Utilization of absolute monocyte counts to predict cardiovascular events in people living with HIV.

OBJECTIVES: Cardiovascular risk is increased in people living with HIV (PLWH). In HIV-uninfected populations, total absolute monocyte count (AMC) has been shown to be predictive of future cardiovascular events (CVEs). We sought to determine whether AMC predicts CVEs in PLWH independent of established and HIV-related cardiovascular risk factors. METHODS: We identified all PLWH within the Partners HIV Cohort without factors that could confound the monocyte count. CVE was defined as fatal or non-fatal acute myocardial infarction or ischaemic stroke. Baseline-measured AMC was defined as the average of all outpatient AMC counts a year before and after the baseline date. Multivariable Cox proportional hazards models were used to assess the association of baseline AMC with CVEs. RESULTS: Our cohort consisted of 1980 patients, with median follow-up of 10.9 years and 182 CVEs. Mean (± SD) age was 41.9 ± 9.3 years; 73.0% were male. Mean CD4 count was 506.3 ± 307.1 cells/µL, 48% had HIV viral load (VL) < 400 copies/mL, and 87% were on antiretroviral therapy. Mean AMC was 0.38 × 103  ± 0.13 cells/µL. In multivariable modelling adjusted for traditional CV risk factors, CD4 cell count, and HIV VL, AMC quartile 2 (Q2) (HR = 1.01, P = 0.98), Q3 (HR = 1.07, P = 0.76), and Q4 (HR = 0.97, P = 0.89) were not significantly predictive of CVE compared with Q1. DISCUSSION: Baseline AMC was not associated with long-term CVEs in PLWH. AMC obtained in routine clinical encounters does not appear to enhance CV risk stratification in PLWH.

Predicting 10-year risk of recurrent cardiovascular events andcardiovascular interventions in patients with established cardiovascular disease: results from UCC-SMART and REACH.

BACKGROUND: Existing cardiovascular risk scores for patients with established cardiovascular disease (CVD) estimate residual risk of recurrent major cardiovascular events (MACE). The aim of the current study is to develop and externally validate a prediction model to estimate the 10-year combined risk of recurrent MACE and cardiovascular interventions (MACE+) in patients with established CVD. METHODS: Data of patients with established CVD from the UCC-SMART cohort (N = 8421) were used for model development, and patient data from REACH Western Europe (N = 14,528) and REACH North America (N = 19,495) for model validation. Predictors were selected based on the existing SMART risk score. A Fine and Gray competing risk-adjusted 10-year risk model was developed for the combined outcome MACE+. The model was validated in all patients and in strata of coronary heart disease (CHD), cerebrovascular disease (CeVD), peripheral artery disease (PAD). RESULTS: External calibration for 2-year risk in REACH Western Europe and REACH North America was good, c-statistics were moderate: 0.60 and 0.58, respectively. In strata of CVD at baseline good external calibration was observed in patients with CHD and CeVD, however, poor calibration was seen in patients with PAD. C-statistics for patients with CHD were 0.60 and 0.57, for patients with CeVD 0.62 and 0.61, and for patients with PAD 0.53 and 0.54 in REACH Western Europe and REACH North America, respectively. CONCLUSIONS: The 10-year combined risk of recurrent MACE and cardiovascular interventions can be estimated in patients with established CHD or CeVD. However, cardiovascular interventions in patients with PAD could not be predicted reliably.

Hexarelin protects cardiac H9C2 cells from angiotensin II-induced hypertrophy via the regulation of autophagy.

