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The geographic variation patterns of vascular mortality and their major risk factors from 68 samples in Eurasia are described in this work. The goodness-of-fit tests and analysis of variance indicate significant differences in incidence of mortality from the studied diseases, as well as in risk factors among the various geographic regions in four age groups. Correlation analysis points out the two general tendencies for the majority of studied traits: (a) significant positive association with latitude and (b) significant negative correlation with longitude. In turn, one-dimensional correlograms showed no specific geographic pattern at least up to 3000 km for all studied variables. However, at the large geographic scale a long-distance differentiation pattern was indicated for total serum cholesterol and body mass index; regional patches--for total death rate, mortality rate from cardiovascular and ischaemic heart diseases, systolic and diastolic blood pressure; and a local patches pattern was detected for mortality rate from stroke. Two-dimensional correlograms uncovered three distinct and significant patterns of variation: (a) a north-south trend for total mortality rate, for death rate from cardiovascular and ischaemic heart diseases, for diastolic and systolic blood pressure and for body mass index; (b) a northwest-southeast pattern for mortality rate from cerebrovascular disease; and (c) local patches for total serum cholesterol.

Estrogen and related hormones activate estrogen receptors, which in turn regulate genes for some cardiovascular disease (CVD) risk factors. Variants of the estrogen receptor a gene (ESR1) have been investigated, in small study populations, in relation to coronary artery disease and variation in levels of high-density lipoprotein (HDL) cholesterol, but little is known about the effects of genetic variation in ESR1 on CVD risk. This prospective study attempted to determine whether the ESR1 c.454-397T>C polymorphism correlates with CVD risk. Participants were 1739 unrelated men and women, ranging in age from 28 to 62 years, from the population-based offspring cohort of the Framingham Heart Study. Total atherosclerotic CVD events (recognized or unrecognized myocardial infarction [MI], angina pectoris, coronary insufficiency, intermittent claudication, death from coronary heart disease, atherothrombotic stroke) were present in 178 individuals followed up from 1971 to 1998. Major atherosclerotic CVD (recognized acute MI, coronary insufficiency, fatal coronary heart disease, atherothrombotic stroke) was found in 83 participants. Fifty-nine persons were recognized as having acute MI. One fifth of those participating in this study were homozygous for the ESR1c.454-397C allele. After adjusting for numerous covariates (age, gender, body mass index, hypertension, diabetes, total and high-density lipoprotein cholesterol, cigarette smoking), the CC genotype correlated significantly with major atherosclerotic CVD (odds ratio, 2.0; 95% confidence interval [CI], 1.3-3.2) compared with the CT or TT genotype. The risk of MI was 3-fold greater in those with the CC genotype (95% CI, 1.7-5.2). Comparable results were obtained when analyzing only men. Total atherosclerotic CVD did not appear to be associated with genotype. This study disclosed an association between a common estrogen receptor genotype, ESR1c.454-397CC, and an increased risk of MI. It seems likely that variations in estrogen receptors could help to explain the apparent effects of combination hormone therapy on the risk of CVD in women.

Background: The efficacy of Tam varies widely. Tam is biotransformed to endoxifen and 4-OH-Tam by the cytochrome P450 2D6 enzymes. Studies on 2D6 variants as a predictor of Tam's efficacy show conflicting results. Compared to Tam, endoxifen and 4-OH-Tam have a 100-fold greater affinity for the estrogen receptor (ER) and in suppressing estrogen-dependent cell proliferation. However, endoxifen is present at concentrations up to 20-fold higher and is pharmacologically distinct from 4-OH-Tam. Endoxifen's effect on ERα-degredation and transcription is concentration dependent; serum endoxifen levels ≥40nM block ERα-mediated transcriptional activation and inhibit estrogen-induced breast cancer cell proliferation. Methods: Pts taking Tam 20 mg/day, for any reason, were potentially eligible. Serum Tam, Tam metabolites and nineteen 2D6 alleles were analyzed. 2D6 genotyping was performed by PCR-based procedures and metabolites were assessed on Liquid Chromatography/Tandem Mass Spectrometry. Pts were stratified by 2D6 function into 3 groups: extensive metabolizers (EM) who had normal enzyme activity; intermediate metabolizers (IM) who had reduced enzyme activity; and poor metabolizers (PM) who had no enzyme activity. Tam was increased to 30 mg/day if serum endoxifen < 40nMol. Serum Tam and Tam metabolites were measured every 2 weeks for 90 days. Results: 121 patients have enrolled to date. Age, ethnicity, BMI, concomitant medications, and adherence were recorded. 25/117 (21%) pts had low serum endoxifen; all of these pts were increased to Tam 30 mg/day. Of this cohort, 36% were PMs (PM/PM and PM/IM), 32% were IMs (EM/PM and IM/IM) and 32% were EMs (EM/EM and EM/IM). All patients’ serum endoxifen increased to >40nMol by day 30 (Diagram below). Conclusions: 1) Sub-therapeutic serum endoxifen can be found in any 2D6 phenotype. 2) It is possible to achieve therapeutic serum endoxifen/Tam metabolite levels by optimizing the dose of Tam. 3) Subsequently, it is likely that other gene variants are involved in Tam metabolism hence why 2D6 testing may lead to conflicting results. 4). Further investigation is warranted as personalizing the dose of Tam is feasible and may potentially improve the outcome of pts taking Tam regardless of 2D6 genotype. Figure available in online version. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-02-06.

