Methylation Index Research Articles

Association between Arsenic Methylation in Early Pregnancy and Gestational Diabetes Mellitus: A Prospective Cohort Study.

The metabolism of arsenic (As) plays a crucial role in its health effects. However, the impact of arsenic methylation during early pregnancy on gestational diabetes mellitus (GDM) remains unclear. This study aimed to investigate the associations between As methylation in the first and second trimesters and the incidence of GDM by conducting a prospective cohort study in Chongqing, China. Urine samples from first (n = 131) and second (n = 53) trimester pregnant women were analyzed for arsenite (AsIII), arsenate (AsV), monomethylarsonic acid (MMA), and dimethylarsinic acid (DMA) concentrations. Logistic regressions assessed associations between the concentrations of As species, methylation indices, and GDM risk. As species concentrations showed no significant differences between GDM and non-GDM groups. Higher MMA% (OR = 1.11; 95% CI = 1.02, 1.22) and lower secondary methylation index (SMI) (OR = 0.81; 95% CI = 0.71, 0.93) correlated with GDM risk, primarily in the first trimester. MMA% decreased and DMA% and SMI increased from the first to the second trimester. Results of stratified analyses revealed these associations in women under 28 or with normal BMIs (18-24 kg/m2). The study underscores inefficient arsenic methylation as a GDM risk, modified by age and BMI, with the first trimester as a critical period.

Low-to-Moderate Arsenic Exposure and Urothelial Tract Cancers with a Long Latent Period of Follow-Up in an Arseniasis Area

BackgroundArsenic exposure can cause adverse health effects. The effects of long-term low-to-moderate exposure and methylations remain unclear.ObjectiveThis study aims to examine the association between low-to-moderate arsenic exposure and urothelial tract cancers while considering the effects of methylation capacity.MethodsIn this study, 5,811 participants were recruited from an arseniasis area in Taiwan for inorganic arsenic metabolite analysis. This follow-up study was conducted between August 1995 and December 2017. We identified 85 urothelial tract cancers in these participants, including 49 bladder and 36 upper urothelial tract cancer cases. A Cox proportional hazards model was employed.ResultsThe analyses revealed a significant association between concentrations of inorganic arsenic in water > 100 ug/L and bladder cancer occurrence, with a hazard ratio (HR) of 4.88 (95% CI 1.35–17.61). A monotonic trend was observed between concentrations of inorganic arsenic in water (from 0 to > 100 ug/L) and the incidence of urothelial tract cancer, including bladder cancer (p < 0.05) and upper urothelial tract cancers (p < 0.05). Participants with a lower primary methylation index or higher secondary methylation index had a prominent effect.ConclusionsRigorous regulations and active interventions should be considered for populations with susceptible characteristics.

Anthropometric measures and arsenic methylation among pregnant women in rural northern Bangladesh

IntroductionArsenic methylation converts inorganic arsenic (iAs) to monomethyl (MMA) and dimethyl (DMA) arsenic compounds. Body mass index (BMI) has been positively associated with arsenic methylation efficiency (higher DMA%) in adults, but evidence in pregnancy is inconsistent. We estimated associations between anthropometric measures and arsenic methylation among pregnant women in rural northern Bangladesh. MethodsWe enrolled pregnant women (n = 784) (median [IQR] gestational week: 14 [13, 15]) in Gaibandha District, Bangladesh from 2018 to 2019. Anthropometric measures were BMI, subscapular and triceps skinfold thicknesses, and mid-upper arm circumference (MUAC), fat area (MUAFA), and muscle area (MUAMA). Arsenic methylation measures were urinary iAs, MMA, and DMA divided by their sum and multiplied by 100 (iAs%, MMA%, and DMA%), primary methylation index (MMA/iAs; PMI), and secondary methylation index (DMA/MMA; SMI). In complete cases (n = 765 [97.6%]), we fitted linear, beta, and Dirichlet regression models to estimate cross-sectional differences in iAs%, MMA%, DMA%, PMI, and SMI per IQR-unit difference in each anthropometric measure, adjusting for drinking water arsenic, age, gestational age, education, living standards index, and plasma folate, vitamin B12, and homocysteine. ResultsMedian (IQR) BMI, subscapular skinfold thickness, triceps skinfold thickness, MUAC, MUAFA, and MUAMA were 21.5 (19.4, 23.8) kg/m2, 17.9 (13.2, 24.2) mm, 14.2 (10.2, 18.7) mm, 25.9 (23.8, 28.0) cm, 15.3 (10.5, 20.3) cm2, and 29.9 (25.6, 34.2) cm2, respectively. Median (IQR) iAs%, MMA%, DMA%, PMI, and SMI were 12.0 (9.3, 15.2)%, 6.6 (5.3, 8.3)%, 81.0 (77.1, 84.6)%, 0.6 (0.4, 0.7), and 12.2 (9.3, 15.7), respectively. In both unadjusted and adjusted linear models, all anthropometric measures were negatively associated with iAs%, MMA%, and PMI and positively associated with DMA% and SMI. For example, fully adjusted mean differences (95% CI) in DMA% per IQR-unit difference in BMI, subscapular skinfolds thickness, triceps skinfold thickness, MUAC, MUAFA, and MUAMA were 1.72 (1.16, 2.28), 1.58 (0.95, 2.21), 1.74 (1.11, 2.37), 1.45 (0.85, 2.06), 1.70 (1.08, 2.31), and 0.70 (0.13, 1.27) pp, respectively. ConclusionsAnthropometric measures were positively associated with arsenic methylation efficiency among pregnant women in the early second trimester.