Hexarelin is a synthetic growth hormone-releasing peptide that exerts cardioprotective effects. Regulation of autophagy is known to be cardioprotective so this study examined the role of autophagy and potential regulatory mechanisms in hexarelin-elicited anti-cardiac hypertrophic action in cardiomyocytes subjected to hypertrophy. H9C2 cardiomyocytes were subjected to hypertrophy by angiotensin-II (Ang-II). Autophagic light chain-3 (LC3) and cytoskeletal proteins were determined by immunofluorescence assay. Autophagy was also detected using monodansylcadaverine (MDC) for autophagic vacuole visualization and Cyto-ID staining for autophagic flux measurement. Molecular changes were analysed by Western blotting and qRT-PCR. Apoptosis was evaluated using flow cytometry and TUNEL assay. ATP content and CCK-8 assay were used in assessing enhanced cell survival whilst oxidative stress was analysed by measuring malondialdehyde(MDA) and superoxide dismutase(SOD) levels. Ang-II induced cardiomyocyte hypertrophy, oxidative stress, apoptosis and decreased cell survival, all of which were significantly suppressed by hexarelin treatment which also enhanced autophagy in hypertrophic H9C2 cells. Furthermore, inhibition of hexarelin induced autophagy by 3-methyladenine (3MA) abolished the anti-hypertrophic function of hexarelin and also abrogated the protection of hexarelin against cell survival inhibition and apoptosis. Conversely, the application of autophagy stimulator rapamycin in H9C2 hypertrophic cells inhibited apoptosis, cell survival and reduced cell size as well. Additionally, hexarelin regulated the upstream signalling of autophagy by inhibiting the phosphorylation of mammalian target of rapamycin(mTOR). We propose that hexarelin plays a novel role of attenuating cardiomyocyte hypertrophy and apoptosis via an autophagy-dependent mechanism associated with the suppression of the mTOR signalling pathway.

Extraskeletal Calcifications in Hutchinson-Gilford Progeria Syndrome.

PURPOSE: Children with Hutchinson-Gilford progeria syndrome (HGPS), a rare premature aging disease, exhibit extraskeletal calcifications detected by radiographic analysis and on physical examination. The aim of this study was to describe the natural history and pathophysiology of these abnormal calcifications in HGPS, and to determine whether medications and/or supplements tested in clinical trials alter their development. METHODS: Children from two successive clinical trials administering 1) lonafarnib (n = 26) and 2) lonafarnib + pravastatin + zoledronic acid (n = 37) were studied at baseline (pre-therapy), one year on therapy, and at end-of-therapy (3.3-4.3 years after the baseline visit). Calcium supplementation (oral calcium carbonate) was administered during the first year of the second trial and was subsequently discontinued. Information on calcifications was obtained from physical examinations, radiographs, and serum and urinary biochemical measures. The mineral content of two skin-derived calcifications was determined by x-ray diffraction. RESULTS: Extraskeletal calcifications were detected radiographically in 12/39 (31%) patients at baseline. The odds of exhibiting calcifications increased with age (p = 0.045). The odds were unaffected by receipt of lonafarnib, pravastatin, and zoledronate therapies. However, administration of calcium carbonate supplementation, in conjunction with all three therapeutic agents, significantly increased the odds of developing calcifications (p = 0.009), with the odds plateauing after the supplement's discontinuation. Composition analysis of calcinosis cutis showed hydroxyapatite similar to bone. Although serum calcium, phosphorus, and parathyroid hormone (PTH) were within normal limits at baseline and on-therapy, PTH increased significantly after lonafarnib initiation (p < 0.001). Both the urinary calcium/creatinine ratio and tubular reabsorption of phosphate (TRP) were elevated at baseline in 22/39 (56%) and 31/37 (84%) evaluable patients, respectively, with no significant changes while on-therapy. The mean calcium × phosphorus product (Ca × Pi) was within normal limits, but plasma magnesium decreased over both clinical trials. Fibroblast growth factor 23 (FGF23) was lower compared to age-matched controls (p = 0.03). CONCLUSIONS: Extraskeletal calcifications increased with age in children with HGPS and were composed of hydroxyapatite. The urinary calcium/creatinine ratio and TRP were elevated for age while FGF23 was decreased. Magnesium decreased and PTH increased after lonafarnib therapy which may alter the ability to mobilize calcium. These findings demonstrate that children with HGPS with normal renal function and an unremarkable Ca × Pi develop extraskeletal calcifications by an unidentified mechanism that may involve decreased plasma magnesium and FGF23. Calcium carbonate accelerated their development and is, therefore, not recommended for routine supplementation in these children.

HFE gene polymorphism defined by sequence-based typing of the Brazilian population and a standardized nomenclature for HFE allele sequences.

