The prognostic significance of whole blood global and specific DNA methylation levels in gastric adenocarcinoma

Background

Epigenetics, particularly DNA methylation, has recently been elucidated as important in gastric cancer (GC) initiation and progression. We investigated the clinical and prognostic importance of whole blood global and site-specific DNA methylation in GC.

Methods

Genomic DNA was extracted from the peripheral blood of 105 Omani GC patients at diagnosis. DNA methylation was quantified by pyrosequencing of global DNA and specific gene promoter regions at 5 CpG sites for CDH1, 7 CpG sites for p16, 4 CpG sites for p53, and 3 CpG sites for RUNX3. DNA methylation levels in patients were categorized into low, medium, and high tertiles. Associations between methylation level category and clinicopathological features were evaluated using χ² tests. Survival analyses were carried out using the Kaplan-Meier method and log rank test. A backward conditional Cox proportional hazards regression model was used to identify independent predictors of survival.

Results

Older GC patients had increased methylation levels at specific CpG sites within the CDH1, p53, and RUNX-3 promoters. Male gender was significantly associated with reduced global and increased site-specific DNA methylation levels in CDH1, p16, and p53 promoters. Global DNA low methylation level was associated with better survival on univariate analysis. Patients with high and medium methylation vs. low methylation levels across p16 promoter CpG sites, site 2 in particular, had better survival. Multivariate analysis showed that global DNA hypermethylation was a significant independent predictor of worse survival (hazard ratio (HR) = 2.0, 95% CI: 1.1–3.8; p = 0.02) and high methylation mean values across p16 promoter sites 1–7 were associated with better survival with HR of 0.3 (95% CI, 0.1–0.8; p = 0.02) respectively.

Conclusions

Analysis of global and site-specific DNA methylation in peripheral blood by pyrosequencing provides quantitative DNA methylation values that may serve as important prognostic indicators.     

Methylomic analysis identifies frequent DNA methylation of zinc finger protein 582 (ZNF582) in cervical neoplasms

Background

Despite of the trend that the application of DNA methylation as a biomarker for cancer detection is promising, clinically applicable genes are few. Therefore, we looked for novel hypermethylated genes for cervical cancer screening.

Methods and Findings

At the discovery phase, we analyzed the methylation profiles of human cervical carcinomas and normal cervixes by methylated DNA immunoprecipitation coupled to promoter tiling arrays (MeDIP-on-chip). Methylation-specific PCR (MSP), quantitative MSP and bisulfite sequencing were used to verify the methylation status in cancer tissues and cervical scrapings from patients with different severities. Immunohistochemical staining of a cervical tissue microarray was used to confirm protein expression. We narrowed to three candidate genes: DBC1, PDE8B, and ZNF582; their methylation frequencies in tumors were 93%, 29%, and 100%, respectively. At the pre-validation phase, the methylation frequency of DBC1 and ZNF582 in cervical scraping correlated significantly with disease severity in an independent cohort (n = 330, both P<0.001). For the detection of cervical intraepithelial neoplasia 3 (CIN3) and worse, the area under the receiver operating characteristic curve (AUC) of ZNF582 was 0.82 (95% confidence interval  = 0.76–0.87).

Conclusions

Our study shows ZNF582 is frequently methylated in CIN3 and worse lesions, and it is demonstrated as a potential biomarker for the molecular screening of cervical cancer.     

Global DNA hypomethylation in peripheral blood leukocytes as a biomarker for cancer risk: a meta-analysis

Background

Good biomarkers for early detection of cancer lead to better prognosis. However, harvesting tumor tissue is invasive and cannot be routinely performed. Global DNA methylation of peripheral blood leukocyte DNA was evaluated as a biomarker for cancer risk.

Methods

We performed a meta-analysis to estimate overall cancer risk according to global DNA hypomethylation levels among studies with various cancer types and analytical methods used to measure DNA methylation. Studies were systemically searched via PubMed with no language limitation up to July 2011. Summary estimates were calculated using a fixed effects model.