Higher β cell death in pregnant women, measured by DNA methylation patterns of cell-free DNA, compared to new-onset type 1 and type 2 diabetes subjects: a cross-sectional study

Diabetes is a metabolic disorder of glucose homeostasis in which β cell destruction occurs silently and is detected mainly when symptoms appear. In the last few years, it has emerged a great interest in developing markers capable of detecting pancreatic β cell death focused on improving early diagnosis and getting a better treatment response, mainly in type 1 diabetes. But other types of diabetes would also benefit from early detection of β cell death. Differentially methylated circulating DNA is being studied as minimally invasive biomarker of cell death. We aimed to explore whether the unmethylated/methylated ratio of the insulin and amylin genes might be a good biomarker of β cell death in different types of diabetes. A lower index ∆Ct indicates a higher rate of β-cell death. Plasma samples from subjects without diabetes, pregnant women, pregnant with gestational diabetes (GDM), type 1 diabetes and type 2 diabetes were analyzed. A qPCR reaction with specific primers for both methylated and unmethylated fragments of insulin and amylin genes were carried out. Pregnant women, GDM and non- GDM, showed a higher β-cell death for both markers (∆INS = 3.8 ± 2.1 and ∆Amylin = 8.5 ± 3.6), whereas T1D presented lower rate (∆INS = 6.2 ± 2.1 and ∆Amylin = 10.7 ± 2.9) comparable to healthy subjects. The insulin methylation index was associated with the newborn birth weight (r = 0.46; p = 0.033) and with insulin resistance (r = -0.533; p = 0.027) in the GDM group. The higher rate of β-cell death was observed in pregnant women independently of their metabolic status. These indexes could be a good indicator of β cell death in processes caused by defects on insulin secretion, insulin action, or both.

Association of Relative Telomere Length and LINE-1 Methylation with Autism but not with Severity

Autism is associated with genomic instability, which is regulated by telomere length (TL) and index of global methylation (LINE-1). This study will determine relative TL (RTL) and LINE-1 methylation percentage for 69 patients and 33 control subjects to evaluate their potential role as biomarkers for autism. The results displayed a significant decrease of both RTL and LINE-1 methylation in autistic cases relative to controls (P < 0.001). Analysis of receiver operating characteristics curve revealed that both of RTL and LINE-1 methylation percentage have the ability to serve as autism biomarkers (area under the curve = 0.817 and 0.889, respectively). The statistical analysis revealed positive correlation between the two biomarkers (correlation coefficient = 0.439 and P < 0.001).

Holocene warming trend based on peat brGDGTs records from southeastern humid to northwestern arid China

Recent syntheses of marine paleoclimate records have revealed an overall long-term Holocene warming trend, especially in mean annual temperature (MAT), which is inconsistent with most terrestrial records on continental and global scales. We present a record of branched glycerol dialkyl glycerol tetraethers (brGDGTs) from the Shiwangutian (SWGT) peatland in central-southern China. Comparative results from surface peat samples from the SWGT peatland and the surrounding surface soils demonstrate that the brGDGTs are primarily of in-situ origin, demonstrating their potential as a record of local environmental changes. Monthly observational results show no seasonal variations in the methylation index of 5-methyl brGDGTs (MBT'5MEwhich has been proposed as an indicator of local environmental temperature in peatlands) in the surface peat samples, suggesting that it is an indicator of MAT. Thus, we conclude that the long-term increasing trend in the brGDGTs-based MBT'5ME records of two parallel independently-dated peat cores from the SWGT peatland clearly demonstrate a long-term warming trend during the Holocene. Additionally, we found that the brGDGTs-based records from five peatlands along an environmental gradient from southeastern (SE) humid to northwestern (NW) arid China also show a long-term Holocene warming trend. However, this trend is the opposite to the long-term Holocene cooling trend indicated by brGDGTs records from two peatlands in northeastern (NE) China, highlighting the complexity of peat brGDGTs as a climatic indicator. We therefore appeal for more peat brGDGTs studies, which are important for understanding the climate history of the Holocene and its potential relationship with human activities.