The HFE molecule controls iron uptake from gut, and defects in the molecule have been associated with iron overload, particularly in hereditary hemochromatosis. The HFE gene including both coding and boundary intronic regions were sequenced in 304 Brazilian individuals, encompassing healthy individuals and patients exhibiting hereditary or acquired iron overload. Six sites of variation were detected: (1) H63D C>G in exon 2, (2) IVS2 (+4) T>C in intron 2, (3) a C>G transversion in intron 3, (4) C282Y G>A in exon 4, (5) IVS4 (-44) T>C in intron 4, and (6) a new guanine deletion (G>del) in intron 5, which were used for haplotype inference. Nine HFE alleles were detected and six of these were officially named on the basis of the HLA Nomenclature, defined by the World Health Organization (WHO) Nomenclature Committee for Factors of the HLA System, and published via the IPD-IMGT/HLA website. Four alleles, HFE*001, *002, *003, and *004 exhibited variation within their exon sequences.

Plasminogen activator inhibitor and the risk of cardiovascular disease: The Framingham Heart Study.

INTRODUCTION: Although plasminogen activator inhibitor (PAI-1) plays a key regulatory role in fibrinolysis, it has not been clearly shown to independently predict cardiovascular disease (CVD) among individuals without prior CVD. We investigated, in the Framingham Heart Study offspring cohort, whether PAI-1 predicted CVD risk among individuals without prior CVD. METHODS: Plasma PAI-1 antigen and tissue plasminogen activator (TPA) antigen were measured in 3203 subjects without prior CVD between 1991 and 1995; average follow-up of 10 years. PAI-1 was remeasured 4 years after baseline, to determine the effect of serial change on risk. RESULTS: PAI-1 levels (mean ± SD) were 29.1 ng/ml (19.2) versus 22.1 (16.5) for those and without incident CVD; p<0.001, and TPA levels were 12.0 ng/ml (5.7) versus 9.0 (4.7); p<0.001. PAI-1 and TPA antigen levels had a strong unadjusted linear relation with incident CVD (p<0.001). After adjustment for conventional risk factors, the hazard ratios (HRs) for higher quartiles of PAI-1, compared with the lowest, were 1.9, 1.9, 2.6 (linear trend p=0.006), and 1.6, 1.6, 2.9 (p<0.001) for TPA antigen. The adjusted HRs for increasing quartiles of serial change in PAI-1 at 4 years, compared with the lowest, were 0.9, 0.8, 1.3 (p=0.050). C statistic assessment showed that adding PAI-1 or TPA to conventional risk factors resulted in small increases in discrimination and modest reclassification of risk, which was statistically significant for TPA (net reclassification 6.8%, p=0.037) but not PAI-1 (4.8%, p=0.113). CONCLUSION: PAI-1 and TPA antigen levels are predictive of CVD events after accounting for established risk factors. A serial increase in PAI-1 is associated with a further increase in risk. These findings support the importance of fibrinolytic potential in CVD.

Increased visit-to-visit blood pressure variability is associated with worse cardiovascular outcomes in low ejection fraction heart failure patients: Insights from the HEAAL study.

BACKGROUND: Recent data have highlighted shortcomings of the usual blood pressure (BP) hypothesis in several populations, and emphasized the importance of visit-to-visit variability of BP in predicting cardiovascular events. Herein, we aimed at assessing the association between visit-to-visit BP variability and outcomes in chronic heart failure (CHF) patients enrolled in the Heart failure Endpoint evaluation of Angiotensin II Antagonist Losartan (HEAAL). METHODS AND RESULTS: The HEAAL study randomized 3834 patients with HF and reduced ejection fraction administered 150 mg or 50mg losartan daily in a double blind, randomized, controlled trial. The patients were followed up for up to 6.8 years after randomization, and BP was measured at 3 time points in the first year and at semi-annual visits in the years thereafter. Three measures of visit-to-visit BP variability were computed for each subject: the standard deviation, the coefficient of variation and the average absolute visit-to-visit variation. Cox proportional hazard models were used to investigate the relationship between variations in systolic blood pressure, baseline covariates and the time to death or heart failure hospitalization (i.e. primary outcome). In multivariate analyses stratified on baseline BP, the patients with higher visit-to visit BP variability exhibited poorer outcomes (average absolute difference in SBP in mmHg:hazard ratio: 1.023 [95% CI (1.013, 1.034), P<0.0001]), independent from high dose losartan (still beneficial). CONCLUSIONS: For the first time, visit-to-visit BP variability was found elevated in CHF patients with reduced ejection fraction, and associated with poorer cardiovascular outcomes. Such assessments should be prioritized for testing prevention strategies in CHF. CLINICAL TRIAL REGISTRATION: This study is registered with the ClinicalTrials.gov, number NCT00090259.