Results

The subgroup analyses by experimental methods to determine DNA methylation level were performed due to heterogeneity within the selected studies (p<0.001, I²: 80%). Heterogeneity was not found in the subgroup of %5-mC (p = 0.393, I²: 0%) and LINE-1 used same target sequence (p = 0.097, I²: 49%), whereas considerable variance remained in LINE-1 (p<0.001, I²: 80%) and bladder cancer studies (p = 0.016, I²: 76%). These results suggest that experimental methods used to quantify global DNA methylation levels are important factors in the association study between hypomethylation levels and cancer risk. Overall, cancer risks of the group with the lowest DNA methylation levels were significantly higher compared to the group with the highest methylation levels [OR (95% CI): 1.48 (1.28–1.70)].

Conclusions

Global DNA hypomethylation in peripheral blood leukocytes may be a suitable biomarker for cancer risk. However, the association between global DNA methylation and cancer risk may be different based on experimental methods, and region of DNA targeted for measuring global hypomethylation levels as well as the cancer type. Therefore, it is important to select a precise and accurate surrogate marker for global DNA methylation levels in the association studies between global DNA methylation levels in peripheral leukocyte and cancer risk.     

DNA methylation profiles of primary colorectal carcinoma and matched liver metastasis

Background

The contribution of DNA methylation to the metastatic process in colorectal cancers (CRCs) is unclear.

Methods

We evaluated the methylation status of 13 genes (MINT1, MINT2, MINT31, MLH1, p16, p14, TIMP3, CDH1, CDH13, THBS1, MGMT, HPP1 and ERα) by bisulfite-pyrosequencing in 79 CRCs comprising 36 CRCs without liver metastasis and 43 CRCs with liver metastasis, including 16 paired primary CRCs and liver metastasis. We also performed methylated CpG island amplification microarrays (MCAM) in three paired primary and metastatic cancers.

Results

Methylation of p14, TIMP3 and HPP1 in primary CRCs progressively decreased from absence to presence of liver metastasis (13.1% vs. 4.3%; 14.8% vs. 3.7%; 43.9% vs. 35.8%, respectively) (P<.05). When paired primary and metastatic tumors were compared, only MGMT methylation was significantly higher in metastatic cancers (27.4% vs. 13.4%, P = .013), and this difference was due to an increase in methylation density rather than frequency in the majority of cases. MCAM showed an average 7.4% increase in DNA methylated genes in the metastatic samples. The numbers of differentially hypermethylated genes in the liver metastases increased with increasing time between resection of the primary and resection of the liver metastasis. Bisulfite-pyrosequencing validation in 12 paired samples showed that most of these increases were not conserved, and could be explained by differences in methylation density rather than frequency.

Conclusions

Most DNA methylation differences between primary CRCs and matched liver metastasis are due to random variation and an increase in DNA methylation density rather than de-novo inactivation and silencing. Thus, DNA methylation changes occur for the most part before progression to liver metastasis.     

A Prospective Study of LINE-1DNA Methylation and Development of Adiposity in School-Age Children

Background

Repetitive element DNA methylation is related to prominent obesity-related chronic diseases including cancer and cardiovascular disease; yet, little is known of its relation with weight status. We examined associations of LINE-1 DNA methylation with changes in adiposity and linear growth in a longitudinal study of school-age children from Bogotá, Colombia.

Methods

We quantified methylation of LINE-1 elements from peripheral leukocytes of 553 children aged 5–12 years at baseline using pyrosequencing technology. Anthropometric characteristics were measured periodically for a median of 30 months. We estimated mean change in three age-and sex-standardized indicators of adiposity: body mass index (BMI)-for-age Z-score, waist circumference Z-score, and subscapular-to-triceps skinfold thickness ratio Z-score according to quartiles of LINE-1 methylation using mixed effects regression models. We also examined associations with height-for-age Z-score.

Results

There were non-linear, inverse relations of LINE-1 methylation with BMI-for-age Z-score and the skinfold thickness ratio Z-score. After adjustment for baseline age and socioeconomic status, boys in the lowest quartile of LINE-1 methylation experienced annual gains in BMI-for-age Z-score and skinfold thickness ratio Z-score that were 0.06 Z/year (P = 0.04) and 0.07 Z/year (P = 0.03), respectively, higher than those in the upper three quartiles. The relation of LINE-1 methylation and annual change in waist circumference followed a decreasing monotonic trend across the four quartiles (P trend = 0.02). DNA methylation was not related to any of the adiposity indicators in girls. There were no associations between LINE-1 methylation and linear growth in either sex.