Expression of the NUP153 and YWHAB genes from their canonical promoters and alternative promoters of the LINE-1 retrotransposon in the placenta of the first trimester of pregnancy.

The placenta has a unique hypomethylated genome. Due to this feature of the placenta, there is a potential possibility of using regulatory elements derived from retroviruses and retrotransposons, which are suppressed by DNA methylation in the adult body. In addition, there is an abnormal increase in the level of methylation of the LINE-1 retrotransposon in the chorionic trophoblast in spontaneous abortions with both normal karyotype and aneuploidy on different chromosomes, which may be associated with impaired gene transcription using LINE-1 regulatory elements. To date, 988 genes that can be expressed from alternative LINE-1 promoters have been identified. Using the STRING tool, genes (NUP153 and YWHAB) were selected, the products of which have significant functional relationships with proteins highly expressed in the placenta and involved in trophoblast differentiation. This study aimed to analyze the expression of the NUP153 and YWHAB genes, highly active in the placenta, from canonical and alternative LINE-1 promoters in the germinal part of the placenta of spontaneous and induced abortions. Gene expression analysis was performed using real-time PCR in chorionic villi and extraembryonic mesoderm of induced abortions (n = 10), adult lymphocytes (n = 10), spontaneous abortions with normal karyotype (n = 10), and with the most frequent aneuploidies in the first trimester of pregnancy (trisomy 16 (n = 8) and monosomy X (n = 6)). The LINE-1 methylation index was assessed in the chorionic villi of spontaneous abortions using targeted bisulfite massive parallel sequencing. The level of expression of both genes from canonical promoters was higher in blood lymphocytes than in placental tissues (p < 0.05). However, the expression level of the NUP153 gene from the alternative LINE-1 promoter was 17 times higher in chorionic villi and 23 times higher in extraembryonic mesoderm than in lymphocytes (p < 0.05). The expression level of NUP153 and YWHAB from canonical promoters was higher in the group of spontaneous abortions with monosomy X compared to all other groups (p <0.05). The LINE-1 methylation index negatively correlated with the level of gene expression from both canonical (NUP153 - R = -0.59, YWHAB - R = -0.52, p < 0.05) and alternative LINE-1 promoters (NUP153 - R = -0.46, YWHAB - R = -0.66, p < 0.05). Thus, the observed increase in the LINE-1 methylation index in the placenta of spontaneous abortions is associated with the level of expression of the NUP153 and YWHAB genes not only from alternative but also from canonical promoters, which can subsequently lead to negative consequences for normal embryogenesis.

Diagnostic value of cell-free DNA in thyroid cancer: A systematic review and meta-analysis.

An increasing number of studies have shown the potential diagnostic value of cell-free DNA (cfDNA) as a new biomarker in the management of thyroid cancer (TC); however, the accuracy of research results is inconsistent. This meta-analysis is the first to synthesize published results and evaluate the application value of circulating cfDNA in the diagnosis of TC. A search strategy was developed according to PICO (P: Patient; I: Intervention; C: Comparison; O: Outcome) principles. We searched 5 databases until October 2022. Original studies that examined cfDNA for the diagnosis of TC and used pathology as the gold standard were included in this meta-analysis. A random-effects model was used to pool the data extracted from individual studies, including the number of patients and the numbers of true positives, false positives, true negatives, and false negatives. A total of 622 patients with TC, 547 patients with benign thyroid nodules, and 98 healthy individuals were included in 20 studies reported in 14 articles. The types of cfDNA included in the research include specific mutations of cfDNA, methylation of cfDNA, the content of cfDNA, and cfDNA index. After rigorous statistical analysis, the pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, diagnostic odds ratio, and area under the summary receiver operating characteristic curve were 0.76 (95% confidence interval [CI] 0.62-0.85), 0.87 (95% CI 0.78-0.93), 5.08 (95% CI 3.3-10.3), 0.28 (95% CI 0.17-0.46), 21 (95% CI 9-49), and 0.89 (95% CI 0.86-0.91), respectively. The meta-regression results showed that the number of cfDNAs, cfDNA methylation status, and sample size were the sources of heterogeneity in the specificity of the study. A subgroup analysis showed that the quantitative analysis group (cfDNA level) had a higher diagnostic accuracy than that of the qualitative analysis group (cfDNA methylation, mutation, or integrity index), with a sensitivity of 0.84, specificity of 0.89, and area under the curve of 0.91. The results of this meta-analysis suggest that cfDNA has value as an adjunct for the diagnosis of TC. Quantitative detection of cfDNA can achieve relatively high diagnostic accuracy. However, due to heterogeneity, the test results based on cfDNA for TC should be interpreted with caution.