Incident hyperkalemia may be an independent therapeutic target in low ejection fraction heart failure patients: insights from the HEAAL study.

BACKGROUND: Angiotensin receptor antagonists (ARBs) improve outcomes in patients with heart failure (HF) with reduced left ventricular ejection fraction, but may induce hyperkalemia (HK) and/or a worsening of renal function (WRF). METHODS AND RESULTS: The incidence and risk factors of HK and its inter-relationship with WRF, as well as associations with clinical outcome (death or admission for HF i.e. the primary outcome) in 3846 HF patients enrolled in the double blind HEAAL trial (losartan 150 mg/d vs. 50 mg/d) were assessed. Worsening of renal function was defined as a decrease in eGFR >20% from baseline and HK as serum K >5.5 or >5 mmol/L. Higher dose of losartan increased serum potassium. Episodes of HK >5 mmol/L or WRF occurred at least once in about half of the patients. WRF was associated with higher occurrence of HK (HR 1.19 (1.06-1.34)) and vice versa (HR 1.35 (1.19-1.53)), but preceded HK in only about half of the events. High dose losartan improved outcome despite more frequent WRF and HK, both being independently associated with adverse outcomes in multivariate analyses. CONCLUSIONS: HK and WRF are common in HF patients. Both can be predicted from baseline risk factors and are therefore potentially preventable. Although associated with worse outcome, occurrence of any does not hinder the efficacy of high dose losartan. HK was associated with WRF and worse outcomes. Whether therapy targeting specifically HK may maximize the survival benefit derived from renin angiotensin aldosterone inhibitor use should be appropriately tested in future trials.

A systems analysis of age-related changes in some cardiac aging traits.

Aging process or senescence affects the expression of a wide range of phenotypic traits throughout the life span of organisms. These traits often show modular, synergistic, and even antagonistic relationships, and are also influenced by genomic, developmental, physiological and environmental factors. The cardiovascular system (CVS) in humans represents a major modular system in which the relationships among physiological, anatomical and morphological traits undergo continuous remodeling throughout the life span of an individual. Here we extend the concept of developmental plasticity in order to study the relationships among 14 traits measured on 3,412 individuals from the Framingham Heart Study cohort, relative to age and gender, using exploratory structural equation modeling-a form of systems analysis. Our results reveal differing patterns of association among cardiac traits in younger and older persons in both sexes, indicating that physiological and developmental factors may be channeled differentially in relation to age and gender during the remodeling process. We suggest that systems approaches are necessary in order to understand the coordinated functional relationships among traits of the CVS over the life course of individuals.

Worldwide genetic variation at the 3' untranslated region of the HLA-G gene: balancing selection influencing genetic diversity.

The HLA-G (human leukocyte antigen-G) molecule plays a pivotal role in immune tolerance by inhibiting different cell subsets involved in both innate and adaptive immunity. Besides its primary function in maintaining the maternal-fetal tolerance, HLA-G has been involved in a wide range of pathological conditions where it can be either favorable or detrimental to the patient, depending on the nature of the pathology. Although several studies have demonstrated the utmost importance of the 3' untranslated region (3'UTR) in the HLA-G expression profile, limited data exist on the sequence variability of this gene region in human populations. In this study, we characterized the genetic diversity and haplotype structure of the HLA-G 3'UTR by resequencing 444 individuals from three sub-Saharan African populations and retrieving data from the 1000 Genomes project and the literature. A total of 1936 individuals representing 21 worldwide populations were combined and jointly analyzed. Our data revealed a high level of nucleotide diversity, an excess of intermediate frequency variants and an extremely low population differentiation, strongly supporting a history of balancing selection at this locus. The 14-bp insertion/deletion polymorphism was further pointed out as the likely target of selection, emphasizing its potential role in the post-transcriptional regulation of HLA-G expression.