Conclusions

Lower LINE-1 DNA methylation is related to development of adiposity in boys.     

Frequent and distinct aberrations of DNA methylation patterns in fibrolamellar carcinoma of the liver

Background

Gene silencing due to aberrant DNA methylation is a frequent event in hepatocellular carcinoma (HCC) and also in hepatocellular adenoma (HCA). However, very little is known about epigenetic defects in fibrolamellar carcinoma (FLC), a rare variant of hepatocellular carcinoma that displays distinct clinical and morphological features.

Methodology/Principal Findings

We analyzed the methylation status of the APC, CDH1, cyclinD2, GSTπ1, hsa-mir-9-1, hsa-mir-9-2, and RASSF1A gene in a series of 15 FLC and paired normal liver tissue specimens by quantitative high-resolution pyrosequencing. Results were compared with common HCC arising in non-cirrhotic liver (n = 10). Frequent aberrant hypermethylation was found for the cyclinD2 (19%) and the RASSF1A (38%) gene as well as for the microRNA genes mir-9-1 (13%) and mir-9-2 (33%). In contrast to common HCC the APC and CDH1 (E-cadherin) genes were found devoid of any DNA methylation in FLC, whereas the GSTπ1 gene showed comparable DNA methylation in tumor and surrounding tissue at a moderate level. Changes in global DNA methylation level were measured by analyzing methylation status of the highly repetitive LINE-1 sequences. No evidence of global hypomethylation could be found in FLCs, whereas HCCs without cirrhosis showed a significant reduction in global methylation level as described previously.

Conclusions

FLCs display frequent and distinct gene-specific hypermethylation in the absence of significant global hypomethylation indicating that these two epigenetic aberrations are induced by different pathways and that full-blown malignancy can develop in the absence of global loss of DNA methylation. Only quantitative DNA methylation detection methodology was able to identify these differences.     

Prolactin secretion in healthy adults is determined by gender, age and body mass index

Background

Prolactin (PRL) secretion is quantifiable as mean, peak and nadir PRL concentrations, degree of irregularity (ApEn, approximate entropy) and spikiness (brief staccato-like fluctuations).

Hypothesis

Distinct PRL dynamics reflect relatively distinct (combinations of) subject variables, such as gender, age, and BMI.

Location

Clinical Research Unit.

Subjects

Seventy-four healthy adults aged 22–77 yr (41 women and 33 men), with BMI 18.3–39.4 kg/m².

Measures

Immunofluorometric PRL assay of 10-min samples collected for 24 hours.

Results

Mean 24-h PRL concentration correlated jointly with gender (P<0.0001) and BMI (P = 0.01), but not with age (overall R² = 0.308, P<0.0001). Nadir PRL concentration correlated with gender only (P = 0.017) and peak PRL with gender (P<0.001) and negatively with age (P<0.003), overall R² = 0.325, P<0.0001. Forward-selection multivariate regression of PRL deconvolution results demonstrated that basal (nonpulsatile) PRL secretion tended to be associated with BMI (R² = 0.058, P = 0.03), pulsatile secretion with gender (R² = 0.152, P = 0.003), and total secretion with gender and BMI (R² = 0.204, P<0.0001). Pulse mass was associated with gender (P = 0.001) and with a negative tendency to age (P = 0.038). In male subjects older than 50 yr (but not in women) approximate entropy was increased (0.942±0.301 vs. 1.258±0.267, P = 0.007) compared with younger men, as well as spikiness (0.363±0.122 vs. 0463±2.12, P = 0.031). Cosinor analysis disclosed higher mesor and amplitude in females than in men, but the acrophase was gender-independent. The acrophase was determined by age and BMI (R² = 0.186, P = 0.001).

Conclusion

In healthy adults, selective combinations of gender, age, and BMI specify distinct PRL dynamics, thus requiring balanced representation of these variables in comparative PRL studies.     

LINE-1 methylation levels in leukocyte DNA and risk of renal cell cancer

Purpose

Leukocyte global DNA methylation levels are currently being considered as biomarkers of cancer susceptibility and have been associated with risk of several cancers. In this study, we aimed to examine the association between long interspersed nuclear elements (LINE-1) methylation levels, as a biomarker of global DNA methylation in blood cell DNA, and renal cell cancer risk.