Reduced methylation correlates with diabetic nephropathy risk in type 1 diabetes.

Diabetic nephropathy (DN) is a polygenic disorder with few risk variants showing robust replication in large-scale genome-wide association studies. To understand the role of DNA methylation, it is important to have the prevailing genomic view to distinguish key sequence elements that influence gene expression. This is particularly challenging for DN because genome-wide methylation patterns are poorly defined. While methylation is known to alter gene expression, the importance of this causal relationship is obscured by array-based technologies since coverage outside promoter regions is low. To overcome these challenges, we performed methylation sequencing using leukocytes derived from participants of the Finnish Diabetic Nephropathy (FinnDiane) type 1 diabetes (T1D) study (n = 39) that was subsequently replicated in a larger validation cohort (n = 296). Gene body-related regions made up more than 60% of the methylation differences and emphasized the importance of methylation sequencing. We observed differentially methylated genes associated with DN in 3 independent T1D registries originating from Denmark (n = 445), Hong Kong (n = 107), and Thailand (n = 130). Reduced DNA methylation at CTCF and Pol2B sites was tightly connected with DN pathways that include insulin signaling, lipid metabolism, and fibrosis. To define the pathophysiological significance of these population findings, methylation indices were assessed in human renal cells such as podocytes and proximal convoluted tubule cells. The expression of core genes was associated with reduced methylation, elevated CTCF and Pol2B binding, and the activation of insulin-signaling phosphoproteins in hyperglycemic cells. These experimental observations also closely parallel methylation-mediated regulation in human macrophages and vascular endothelial cells.

Epigenome-wide meta-analysis of BMI in nine cohorts: Examining the utility of epigenetically predicted BMI

This study sought to examine the association between DNA methylation and body mass index (BMI) and the potential of BMI-associated cytosine-phosphate-guanine (CpG) sites to provide information about metabolic health. We pooled summary statistics from six trans-ethnic epigenome-wide association studies (EWASs) of BMI representing nine cohorts (n= 17,034), replicated these findings in the Women's Health Initiative (WHI, n= 4,822), and developed an epigenetic prediction score of BMI. In the pooled EWASs, 1,265 CpG sites were associated with BMI (p<1E-7) and 1,238 replicated in the WHI (FDR<0.05). We performed several stratified analyses to examine whether these associations differed between individuals of European and African descent, as defined by self-reported race/ethnicity. We found that five CpG sites had a significant interaction with BMI by race/ethnicity. To examine the utility of the significant CpG sites in predicting BMI, we used elastic net regression to predict log-normalized BMI in the WHI (80% training/20% testing). This model found that 397 sites could explain 32% of the variance in BMI in the WHI test set. Individuals whose methylome-predicted BMI overestimated their BMI (high epigenetic BMI) had significantly higher glucose and triglycerides and lower HDL cholesterol and LDL cholesterol compared to accurately predicted BMI. Individuals whose methylome-predicted BMI underestimated their BMI (low epigenetic BMI) had significantly higher HDL cholesterol and lower glucose and triglycerides. This study confirmed 553 and identified 685 CpG sites associated with BMI. Participants with high epigenetic BMI had poorer metabolic health, suggesting that the overestimation may be driven in part by cardiometabolic derangements characteristic of metabolic syndrome.

Locus-Specific Bisulfate NGS Sequencing of GSTP1, RNF219, and KIAA1539 Genes in the Total Pool of Cell-Free and Cell-Surface-Bound DNA in Prostate Cancer: A Novel Approach for Prostate Cancer Diagnostics.