Background & Aims Hepatopulmonary syndrome (HPS) affects 10%–30% of patients with cirrhosis and portal hypertension and significantly increases mortality. Studies in experimental models indicate that pulmonary angiogenesis contributes to the development of HPS, but pathogenesis in humans is poorly understood. We investigated genetic risk factors for HPS in patients with advanced liver disease. Methods We performed a multicenter case-control study of patients with cirrhosis being evaluated for liver transplantation. Cases had an alveolar-arterial oxygen gradient ≥ 15 mm Hg (or ≥20 mm Hg if age > 64 years) and contrast echocardiography with late appearance of microbubbles after venous injection of agitated saline (intrapulmonary vasodilatation); controls did not meet both criteria for case status. The study sample included 59 cases and 126 controls. We genotyped 1086 common single nucleotide polymorphisms (SNPs) in 94 candidate genes. Results Forty-two SNPs in 21 genes were significantly associated with HPS after adjustments for race and smoking. Eight genes had at least 2 SNPs associated with disease: CAV3 , ENG , NOX4 , ESR2 , VWF , RUNX1 , COL18A1 , and TIE1 . For example, rs237872 in CAV3 showed an odds ratio of 2.75 (95% confidence interval: 1.65–4.60, P = .0001) and rs4837192 in ENG showed an odds ratio of 0.35 (95% confidence interval: 0.14–0.89, P = .027). Furthermore, variation in CAV3 and RUNX1 was associated with HPS in gene-based analyses. Conclusions Polymorphisms in genes involved in the regulation of angiogenesis are associated with the risk of HPS. Further investigation of these biologic pathways might elucidate the mechanisms that mediate the development of HPS in certain patients with severe liver disease.

e13507 Background: The efficacy of Tam varies widely. Tam is biotransformed to endoxifen and 4-OH-Tam by the cytochrome P450 2D6 enzymes. Compared to Tam, endoxifen and 4-OH-Tam have a 100-fold gre...

The degeneration of intervertebral discs (IVD) causes a great deal of morbidity, loss of work, and medical expense each year. Ongoing research is directed toward treatment modalities which slow the catabolic process in the degenerating disc with a focus on gene therapy and molecular therapy. While these avenues do have promise, they are plagued with problems including difficulty with persistence of growth factor production and the immunogenic and oncogenic properties of viral

ContextEstrogen and related hormone therapies activate estrogen receptors, which in turn regulate genes for several cardiovascular disease (CVD) risk factors. Relatively little is known, however, about the impact of genetic variation in estrogen receptor α (ESR1) on CVD risk.ObjectiveTo investigate whether the ESR1 c.454-397T>C polymorphism is associated with CVD risk.Design, Setting, and ParticipantsProspective study of 1739 unrelated men and women from the population-based offspring cohort of the Framingham Heart Study, who were followed up from 1971 to 1998.Main Outcome MeasuresTotal atherosclerotic CVD events, defined as recognized or unrecognized myocardial infarction (MI), angina pectoris, coronary insufficiency, intermittent claudication, coronary heart disease death, or atherothrombotic stroke (n = 178); major atherosclerotic CVD, defined as recognized acute MI, coronary insufficiency, coronary heart disease death, or atherothrombotic stroke (n = 83); and recognized acute MI (n = 59).ResultsTwenty percent of participants (n = 352) were homozygous for the ESR1 c.454-397C allele. After adjustment for covariates (age, sex, body mass index, hypertension, diabetes mellitus, total cholesterol, high-density lipoprotein cholesterol, and cigarette smoking), the CC genotype was significantly associated with major atherosclerotic CVD, with an odds ratio of 2.0 (95% confidence interval [CI], 1.3-3.2; P = .004) compared with individuals with the CT or TT genotypes. Participants with the CC genotype had 3.0-fold greater odds of MI (95% CI, 1.7-5.2; PC is causally related to MI risk or in linkage disequilibrium with 1 or more causal variants remains to be determined. These findings support the importance of estrogen receptors in CVD susceptibility, especially in men. Estrogen receptor variation also has potential to explain recent conflicting data regarding the effects of hormone therapy on CVD susceptibility in women.

