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Background Although a higher heart rate is associated with an increased risk of cardiovascular disease, the mechanism is not well understood. As thrombosis has an important role in plaque development and acute coronary syndromes, the increase related to heart rate may result from a prothrombotic imbalance. Methods We investigated the relation between heart rate and thrombotic potential in 3451 participants from the Offspring Cohort of the Framingham Heart Study (mean age 54 years, 55% women). Participants were divided into quintiles based on heart rate derived from a resting electrocardiogram. Results Higher heart rates were associated with significant age-adjusted increases in fibrinogen, viscosity, factor VII antigen, and impaired fibrinolytic potential (plasminogen activator inhibitor and tissue plasminogen activator antigen) among men and women, and von Willebrand factor antigen among men. Fibrinogen levels were 9% higher among men with a heart rate of 80.9 ± 8.1 beats/min (quintile 5) vs. 50.0 ± 3.9 beats/min (quintile 1) (314 vs. 287 mg/dl, p 0.001 for linear trend) and 13% higher among women (83.5 ± 7.7 beats/min vs. 53.7 ± 3.5 beats/min (330 vs. 291 mg/dl, p 0.001). The significant relations persisted after multivariate adjustment, other than among men, in whom factor VII was not significant and fibrinogen was borderline significant ( p = 0.065). Conclusions Higher heart rates are associated with a prothrombotic state. Because these factors are also associated with endothelial dysfunction and inflammation, these findings are consistent with an injurious effect of higher heart rates on the endothelium. Measures to reduce thrombotic potential may be of particular value in people with higher heart rates.

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.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.PAI-1 levels (mean ± SD) were 29.1 ng/ml (19.2) versus 22.1 (16.5) for those and without incident CVD; p0.001, and TPA levels were 12.0 ng/ml (5.7) versus 9.0 (4.7); p0.001. PAI-1 and TPA antigen levels had a strong unadjusted linear relation with incident CVD (p0.001). 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 (p0.001) 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).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.

SummaryObjective and design We explored the cross-sectional and longitudinal associations of thyroid function within the normal range with cardiovascular disease (CVD) risk factors and adiposity measures. Patients and measurements A total of 3483 (50·4% women) participants for the cross-sectional CVD study and 1630 (41·2% women) participants for the cross-sectional body composition substudy were drawn from the Framingham Third Generation Exam 1; 2912 participants (50·1% women) for the longitudinal CVD study and 713 participants (35·9% women) for the longitudinal body composition substudy were drawn from the Framingham Third Generation Exams 1–2. Thyroid function was assessed by thyrotropin [thyroid-stimulating hormone (TSH)] and free thyroxine (fT4) concentrations within the reference range at Exam 1. The associations between thyroid function and CVD risk factors were modelled via multivariable-adjusted regression models. Multivariable adjustment included age, sex, current smoking, postmenopausal status and BMI. Results Cross-sectionally, higher TSH concentration was associated with increased odds of hypertriglyceridaemia [odds ratio (OR)=1·10], and higher BMI (β = 0·19 kg/m2), total cholesterol (β = 0·05 mmol/l), triglycerides (β = 0·0006 mmol/l) and subcutaneous adipose tissue (SAT) volume (β = 38·8 cm3) (all P < 0·05). Cross-sectionally, fT4 was inversely associated with metabolic and adiposity-related CVD risk factors, including obesity (OR = 1·17), hypertriglyceridaemia (OR = 1·09), BMI (β = 0·42 kg/m2), total cholesterol (β = 0·05 mmol/l), triglycerides (β = 0·0002 mmol/l), visceral adipose tissue (VAT) volume (β = −20·7 cm3) and attenuation (0·17 HU) and VAT/SAT ratio (β = −0·01) (all P < 0·05). However, during 6·1 years of follow-up, baseline TSH and fT4 levels were not longitudinally associated with CVD risk factors and adiposity measures. Conclusions Thyroid function within the normal range is cross-sectionally, but not longitudinally, associated with CVD risk factors and adiposity measures.