The locus-specific methylation of three genes (GSTP1, RNF219, and KIAA1539, also known as FAM214B) in the total pool of blood cell-free DNA, including cell-free DNA from plasma and cell-surface-bound DNA, of patients with prostate cancer and healthy donors was studied on the MiSeq platform. Our study found a higher methylation index of loci for total cell-free DNA compared with cell-free DNA. For total cell-free DNA, the methylation of GSTP1 in each of the 11 positions provided a complete separation of cancer patients from healthy donors, whereas for cell-free DNA, there were no positions in the three genes allowing for such separation. Among the prostate cancer patients, the minimum proportion of GSTP1 genes methylated in any of the 17 positions was 12.1% of the total circulated DNA fragments, and the minimum proportion of GSTP1 genes methylated in any of the 11 diagnostically specific positions was 8.4%. Total cell-free DNA was shown to be more convenient and informative as a source of methylated DNA molecules circulating in the blood than cell-free DNA.

Global and promoter specific hypermethylation of tumor suppressor genes P16, SOCS1, and SHP1 in oral squamous cell carcinoma and oral submucous fibrosis.

Aberrant methylation pattern leads to altered gene expression, that is, involved in the transformation of various cancers, including oral squamous cell carcinoma (OSCC). In the present study, an attempt has been made to examine the association of global and promoter-specific methylation of tumor suppressor genes in patients with OSCC and oral submucous fibrosis (OSMF). Promoter-specific methylation of tumor suppressor genes P16, SOCS1, and SHP1 had been studied earlier for their aberrant methylation patterns in other cancers; however, these studies were mainly conducted in-vitro or in animal models, and as such, only a few studies are available on human samples. In the present study evaluation of promoter-specific methylation of genes P16, SOCS1, and SHP1 in 76 patients' blood and tissue samples was done and compared with methylation of 35 healthy control samples using qPCR. Further, these samples were analyzed for global methylation patterns using ELISA. The results have shown a significant decreasing trend of promoter methylation (OSCC > OSMF > Controls); the methylation indices (MI) were significantly higher in OSCC than in the controls. The median MI of three genes for OSCC were P16MI (0.96), SHP1MI (0.79), and SOCS1 (0.80). Similarly, median MIs for OSMF were P16MI (0.18), SHP1 MI (0.19), and SOCS1 MI (0.5) against controls with MI (0) for each of the three genes. The global methylation %mC values were 1.9, 0.5, and 0.1, respectively. The values of MI and %mC were found to correlate with various risk factors such as tobacco, smoking, and alcohol consumption, which are positively involved in OSMF pathogenesis followed by oral cancer progression. Further, the methylation trend in tissue was reflected in blood samples, proving a window for methylation load to be used as a lesser invasive biomarker. The sensitivity and specificity of methylation load were also found reasonable. Therefore, the current study suggests that there may be a role of global and promoter-specific methylation load in the transition of OSMF to OSCC.

Quantitative analysis of MGMT promoter methylation in glioblastoma suggests nonlinear prognostic effect.

Epigenetic inhibition of the O6-methylguanine-DNA-methyltransferase (MGMT) gene has emerged as a clinically relevant prognostic marker in glioblastoma (GBM). Methylation of the MGMT promoter has been shown to increase chemotherapy efficacy. While traditionally reported as a binary marker, recent methodological advancements have led to quantitative methods of measuring promoter methylation, providing clearer insight into its functional relationship with survival. A CLIA assay and bisulfite sequencing was utilized to develop a quantitative, 17-point, MGMT promoter methylation index. GBMs of 240 newly diagnosed patients were sequenced and risk for mortality was assessed. Nonlinearities were captured by fitting splines to Cox proportional hazard models and plotting smoothed residuals. Covariates included age, Karnofsky performance status, IDH1 mutation, and extent of resection. Median follow-up time and progression-free survival were 16 and 9 months, respectively. A total of 176 subjects experienced death. A one-unit increase in promoter CpG methylation resulted in a 4% reduction in hazard (95% CI 0.93-0.99, P < .005). GBM patients with low levels of promoter methylation (1-6 CpG sites) fared markedly worse (HR = 1.62, 95% CI 1.03-2.54, P < .036) than individuals who were unmethylated. Subjects with medium levels of promoter methylation (7-12 sites) had the greatest reduction in hazard (HR = 0.48, 95% CI 0.29-0.80, P < .004), followed by individuals in the highest promoter methylation tertile (HR = 0.62, 95% CI 0.40-0.97, P < .035). Our findings suggest that the relationship between the extent of MGMT promoter methylation and survival in GBM may be nonlinear. These findings challenge the current understanding of MGMT and underlines the clinical importance of determining its prognostic utility. Potential limitations include censoring, sample size, and extraneous mutations.