Experimental Design

LINE-1 methylation of bisulfite-converted genomic DNA isolated from leukocytes was quantified by pyrosequencing measured in triplicate, and averaged across 4 CpG sites. A total of 328 RCC cases and 654 controls frequency-matched(2∶1) on age(±5years), sex and study center, from a large case-control study conducted in Central and Eastern Europe were evaluated.

Results

LINE-1 methylation levels were significantly higher in RCC cases with a median of 81.97% (interquartile range[IQR]: 80.84–83.47) compared to 81.67% (IQR: 80.35–83.03) among controls (p = 0.003, Wilcoxon). Compared to the lowest LINE-1 methylation quartile(Q1), the adjusted ORs for increasing methylation quartiles were as follows: OR(Q2) = 1.84(1.20−2.81), OR(Q3) = 1.72(1.11−2.65) and OR(Q4) = 2.06(1.34−3.17), with a p-trend = 0.004. The association was stronger among current smokers (p-trend<0.001) than former or never smokers (p-interaction = 0.03). To eliminate the possibility of selection bias among controls, the relationship between LINE-1 methylation and smoking was evaluated and confirmed in a case-only analysis, as well.

Conclusions

Higher levels of LINE-1 methylation appear to be positively associated with RCC risk, particularly among current smokers. Further investigations using both post- and pre-diagnostic genomic DNA is warranted to confirm findings and will be necessary to determine whether the observed differences occur prior to, or as a result of carcinogenesis.     

Association of the European Lactase Persistence Variant (LCT-13910 C>T Polymorphism) with Obesity in the Canary Islands

Background

European lactose tolerance genotype (LCT -13910 C>T, rs4988234) has been positively associated to body mass indexes (BMI) in a meta-analysis of 31,720 individuals of northern and central European descent. A strong association of lactase persistence (LP) with BMI and obesity has also been traced in a Spanish Mediterranean population. The aim of this study was to analyze a potential association of LP compared to lactase non-persistence (LNP) with BMI in inhabitants of the Canary Islands of Spain using Mendelian randomization.

Methods

A representative, randomly sampled population of adults belonging to the Canary Islands Nutrition Survey (ENCA) in Spain, aged 18–75 years (n = 551), was genotyped for the LCT – 13910 C>T polymorphism. Milk consumption was assessed by a validated questionnaire. Anthropometric variables were directly measured. WHO classification of BMI was used.

Results

LP individuals were significantly more obese than LNP subjects (χ² = 10.59; p<0.005). LP showed in a multivariate linear regression analysis showed a positive association of LP with BMI compared to LNP, (β = 0.96; 95% CI: 0.08–1.85, p = 0.033). In a multinomial logistic regression analysis normal range weight LP subjects showed an odds ratio for obesity of 2.41; 95%CI 1.39–418, (p = 0.002) compared to LNP.

Conclusions

The T-13910 of the allele LCT-13910 C>T polymorphism is positively associated with BMI. LP increases significantly the risk to develop obesity in the studied population. The LCT-13910 C>T polymorphism stands proxy for the lifetime exposure pattern, milk intake, that may increase susceptibility to obesity and to obesity related pathologies.     

Epigenetic subgroups of esophageal and gastric adenocarcinoma with differential GATA5 DNA methylation associated with clinical and lifestyle factors

Background

Adenocarcinomas located near the gastroesophageal junction have unclear etiology and are difficult to classify. We used DNA methylation analysis to identify subtype-specific markers and new subgroups of gastroesophageal adenocarcinomas, and studied their association with epidemiological risk factors and clinical outcomes.

Methodology/Principal Findings

We used logistic regression models and unsupervised hierarchical cluster analysis of 74 DNA methylation markers on 45 tumor samples (44 patients) of esophageal and gastric adenocarcinomas obtained from a population-based case-control study to uncover epigenetic markers and cluster groups of gastroesophageal adenocarcinomas. No distinct epigenetic differences were evident between subtypes of gastric and esophageal cancers. However, we identified two gastroesophageal adenocarcinoma subclusters based on DNA methylation profiles. Group membership was best predicted by GATA5 DNA methylation status. We analyzed the associations between these two epigenetic groups and exposure using logistic regression, and the associations with survival time using Cox regression in a larger set of 317 tumor samples (278 patients). There were more males with esophageal and gastric cardia cancers in Cluster Group 1 characterized by higher GATA5 DNA methylation values (all p<0.05). This group also showed associations of borderline statistical significance with having ever smoked (p-value = 0.07), high body mass index (p-value = 0.06), and symptoms of gastroesophageal reflux (p-value = 0.07). Subjects in cluster Group 1 showed better survival than those in Group 2 after adjusting for tumor differentiation grade, but this was not found to be independent of tumor stage.