Association of HLA-G 3' untranslated region polymorphisms with antibody response against Plasmodium falciparum antigens: preliminary results.

Host and Plasmodium interactions result in highly variable clinical phenotypes, partly explained by the nature and level of anti-malarial antibody response. Human leukocyte antigen (HLA)-G can create a tolerogenic environment, allowing parasites to escape from anti-malarial immunity. We performed a family-based association study encompassing 483 Sereer individuals (261 children and their parents), and reported two independent signals at the HLA-G 3' untranslated region associated with antibody response to specific Plasmodium falciparum blood stage antigens, previously associated with malaria protection: (i) +3010G together with +3142C with total IgG and IgG1 against GLURP and (ii) +3196G with IgG3 against MSP2. While these results require further investigation, they suggest for the first time a role of HLA-G in the regulation of humoral immune response in malaria.

The ratio of visceral to subcutaneous fat, a metric of body fat distribution, is a unique correlate of cardiometabolic risk.

AIMS/HYPOTHESIS: The anatomic location of excess body fat has an impact on associated cardiometabolic morbidity, and visceral adipose tissue (VAT) is more pathogenic than subcutaneous adipose tissue (SAT). However, VAT or SAT alone provides little information regarding the relative distribution of body fat. We hypothesised that the propensity to store energy in VAT relative to SAT depots may be a correlate of cardiometabolic risk, and tested this hypothesis using the VAT/SAT ratio as a metric of fat distribution. METHODS: We investigated associations of the VAT/SAT ratio with cardiometabolic traits in 3,223 participants (48% women) from the Framingham Heart Study. Fat depots were quantified by multidetector computed tomography (CT) scanning. RESULTS: In women and men, higher VAT/SAT ratio was associated (p < 0.05) with most assessed cardiovascular risk factors reflecting blood pressure, dyslipidaemia and insulin resistance. Additional adjustment for BMI did not materially change the findings in women, and generally strengthened associations in men. Further adjustment for VAT attenuated some associations in women, but those with lower HDL-cholesterol, higher triacylglycerol (both p < 0.0001) and higher prevalence of hypertension (p = 0.02), diabetes (p = 0.01) and the metabolic syndrome (p = 0.005) remained significant. Similarly, in men, associations with higher systolic (p = 0.006) and diastolic blood pressure (p = 0.03), higher fasting glucose (p = 0.0005), lower HDL-cholesterol and higher triacylglycerol (both p < 0.0001) and higher prevalence of diabetes (p = 0.006) remained significant. CONCLUSIONS/INTERPRETATION: VAT/SAT ratio is a correlate of cardiometabolic risk, above and beyond BMI and VAT. The propensity to store fat viscerally versus subcutaneously may be a unique risk factor independent of absolute fat volumes.

Progesterone vs placebo therapy for women with epilepsy: A randomized clinical trial.

OBJECTIVE: To assess progesterone treatment of intractable seizures in women with partial epilepsy. METHODS: This randomized, double-blind, placebo-controlled, phase III, multicenter, clinical trial compared the efficacy and safety of adjunctive cyclic natural progesterone therapy vs placebo treatment of intractable seizures in 294 subjects randomized 2:1 to progesterone or placebo, stratified by catamenial and noncatamenial status. It compared treatments on proportions of ≥50% responders and changes in seizure frequency from 3 baseline to 3 treated menstrual cycles. RESULTS: There was no significant difference in proportions of responders between progesterone and placebo in the catamenial and noncatamenial strata. Prespecified secondary analysis showed that the level of perimenstrual seizure exacerbation (C1 level) was a significant predictor of responders for progesterone but not placebo. With increasing C1 levels, responders increased from 21% to 57% with progesterone vs 19% to 20% with placebo. Reductions in seizure frequency correlated with increasing C1 levels for progesterone but not placebo, progressing from 26% to 71% for progesterone vs 25% to 26% for placebo. A prespecified clinically important separation between progesterone and placebo responders (37.8% vs 11.1%; p = 0.037) was realized among 21.4% of women who had C1 level ≥3. CONCLUSION: There was no difference in the primary outcome of ≥50% responder rates between progesterone vs placebo for catamenial or noncatamenial groups. Post hoc findings suggest that the level of perimenstrual seizure exacerbation is a significant predictor of responder rate with progesterone and that progesterone may provide clinically important benefit for a subset of women with perimenstrually exacerbated seizures. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that cyclic progesterone is ineffective in women with intractable partial epilepsy. Post hoc analysis identified a subset of women with higher levels of perimenstrual seizure exacerbation that were responsive to treatment.