Background Some benefits of glucose-insulin-potassium (GIK) in patients with acute coronary syndromes (ACS) may be from an anti-inflammatory effect. The primary aim of this study was to assess the impact of GIK administration early in the course of ACS on inflammatory marker C-reactive protein (CRP) levels. A secondary aim was to investigate the association between CRP and 30-day infarct size. Methods and Results Retrospective analysis of participants with ACS randomly assigned to GIK or placebo for at least 8 h in the IMMEDIATE Trial biological mechanism cohort (n = 143). High sensitivity CRP (hs-CRP) was measured at emergency department presentation, and 6 and 12 h into infusion. Logarithmically transformed hs-CRP values at 12-hours were lower with GIK vs. placebo (mean =0.65 mg/L in GIK, 0.84 mg/L in placebo), with a marginal trend toward significance (P = 0.053). Furthermore, using mixed models of hs-CRP, time, and study group, there was a significant increase in hs-CRP levels over time, but the rate of change did not differ between treatment arms (P = 0.3). Multivariable analysis showed that an elevation in hs-CRP, measured at 12 h, was an independent predictor of 30-day infarct size (β coefficient, 6.80; P = 0.04) using sestamibi SPECT imaging. Conclusions The results of this study show no significant effect of GIK on hs-CRP. In addition our results show that in patients with ACS, hs-CRP measured as early as 12 h can predict 30-day infarct size. Electronic supplementary material The online version of this article (doi:10.1186/s12872-015-0153-7) contains supplementary material, which is available to authorized users.

High-throughput bacterial 16S rRNA gene sequencing followed by clustering of short sequences into operational taxonomic units (OTUs) is widely used for microbiome profiling. However, clustering of short 16S rRNA gene reads into biologically meaningful OTUs is challenging, in part because nucleotide variation along the 16S rRNA gene is only partially captured by short reads. The recent emergence of long-read platforms, such as single-molecule real-time (SMRT) sequencing from Pacific Biosciences, offers the potential for improved taxonomic and phylogenetic profiling. Here, we evaluate the performance of long- and short-read 16S rRNA gene sequencing using simulated and experimental data, followed by OTU inference using computational pipelines based on heuristic and complete-linkage hierarchical clustering.In simulated data, long-read sequencing was shown to improve OTU quality and decrease variance. We then profiled 40 human gut microbiome samples using a combination of Illumina MiSeq and Blautia-specific SMRT sequencing, further supporting the notion that long reads can identify additional OTUs. We implemented a complete-linkage hierarchical clustering strategy using a flexible computational pipeline, tailored specifically for PacBio circular consensus sequencing (CCS) data that outperforms heuristic methods in most settings: https://github.com/oscar-franzen/oclust/ .Our data demonstrate that long reads can improve OTU inference; however, the choice of clustering algorithm and associated clustering thresholds has significant impact on performance.

Background DNA methylation has essential roles in transcriptional regulation, imprinting, X chromosome inactivation and other cellular processes, and aberrant CpG methylation is directly involved in the pathogenesis of human imprinting disorders and many cancers. To address the need for a quantitative and highly multiplexed bisulfite sequencing method with long read lengths for targeted CpG methylation analysis, we developed single-molecule real-time bisulfite sequencing (SMRT-BS). Results Optimized bisulfite conversion and PCR conditions enabled the amplification of DNA fragments up to ~1.5 kb, and subjecting overlapping 625–1491 bp amplicons to SMRT-BS indicated high reproducibility across all amplicon lengths (r = 0.972) and low standard deviations (≤0.10) between individual CpG sites sequenced in triplicate. Higher variability in CpG methylation quantitation was correlated with reduced sequencing depth, particularly for intermediately methylated regions. SMRT-BS was validated by orthogonal bisulfite-based microarray (r = 0.906; 42 CpG sites) and second generation sequencing (r = 0.933; 174 CpG sites); however, longer SMRT-BS amplicons (>1.0 kb) had reduced, but very acceptable, correlation with both orthogonal methods (r = 0.836-0.897 and r = 0.892-0.927, respectively) compared to amplicons less than ~1.0 kb (r = 0.940-0.951 and r = 0.948-0.963, respectively). Multiplexing utility was assessed by simultaneously subjecting four distinct CpG island amplicons (702–866 bp; 325 CpGs) and 30 hematological malignancy cell lines to SMRT-BS (average depth of 110X), which identified a spectrum of highly quantitative methylation levels across all interrogated CpG sites and cell lines. Conclusions SMRT-BS is a novel, accurate and cost-effective targeted CpG methylation method that is amenable to a high degree of multiplexing with minimal clonal PCR artifacts. Increased sequencing depth is necessary when interrogating longer amplicons (>1.0 kb) and the previously reported bisulfite sequencing PCR bias towards unmethylated DNA should be considered when measuring intermediately methylated regions. Coupled with an optimized bisulfite PCR protocol, SMRT-BS is capable of interrogating ~1.5 kb amplicons, which theoretically can cover ~91% of CpG islands in the human genome. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-1572-7) contains supplementary material, which is available to authorized users.