Biomarker-based quantitative constraints on maximal soil-derived brGDGTs in modern lake sediments

Branched glycerol dialkyl glycerol tetraethers (brGDGTs) are emerging as a powerful tool for quantifying past terrestrial temperature variations. In lake sediments, mixed lacustrine vs. soil sources of brGDGTs pose a challenge to quantitative application of this paleothermometer, and this effect has not yet been well-constrained. In this work, we analyzed concentrations and distributions of brGDGTs along with two other types of typical lipid biomarkers (isoGDGTs and long-chain n-alkanes) in modern sediments and catchment soils from 26 park lakes across a large climatic gradient in China. BrGDGTs in these lakes may largely represent the lacustrine signal due to the relatively small catchment area and high eutrophic condition. This is confirmed by the combined use of crenarchaeol, its isomer crenarchaeol′, and C33n-alkane as tracers of soil input for brGDGTs. This novel method quantifies the upper limit of soil brGDGT contribution to be less than 10% for most investigated lakes, with an average value of 7% ± 10%. Subsequently, the brGDGT signal of the assumed pure lacustrine origin was constrained by the subtle difference between the original sedimentary brGDGTs and those corrected by the maximum possible soil brGDGT input for those park lakes. A compilation of global freshwater lakes further shows that the correlation between the methylation index of brGDGTs (MBT or MBT′5ME) of their sediments and temperature matches that for the assumed pure lacustrine signals. Hence, soil contribution might be generally minor and may not significantly impact the quantitative application of the brGDGT paleothermometer for worldwide lakes. This study reinforces a foundation for this novel lacustrine paleothermometer to be applied in paleoclimate reconstructions, although factors affecting lacustrine brGDGT distributions within lakes still need to be better constrained.

Associations of arsenic exposure with blood pressure and platelet indices in pregnant women: A cross-sectional study in Wuhan, China

BackgroundEnvironmental inorganic arsenic (iAs) exposure is potentially related to abnormal blood pressure (BP) changes and abnormal platelet activation. However, limited epidemiological studies have explored the impacts of iAs exposure on platelet change mediated by BP, especially for pregnant women. ObjectivesOur purpose was to investigate the associations of arsenic exposure with blood pressure and platelet indices among pregnant women. MethodsThe present study population included 765 pregnant women drawn from a prospective birth cohort study in Wuhan, China, recruited between October 2013 and April 2016. Urine sampled in the second trimester were used to assess arsenic species concentrations. The relative distribution of urinary arsenic species was used to measure human methylation capacity. BP parameters and platelet indices originated from the medical record. We applied multivariable linear regression models to explore the cross-sectional relationships between urinary arsenic metabolites, BP parameters, and platelet indices. We utilized mediation analysis to investigate the impacts of arsenic exposure on platelet indices through BP as mediator variables. ResultsWe observed significant positive correlations between iAs and systolic BP (SBP), diastolic BP (DBP), and mean arterial pressure (MAP). Pregnant women with higher methylation capacity to metabolize iAs characterized by higher secondary methylation index (SMI) and total methylation index (TMI) had a more significant reduction in SBP, DBP, and MAP. Pregnant women with higher DBP and MAP had higher platelet counts (PLC). A decreased PLC was found in subjects wither higher SMI. Additionally, SMI was negatively linked to PLC mediated through MAP. ConclusionsObtained results suggested that higher methylation capacity to metabolize iAs might contribute to decreased PLC among pregnant women, and MAP might mediate the influence of SMI on PLC.

Plasma EBV DNA Methylation: Evidence of EBV Lytic Infection in HIV(+) EBV(+) Plasmablastic Lymphoma