Analysis of five polymorphic DNA markers for indirect genetic diagnosis of haemophilia A in the Brazilian population.

Hemophilia A is an X-linked, inherited, bleeding disorder caused by the partial or total inactivity of the coagulation factor VIII (FVIII). Due to difficulties in the direct recognition of the disease-associated mutation in the F8 gene, indirect diagnosis using polymorphic markers located inside or close to the gene is used as an alternative for determining the segregation of the mutant gene within families and thus for detecting carrier individuals and/or assisting in prenatal diagnosis. This study characterizes the allelic and haplotype frequencies, genetic diversity, population differentiation and linkage disequilibrium of five microsatellites (F8Int1, F8Int13, F8Int22, F8Int25.3 and IKBKG) in samples of healthy individuals from São Paulo, Rio Grande do Sul and Pernambuco and of patients from São Paulo with haemophilia A to determine the degree of informativeness of these microsatellites for diagnostic purposes. The interpopulational diversity parameters highlight the differences among the analyzed population samples. Regional differences in allelic frequencies must be taken into account when conducting indirect diagnosis of haemophilia A. With the exception of IKBKG, all of the microsatellites presented high heterozygosity levels. Using the markers described, diagnosis was possible in 10 of 11 families. The F8Int22, F8Int1, F8Int13, F8Int25.3 and IKBKG microsatellites were informative in seven, six, five and two of the cases, respectively, demonstrating the effectiveness of using these microsatellites in prenatal diagnosis and in carrier identification in the Brazilian population.

Association between single-slice measurements of visceral and abdominal subcutaneous adipose tissue with volumetric measurements: the Framingham Heart Study.

OBJECTIVE: Volumetric visceral abdominal adipose tissue (VAT) and subcutaneous abdominal adipose tissue (SAT) as measured by computed tomography (CT) are associated with metabolic risk factors. We sought to identify the correlations of VAT and SAT between area-based measures at different anatomic locations with volumetric measurements to identify the optimal anatomic site, and to relate measurements at this site with metabolic risk factors. METHODS: We measured SAT and VAT volumes across the total imaging volume, whereas we measured SAT and VAT area at seven predefined anatomic landmarks in 200 participants from the Framingham Heart Study (mean age 54 years, 50% women) who underwent abdominal multi-detector CT. Correlation coefficients were used to assess the association between area measurements and volumes as well as metabolic risk factors stratified by gender. RESULTS: Area-based measurements of SAT and VAT obtained at all anatomic landmarks were strongly associated with SAT and VAT volumes (all r>0.93, P<0.0001 and r>0.87, P<0.0001, for women and men; respectively). Consistently, area-based measurements of SAT and VAT obtained at L(3/4) were most strongly associated with volumetric measured VAT and SAT independent of age (both r=0.99 in men, r=0.96 for SAT and r=0.99 for VAT in women, all P-value <0.0001) and were similarly correlated with risk factors compared with SAT and VAT volumes (all P<0.05 for fasting plasma glucose, triglycerides, high-density lipoprotein, systolic blood pressure). CONCLUSION: Among area-based measurements of SAT and VAT, those obtained at the level of L(3/4) were strongly associated with SAT and VAT volumes and cardio-metabolic risk factors in both men and women.

Novel measurements of periaortic adipose tissue in comparison to anthropometric measures of obesity, and abdominal adipose tissue.