Conclusions/Significance

DNA methylation profiling can be used in population-based studies to identify epigenetic subclasses of gastroesophageal adenocarcinomas and class-specific DNA methylation markers that can be linked to epidemiological data and clinical outcome. Two new epigenetic subgroups of gastroesophageal adenocarcinomas were identified that differ to some extent in their survival rates, risk factors of exposure, and GATA5 DNA methylation.     

Genetic variant of AMD1 is associated with obesity in urban Indian children

Background

Hyperhomocysteinemia is regarded as a risk factor for cardiovascular diseases, diabetes and obesity. Manifestation of these chronic metabolic disorders starts in early life marked by increase in body mass index (BMI). We hypothesized that perturbations in homocysteine metabolism in early life could be a link between childhood obesity and adult metabolic disorders. Thus here we investigated association of common variants from homocysteine metabolism pathway genes with obesity in 3,168 urban Indian children.

Methodology/Principal Findings

We genotyped 90 common variants from 18 genes in 1,325 children comprising of 862 normal-weight (NW) and 463 over-weight/obese (OW/OB) children in stage 1. The top signal obtained was replicated in an independent sample set of 1843 children (1,399 NW and 444 OW/OB) in stage 2. Stage 1 association analysis revealed association between seven variants and childhood obesity at P<0.05, but association of only rs2796749 in AMD1 [OR = 1.41, P = 1.5×10^-4] remained significant after multiple testing correction. Association of rs2796749 with childhood obesity was validated in stage 2 [OR = 1.28, P = 4.2×10^-3] and meta-analysis [OR = 1.35, P = 1.9×10^-6]. AMD1 variant rs2796749 was also associated with quantitative measures of adiposity and plasma leptin levels that was also replicated and corroborated in combined analysis.

Conclusions/Significance

Our study provides first evidence for the association of AMD1 variant with obesity and plasma leptin levels in children. Further studies to confirm this association, its functional significance and mechanism of action need to be undertaken.     

LINE-1 hypomethylation during primary colon cancer progression

Background

Methylation levels of genomic repeats such as long interspersed nucleotide elements (LINE-1) are representative of global methylation status and play an important role in maintenance of genomic stability. The objective of the study was to assess LINE-1 methylation status in colorectal cancer (CRC) in relation to adenomatous and malignant progression, tissue heterogeneity, and TNM-stage.

Methodology/Principal Findings

DNA was collected by laser-capture microdissection (LCM) from normal, adenoma, and cancer tissue from 25 patients with TisN0M0 and from 92 primary CRC patients of various TNM-stages. The paraffin-embedded tissue sections were treated by in-situ DNA sodium bisulfite modification (SBM). LINE-1 hypomethylation index (LHI) was measured by absolute quantitative analysis of methylated alleles (AQAMA) realtime PCR; a greater index indicated enhanced hypomethylation. LHI in normal, cancer mesenchymal, adenoma, and CRC tissue was 0.38 (SD 0.07), 0.37 (SD 0.09), 0.49 (SD 0.10) and 0.53 (SD 0.08), respectively. LHI was significantly greater in adenoma tissue compared to its contiguous normal epithelium (P = 0.0003) and cancer mesenchymal tissue (P<0.0001). LHI did not differ significantly between adenoma and early cancer tissue of Tis stage (P = 0.20). LHI elevated with higher T-stage (P<0.04), was significantly greater in node-positive than node-negative CRC patients (P = 0.03), and was significantly greater in stage IV than all other disease stages (P<0.05).

Conclusion/Significance

By using in-situ SBM and LCM cell selection we demonstrated early onset of LINE-1 demethylation during adenomatous change of colorectal epithelial cells and demonstrated that LINE-1 demethylation progression is linear in relation to TNM-stage progression.     