Background: Among people living with HIV, plasmablastic lymphoma (PBL) and classical Hodgkin lymphoma (HL) are almost uniformly associated with Epstein-Barr virus (EBV). Immunohistochemical studies have demonstrated high levels of lytic gene expression in PL,1 whereas EBV(+) HL shows exclusively latent viral gene expression. EBV virion DNA is never CpG methylated, but tumor-derived plasma EBV is CpG methylated as previously shown by our group in EBV(+) HL and nasopharyngeal carcinoma (NPC).2 Others have also presented evidence that patterns of CpG methylation of plasma EBV DNA varies between people with NPC and non-malignant EBV(+) conditions.3 To better characterize the nature of plasma EBV in patients with lymphoma, we investigated EBV DNA copy number and EBV DNA methylation in HIV-associated PBL and HL. Methods: With appropriate approvals from the Human Research Ethics Committee of the University of the Witwatersrand and the Johns Hopkins Institutional Review Boards, we collected plasma from newly diagnosed untreated patients with HIV and PBL or HL in Johannesburg, South Africa. Diagnostic biopsy specimens were reviewed in Baltimore by a hematopathologist to confirm the diagnosis and Epstein-Barr encoding region (EBER) in situ hybridization was performed to determine EBV association. Plasma EBV copies were measured by standard qPCR targeting the BamW repeats. qPCR using the same primers was also performed after capture with methyl-DNA binding protein 2 (MBD2) magnetic beads, where the captured fraction represents methylated DNA and the flow-through represents unmethylated DNA. Results: We studied plasma from 7 patients with PBL and 12 patients with HL. Four of five PBL tumors were EBV (+) and eight of 10 HL tumors were EBV (+) by tissue EBER in situ hybridization. Analysis of plasma EBV DNA copy number showed overlapping levels of EBV DNA (median 2.7 Log10 copy/mL in PBL and 3.2 Log10 copy/mL in HL) (Fig. 1). Patients with at least 1.5 Log10 copy/mL of EBV underwent CpG methylation evaluation, which showed very different EBV CpG methylation indices. The EBV methylation index was highly variable in PBL (median of 51%) and was uniformly high in HL (median of 97%). Conclusions: Although plasma from plasmablastic lymphoma patients show high EBV copy number, the characteristic of that EBV DNA is very different from the plasma EBV detected in Hodgkin lymphoma patients. This difference is hypothesized to be related to evidence of viral lytic gene expression seen in PBL. The degree of plasma EBV DNA methylation may have broad implications for lymphoma diagnosis and subtyping.

Evaluation of cross-platform compatibility of a DNA methylation-based glucocorticoid response biomarker

BackgroundIdentifying blood-based DNA methylation patterns is a minimally invasive way to detect biomarkers in predicting age, characteristics of certain diseases and conditions, as well as responses to immunotherapies. As microarray platforms continue to evolve and increase the scope of CpGs measured, new discoveries based on the most recent platform version and how they compare to available data from the previous versions of the platform are unknown. The neutrophil dexamethasone methylation index (NDMI 850) is a blood-based DNA methylation biomarker built on the Illumina MethylationEPIC (850K) array that measures epigenetic responses to dexamethasone (DEX), a synthetic glucocorticoid often administered for inflammation. Here, we compare the NDMI 850 to one we built using data from the Illumina Methylation 450K (NDMI 450).ResultsThe NDMI 450 consisted of 22 loci, 15 of which were present on the NDMI 850. In adult whole blood samples, the linear composite scores from NDMI 450 and NDMI 850 were highly correlated and had equivalent predictive accuracy for detecting DEX exposure among adult glioma patients and non-glioma adult controls. However, the NDMI 450 scores of newborn cord blood were significantly lower than NDMI 850 in samples measured with both assays.ConclusionsWe developed an algorithm that reproduces the DNA methylation glucocorticoid response score using 450K data, increasing the accessibility for researchers to assess this biomarker in archived or publicly available datasets that use the 450K version of the Illumina BeadChip array. However, the NDMI850 and NDMI450 do not give similar results in cord blood, and due to data availability limitations, results from sample types of newborn cord blood should be interpreted with care.

Epigenetic and Proteomic Biomarkers of Elevated Alcohol Use Predict Epigenetic Aging and Cell-Type variation Better Than Self-Report.