BACKGROUND: Perivascular adipose tissue may be associated with the amount of local atherosclerosis. We developed a novel and reproducible method to standardize volumetric quantification of periaortic adipose tissue by computed tomography (CT) and determined the association with anthropometric measures of obesity, and abdominal adipose tissue. METHODS: Measurements of adipose tissue were performed in a random subset of participants from the Framingham Heart Study (n=100) who underwent multidetector CT of the thorax (ECG triggering, 2.5 mm slice thickness) and the abdomen (helical CT acquisition, 2.5 mm slice thickness). Abdominal periaortic adipose tissue (AAT) was defined by a 5 mm cylindrical region of interest around the aortic wall; thoracic periaortic adipose tissue (TAT) was defined by anatomic landmarks. TAT and AAT were defined as any voxel between -195 and -45 HU and volumes were measured using dedicated semiautomatic software. Measurement reproducibility and association with anthropometric measures of obesity, and abdominal adipose tissue were determined. RESULTS: The intra- and inter-observer reproducibility for both AAT and TAT was excellent (ICC: 0.97 and 0.97; 0.99 and 0.98, respectively). Similarly, the relative intra- and inter-observer difference was small for both AAT (-1.85+/-1.28% and 7.85+/-6.08%; respectively) and TAT (3.56+/-0.83% and -4.56+/-0.85%, respectively). Both AAT and TAT were highly correlated with visceral abdominal fat (r=0.65 and 0.77, P<0.0001 for both) and moderately correlated with subcutaneous abdominal fat (r=0.39 and 0.42, P<0.0001 and P=0.009), waist circumference (r=0.49 and 0.57, P<0.0001 for both) and body mass index (r=0.47 and 0.58, P<0.0001 for both). CONCLUSION: Standardized semiautomatic CT-based volumetric quantification of periaortic adipose tissue is feasible and highly reproducible. Further investigation is warranted regarding associations of periaortic adipose tissue with other body fat deposits, cardiovascular risk factors and clinical outcomes.

Genetic and non-genetic correlates of vitamins K and D.

OBJECTIVE: To assess the genetic and nongenetic correlates of circulating measures of vitamins K and D status in a community-based sample of men and women. SUBJECTS/METHODS: A cross-sectional study of 1762 participants of the Framingham Offspring Study (919 women; mean age 59 years). Vitamin K status was measured as plasma phylloquinone and serum percent undercarboxylated osteocalcin (ucOC), and vitamin D was measured using plasma 25-hydroxyvitamin D (25(OH)D). Associations between vitamin K status and vitamin D status with biologically plausible nongenetic factors were assessed using stepwise regression. Heritability and linkage were determined using Sequential Oligogenic Linkage Analysis Routines (SOLAR). RESULTS: Nongenetic factors accounted for 20.1 and 12.3% of the variability in plasma phylloquinone in men and women respectively, with triglycerides and phylloquinone intake being the primary correlates. In men 12.2% and in women 14.6% of the variability in %ucOC was explained by nongenetic factors in our models. Heritability estimates for these vitamin K status biomarkers were nonsignificant. Season, vitamin D intake, high-density lipoprotein (HDL) cholesterol and waist circumference explained 24.7% (men) and 24.2% (women) of the variability in plasma 25(OH)D. Of the three vitamins examined, only 25(OH)D was significantly heritable (heritability estimate=28.8%, P<0.01), but linkage analysis of 25(OH)D did not achieve genome-wide significance. CONCLUSIONS: Variability in biomarkers of vitamin K status was attributed to nongenetic factors, whereas plasma 25(OH)D was found to be significantly heritable. Further studies are warranted to investigate genetic loci influencing vitamin D status.

Inflammatory biomarkers are associated with total brain volume: the Framingham Heart Study.