Change in DNA Methylation Patterns of SLC6A4 Gene in the Gastric Mucosa in Functional Dyspepsia

Background

The neurochemical serotonin (5-HT) is an important signaling molecule in the gastrointestinal motor and sensory functions. A key regulator of 5-HT levels is the transmembrane serotonin transporter (5-HTT; SLC6A4) that governs the reuptake of 5-HT. Recent studies have indicated 5-HTT expression may be regulated by epigenetic mechanisms. We investigated DNA methylation status of SLC6A4 gene in the gastric mucosa from functional dyspepsia (FD) because of their potential role in dyspeptic symptoms.

Methods

Endoscopic gastric biopsies were obtained from 78 subjects with no upper abdominal symptoms and 79 patients with FD. Bisulfite Pyrosequencing was carried out to determine the methylation status of promoter CpG islands (PCGIs), promoter non-CpG islands (PNCGIs) and gene body non-CpG islands (NPNCGIs) in the SLC6A4 gene. Gene expression was examined by real-time PCR.

Results

In overall, methylation level of PCGIs was significantly lower in FD compared to control subjects (p = 0.04). On the other hand, methylation level of NPNCGIs was significantly higher in FD compared to control subjects (p = 0.03). Lower methylation level in PNCGIs was highlighted in the patients with PDS (p = 0.01), while higher methylation level in NPNCGIs was more prominent in the patients with EPS (p = 0.017). Methylation levels of PCGIs and PNCGIs were inversely correlated, while methylation levels of NPNCGIs was positively correlated with SLC6A4 mRNA levels in FD patients.

Conclusions

Our data suggest that change in DNA methylation pattern of SLC6A4 in the gastric mucosa may have a role for developing FD. A role of epigenetics for developing FD needs to be further evaluated.     

Associations of insulin and insulin-like growth factors with physical performance in old age in the Boyd Orr and Caerphilly studies

Objective

Insulin and the insulin-like growth factor (IGF) system regulate growth and are involved in determining muscle mass, strength and body composition. We hypothesised that IGF-I and IGF-II are associated with improved, and insulin with worse, physical performance in old age.

Methods

Physical performance was measured using the get-up and go timed walk and flamingo balance test at 63–86 years. We examined prospective associations of insulin, IGF-I, IGF-II and IGFBP-3 with physical performance in the UK-based Caerphilly Prospective Study (CaPS; n = 739 men); and cross-sectional insulin, IGF-I, IGF-II, IGFBP-2 and IGFBP-3 in the Boyd Orr cohort (n = 182 men, 223 women).

Results

In confounder-adjusted models, there was some evidence in CaPS that a standard deviation (SD) increase in IGF-I was associated with 1.5% faster get-up and go test times (95% CI: −0.2%, 3.2%; p = 0.08), but little association with poor balance, 19 years later. Coefficients in Boyd Orr were in the same direction as CaPS, but consistent with chance. Higher levels of insulin were weakly associated with worse physical performance (CaPS and Boyd Orr combined: get-up and go time = 1.3% slower per SD log-transformed insulin; 95% CI: 0.0%, 2.7%; p = 0.07; OR poor balance 1.13; 95% CI; 0.98, 1.29; p = 0.08), although associations were attenuated after controlling for body mass index (BMI) and co-morbidities. In Boyd Orr, a one SD increase in IGFBP-2 was associated with 2.6% slower get-up and go times (95% CI: 0.4%, 4.8% slower; p = 0.02), but this was only seen when controlling for BMI and co-morbidities. There was no consistent evidence of associations of IGF-II, or IGFBP-3 with physical performance.

Conclusions

There was some evidence that high IGF-I and low insulin levels in middle-age were associated with improved physical performance in old age, but estimates were imprecise. Larger cohorts are required to confirm or refute the findings.     

Circulating vascular progenitor cells and central arterial stiffness in polycystic ovary syndrome

Objective

Subjects with Polycystic ovarian syndrome (PCOS) are at increased risk of Type 2 diabetes mellitus (T2DM). The mechanism of this enhanced risk is unclear. Circulating vascular progenitor cells (VPC) are immature bone marrow derived cells capable of differentiating into mature endothelial cells. VPC number/function and central arterial stiffness predict cardio-metabolic disease in at-risk populations.