Excessive alcohol consumption (EAC) has a generally accepted effect on morbidity and mortality, outcomes thought to be reflected in measures of epigenetic aging (EA). As the association of self-reported EAC with EA has not been consistent with these expectations, underscoring the need for readily employable non-self-report tools for accurately assessing and monitoring the contribution of EAC to accelerated EA, newly developed alcohol consumption DNA methylation indices, such as the Alcohol T Score (ATS) and Methyl DetectR (MDR), may be helpful. To test that hypothesis, we used these new indices along with the carbohydrate deficient transferrin (CDT), concurrent as well as past self-reports of EAC, and well-established measures of cigarette smoking to examine the relationship of EAC to both accelerated EA and immune cell counts in a cohort of 437 young Black American adults. We found that MDR, CDT, and ATS were intercorrelated, even after controlling for gender and cotinine effects. Correlations between EA and self-reported EAC were low or non-significant, replicating prior research, whereas correlations with non-self-report indices were significant and more substantial. Comparing non-self-report indices showed that the ATS predicted more than four times as much variance in EA, CDT4 cells and B-cells as for both the MDR and CDT, and better predicted indices of accelerated EA. We conclude that each of the non-self-report indices have differing predictive capacities with respect to key alcohol-related health outcomes, and that the ATS may be particularly useful for clinicians seeking to understand and prevent accelerated EA. The results also underscore the likelihood of substantial underestimates of problematic use when self-report is used and a reduction in correlations with EA and variance in cell-types.

Epigenetic potentiation of somatostatin-2 by guadecitabine in neuroendocrine neoplasias as a novel method to allow delivery of peptide receptor radiotherapy

BackgroundSomatostatin receptor-2 (SSTR2) is expressed on cell surface of neuroendocrine neoplasias; its presence is exploited for the delivery of peptide receptor radionuclide therapy (PRRT). Patients with no or low expression of SSTR2 are not candidates for PRRT. SSTR2 promotor undergoes epigenetic modification, known to regulate gene expression. We investigated whether the demethylation agent, guadecitabine, could enhance the expression of SSTR2 in NET models, using radioligand uptake/PET imaging as a biomarker of epigenetic modification. MethodsThe effects of guadecitabine on the transcriptional, translational, and functional regulation of SSTR2 both in vitro and in vivo using low (QGP-1) and high (BON-1) methylated neuroendocrine neoplasia models was characterised. Promotor region methylation profiling of clinical samples (n = 61) was undertaken. Safety of combination guadecitabine and PRRT was assessed in vivo. ResultsPyrosequencing of cell lines illustrated differential methylation indices – BON: 1 94%, QGP: 1 21%. Following guadecitabine treatment, a dose-dependent increase in SSTR2 in BON-1 at a transcriptional, translational, and functional levels using the SSTR2-directed radioligand, 18F-FET-βAG-TOCA ([18F]-FETO) (150% increase [18F]-FETO uptake, p < 0.05) was observed. In vivo, guadecitabine treatment resulted in a 70% increase in [18F]-FETO uptake in BON-1 tumour models compared models with low baseline percentage methylation (p < 0.05). No additive toxicity was observed with the combination treatment of PRRT and guadecitabine in vivo. Methylation index in clinical samples was 10.5% compared to 5.2% in controls (p = 0.03) and correlated with SSTR2 expression (Wilcoxon rank sign −3.75,p < 0.01). ConclusionGuadecitabine increases SSTR2 expression both in vitro and in vivo. The combination of demethylation agents with PRRT warrants further investigation.

Evaluation of low-to-moderate arsenic exposure, metabolism and skin lesions in a Turkish rural population exposed through drinking water.

There is no human data regarding the exposure, metabolism and potential health effects of arsenic (As) contamination in drinking water in the Central Anatolian region of Turkey. Residents in ten villages with drinking water of total As (T-As) level >50μg L-1 and 10-50μg L-1 were selected as an exposed group (n=420) and <10μg L-1 as an unexposed group (n=185). Time-weighted average-As (TWA-As) intake was calculated from T-As analysis of drinking water samples. Concentrations of T-As in urine and hair samples, urinary As species [i.e., As(III), As(V), MMA(V) and DMA(V], and some micronutrients in serum samples of residents of the study area were determined. Primary and secondary methylation indices (PMI and SMI, respectively) were assessed from urinary As species concentrations and the presence of skin lesion was examined. TWA-As intake was found as 75μg L-1 in the exposed group. Urinary and hair T-As and urinary As species concentrations were significantly higher in the exposed group (P<0.05). The PMI and SMI values revealed that methylation capacities of the residents were efficient and that there was no saturation in As metabolism. No significant increase was observed in the frequency of skin lesions (hyperpigmentation, hypopigmentation, keratosis) of the exposed group (P>0.05). Only frequency of keratosis either at the hand or foot was higher in individuals with hair As concentration >1μg g-1 (P<0.05). Individuals living in the study area were chronically exposed to low-to-moderate As due to geological contamination in drinking water. No significant increase was observed in the frequency of skin lesions. Because of the controversy surrounding the health risks of low-to-moderate As exposure, it is critical to initiate long-term follow-up studies on health effects in this region.