BACKGROUND: Systemic inflammation is associated with ischemia and Alzheimer disease (AD). We hypothesized that inflammatory biomarkers would be associated with neuroimaging markers of ischemia (i.e., white matter hyperintensities [WMH]) and AD (i.e., total brain volume [TCB]). METHODS: MRI WMH and TCB were quantified on 1,926 Framingham Offspring participants free from clinical stroke, TIA, or dementia (mean age 60 +/- 9 years; range 35 to 85 years; 54% women) who underwent measurement of a circulating inflammatory marker panel, including CD40 ligand, C-reactive protein, interleukin-6 (IL-6), soluble intracellular adhesion molecule-1, monocyte chemoattractant protein-1, myeloperoxidase, osteoprotegerin (OPG), P-selectin, tumor necrosis factor-alpha (TNFalpha), and tumor necrosis factor receptor II. To account for head size, both TCB (TCBV) and WMH (WMH/TCV) were divided by total cranial volume. We used multivariable linear regression to relate 10 log-transformed inflammatory biomarkers to brain MRI measures. RESULTS: In multivariable models, inflammatory markers as a group were associated with TCBV (p < 0.0001) but not WMH/TCV (p = 0.28). In stepwise models adjusted for clinical covariates with backwards elimination of markers, IL-6 and OPG were inversely associated with TCBV; TNFalpha was inversely related to TCBV in a subset of 1,430 participants. Findings were similar in analyses excluding individuals with prevalent cardiovascular disease. The relations between TCBV and inflammatory markers were modified by both sex and age, and generally were more pronounced in men and in older individuals. CONCLUSIONS: Although our observational cross-sectional data cannot establish causality, they are consistent with the hypothesis that higher inflammatory markers are associated with greater atrophy than expected for age.

Comparison of anthropometric, area- and volume-based assessment of abdominal subcutaneous and visceral adipose tissue volumes using multi-detector computed tomography.

PURPOSE: Cross-sectional imaging may enable accurate localization and quantification of subcutaneous and visceral adipose tissue. The reproducibility of multi-detector computed tomography (MDCT)-based volumetric quantification of abdominal adipose tissue and the ability to depict age- and gender-related characteristics of adipose tissue deposition have not been reported. METHODS: We evaluated a random subset of 100 Caucasian subjects (age range: 37-83 years; 49% women) of the Framingham Heart Study offspring cohort who underwent MDCT scanning. Two readers measured subcutaneous and visceral adipose tissue volumes (SAV and VAV; cm(3)) and areas (SAA and VAA; cm(2)) as well as abdominal sagital diameter (SD) and waist circumference (WC). RESULTS: Inter-reader reproducibility was excellent (relative difference: -0.34+/-0.52% for SAV and 0.59+/-0.93% for VAV, intra-class correlation (ICC)=0.99 each). The mean SAA/VAA ratio was significantly different from the mean SAV/VAV ratio (2.0+/-1.2 vs 1.7+/-0.9; P<0.001). The ratio of SAV/VAV was only weakly inversely associated with SD (ICC=-0.32, P=0.01) and not significantly associated with WC (ICC=-0.14, P=0.14) or body mass index (ICC=-0.17, P=0.09). The mean SAV/VAV ratio was significantly different between participants <60 vs >60 years (1.9+/-1.0 vs 1.5+/-0.7; P<0.001) and between men and women (1.2+/-0.5 vs 2.2+/-0.9; P<0.001). CONCLUSION: This study demonstrates that MDCT-based volumetric quantification of abdominal adipose tissue is highly reproducible. In addition, our results suggest that volumetric measurements can depict age- and gender-related differences of visceral and subcutaneous abdominal adipose tissue deposition. Further research is warranted to assess whether volumetric measurements may substantially improve the predictive value of obesity measures for insulin resistance, type 2 diabetes mellitus and other diseases.

New developments in medical clinical trials.

This paper reviews several new developments and long-standing good practices for conducting clinical trials. Discussion starts with the need for clear statements of study objectives, proceeds to clarify target and sample population, and elaborates on primary vs. secondary variables with the need for alpha adjustment in the presence of multiple outcomes. Here we also review the issue of surrogate endpoints. Study design issues--including blinding, randomization, and multicenter studies--come next. Then we discuss the current trend of the replacement of placebo-controlled trials by active controlled non-inferiority trials, the increasing use of Independent Data Monitoring Committees, the prominence of analysis on Intention-to-Treat samples, and the importance of imputation of missing data. We close with a brief discussion of the unit of analysis, the role of newer statistical analysis methods, safety issues, subset analysis, and, most importantly, clinical significance.