Design

We studied VPC and arterial stiffness measures in non-obese PCOS subjects as compared to age and body mass index (BMI) matched healthy controls in a cross–sectional study.

Methods

Fourteen subjects with PCOS and 12 controls of similar age, BMI (all <30 kg/m²) and metabolic profile were studied. VPC number and in vitro function were studied by flow cytometry and tube formation assays respectively. Augmentation index (AIx), a measure of central arterial stiffness, and central (aortic) blood pressures (BP) were measured by applanation tonometry.

Results

Subjects with PCOS had a reduced number, mean±SEM, of circulating CD34⁺133⁺ VPCs (317.5±51.0 vs. 558.3±101.2, p = 0.03) and impaired in vitro tube formation (completed tube area 1.0±0.06 vs. 1.2±0.05×10⁶ µm² p = 0.02). PCOS subjects had significantly higher AIx (18.4±1.9% vs. 4.9±2.0%) and this difference remained significant even after adjustments for age, BMI and smoking (p = 0.003) in multivariate analyses. Central systolic and pulse pressure were higher in PCOS subjects but these differences were not statistically significant after adjustment for age. Brachial systolic and pulse pressures were similar. VPC number/function and arterial stiffness or BP measures were not correlated.

Conclusions

Non-obese PCOS is characterized by a reduced VPC number, impaired VPC function and increased central arterial stiffness. These changes in novel vascular risk markers may explain the enhanced risk of T2DM and CVD in PCOS.     

DNA methylation analysis of the angiotensin converting enzyme (ACE) gene in major depression

Background

The angiotensin converting enzyme (ACE) has been repeatedly discussed as susceptibility factor for major depression (MD) and the bi-directional relation between MD and cardiovascular disorders (CVD). In this context, functional polymorphisms of the ACE gene have been linked to depression, to antidepressant treatment response, to ACE serum concentrations, as well as to hypertension, myocardial infarction and CVD risk markers. The mostly investigated ACE Ins/Del polymorphism accounts for ∼40%–50% of the ACE serum concentration variance, the remaining half is probably determined by other genetic, environmental or epigenetic factors, but these are poorly understood.

Materials and Methods

The main aim of the present study was the analysis of the DNA methylation pattern in the regulatory region of the ACE gene in peripheral leukocytes of 81 MD patients and 81 healthy controls.

Results

We detected intensive DNA methylation within a recently described, functional important region of the ACE gene promoter including hypermethylation in depressed patients (p = 0.008) and a significant inverse correlation between the ACE serum concentration and ACE promoter methylation frequency in the total sample (p = 0.02). Furthermore, a significant inverse correlation between the concentrations of the inflammatory CVD risk markers ICAM-1, E-selectin and P-selectin and the degree of ACE promoter methylation in MD patients could be demonstrated (p = 0.01 - 0.04).

Conclusion

The results of the present study suggest that aberrations in ACE promoter DNA methylation may be an underlying cause of MD and probably a common pathogenic factor for the bi-directional relationship between MD and cardiovascular disorders.     

Gestational weight gain and body mass index in children: results from three german cohort studies

Introduction

Previous studies suggested potential priming effects of gestational weight gain (GWG) on offspring’s body composition in later life. However, consistency of these effects in normal weight, overweight and obese mothers is less clear.

Methods

We combined the individual data of three German cohorts and assessed associations of total and excessive GWG (as defined by criteria of the Institute of Medicine) with offspring’s mean body mass index (BMI) standard deviation scores (SDS) and overweight at the age of 5–6 years (total: n = 6,254). Quantile regression was used to examine potentially different effects on different parts of the BMI SDS distribution. All models were adjusted for birth weight, maternal age and maternal smoking during pregnancy and stratified by maternal pre-pregnancy weight status.

Results

In adjusted models, positive associations of total and excessive GWG with mean BMI SDS and overweight were observed only in children of non- overweight mothers. For example, excessive GWG was associated with a mean increase of 0.08 (95% CI: 0.01, 0.15) units of BMI SDS (0.13 (0.02, 0.24) kg/m² of ‘real’ BMI) in children of normal-weight mothers. The effects of total and excessive GWG on BMI SDS increased for higher- BMI children of normal-weight mothers.

Discussion

Increased GWG is likely to be associated with overweight in offspring of non-overweight mothers.