients in remission after CC (-0.18 ± 0.1 T/S ratio, n=24, p=0.03) and one year (-0.25 ± 0.1 T/S ratio, n=5, p=0.28). In comparison, in patients with active leukemia, i.e. persistent AML post-IC (-0.61 ± 0.2 T/S ratio, n=8, p=0.006) or relapsed AML post-CC (-1.0 ± 0.2 T/S ratio, n=4, p=0.04), TL was comparable to untreated, diagnostic samples.By using EAS for age prediction, leukemic cells from AML patients with active disease at first diagnosis (+29.6 ± 8.6 years, p=0.001), in persistence (+28.9 ± 11.2 years, p=0.01) and in relapse (+86.8 ± 29.3 years, p=0.01) were found to be significantly and prematurely aged compared to their respective chronological age. In contrast, premature aging based on EAS was substantially less pronounced in CCR post-IC (+4.5 ± 1.8 additional years, p=0.01) and post-CC (+1.4 ± 2.3 additional years, p=0.52).Conclusion: We demonstrate that hematopoietic cells from AML patients at diagnosis are characterized by premature aging indicated both by accelerated telomere shortening and EAS. Not unexpectedly, this premature aging signature of leukemic cells persists in refractory and/or relapsed patients. However interestingly, this phenomenon (although less pronounced) is not completely reverted once patients reach cytogenetic remission either post induction chemotherapy, post consolidation or one year after diagnosis, i.e. when hematopoiesis has shifted from clonal leukemic to non-clonal cells. These data support the conclusion that either normal cells in patients with AML undergo accelerated aging during conventional treatment or alternatively, AML arises from prematurely aged (yet non-clonal cells) hematopoietic stem cells. DisclosuresWagner:Cygenia GmbH: Employment, Equity Ownership, Patents & Royalties.

Leishmania donovani infection causes distinct epigenetic DNA methylation changes in host macrophages.

Infection of macrophages by the intracellular protozoan Leishmania leads to down-regulation of a number of macrophage innate host defense mechanisms, thereby allowing parasite survival and replication. The underlying molecular mechanisms involved remain largely unknown. In this study, we assessed epigenetic changes in macrophage DNA methylation in response to infection with L. donovani as a possible mechanism for Leishmania driven deactivation of host defense. We quantified and detected genome-wide changes of cytosine methylation status in the macrophage genome resulting from L. donovani infection. A high confidence set of 443 CpG sites was identified with changes in methylation that correlated with live L. donovani infection. These epigenetic changes affected genes that play a critical role in host defense such as the JAK/STAT signaling pathway and the MAPK signaling pathway. These results provide strong support for a new paradigm in host-pathogen responses, where upon infection the pathogen induces epigenetic changes in the host cell genome resulting in downregulation of innate immunity thereby enabling pathogen survival and replication. We therefore propose a model whereby Leishmania induced epigenetic changes result in permanent down regulation of host defense mechanisms to protect intracellular replication and survival of parasitic cells.