Background Prolactin is an anterior pituitary hormone that may modulate the adverse effects of obesity. Prolactin has been associated with cardiovascular disease mortality, but less is known about whether prolactin predicts incidence of cardiovascular disease risk factors. Methods and Results Our sample (n=3232, mean age 40.4 years, 52.1% women) was drawn from Framingham Heart Study participants who attended 2 examinations an average of 6.1 years apart. After excluding those with elevated prolactin (>30 mg/dL for women, >20 mg/dL for men), multivariable‐adjusted regressions modeled the associations between baseline prolactin and changes in cardiovascular disease risk factors. Models were adjusted for age, sex, baseline value of the risk factor, smoking status, hormone replacement therapy, and menopausal status and additionally for body mass index. Mean prolactin levels were 11.9 mg/dL ( SD 5.2) in women and 8.0 mg/dL ( SD 2.9) in men. No associations were observed for change in weight, body composition, total cholesterol, triglycerides, or fasting glucose. In women, for example, for each 5‐mg/dL increment in prolactin, odds of incident hypercholesterolemia were 1.06, which was not significant (95% CI 0.91–1.23, P =0.46). Some exceptions were of note. In women, for each 5‐mg/dL increment in prolactin, we observed increased odds of low high‐density lipoprotein cholesterol at follow‐up (odds ratio 1.50, 95% CI 1.18–1.91, P =0.001) that persisted after adjustment for body mass index ( P =0.001). In men, a 5‐mg/dL increment in prolactin was associated with increased odds of incident hypertension (odds ratio 1.61, 95% CI 1.18–2.20 P =0.002) and incident diabetes (odds ratio 1.70, 95% CI 1.04–2.78, P =0.03). Conclusions Prolactin is not associated with a comprehensive panel of incident cardiovascular disease risk factors. Measurement of circulating prolactin levels in the community likely does not provide substantial insight into cardiometabolic risk.

Insulin resistance is associated with central obesity and an increased risk of cardiovascular disease. Our objective is to examine the association between abdominal subcutaneous (SAT) and visceral adipose tissue (VAT) and insulin resistance, to determine which fat depot is a stronger correlate of insulin resistance, and to assess whether there was an interaction between SAT, VAT, and age, sex, or BMI. Participants without diabetes from the Framingham Heart Study (FHS), who underwent multidetector computed tomography to assess SAT and VAT (n = 3,093; 48% women; mean age 50.4 years; mean BMI 27.6 kg/m2), were evaluated. Insulin resistance was measured using the homeostasis model and defined as HOMAIR ≥75th percentile. Logistic regression models, adjusted for age, sex, smoking, alcohol, menopausal status, and hormone replacement therapy use, were used to assess the association between fat measures and insulin resistance. The odds ratio (OR) for insulin resistance per standard deviation increase in SAT was 2.5 (95% confidence interval (CI): 2.2–2.7; P < 0.0001), whereas the OR for insulin resistance per standard deviation increase in VAT was 3.5 (95% CI: 3.1–3.9; P < 0.0001). Overall, VAT was a stronger correlate of insulin resistance than SAT (P < 0.0001 for SAT vs. VAT comparison). After adjustment for BMI, the OR of insulin resistance for VAT was 2.2 (95% CI: 1.9–2.5; P < 0.0001). We observed an interaction between VAT and BMI for insulin (P interaction = 0.0004), proinsulin (P interaction = 0.003), and HOMAIR (P interaction = 0.003), where VAT had a stronger association in obese individuals. In conclusion, SAT and VAT are both correlates of insulin resistance; however, VAT is a stronger correlate of insulin resistance than SAT.

Extensive efforts have been aimed at understanding the genetic underpinnings of complex diseases that affect humans. Numerous genome-wide association studies have assessed the association of genes with human disease, including the Framingham Heart Study (FHS), which genotyped 550,000 SNPs in 9,000 participants. The success of such efforts requires high rates of consent by participants, which is dependent on ethical oversight, communications, and trust between research participants and investigators. To study this we calculated percentages of participants who consented to collection of DNA and to various uses of their genetic information in two FHS cohorts between 2002 and 2009. The data included rates of consent for providing a DNA sample, creating an immortalized cell line, conducting research on various genetic conditions including those that might be considered sensitive, and for notifying participants of clinically significant genetic findings were above 95%. Only with regard to granting permission to share DNA or genetic findings with for-profit companies was the consent rate below 95%. We concluded that the FHS has maintained high rates of retention and consent for genetic research that has provided the scientific freedom to establish collaborations and address a broad range of research questions. We speculate that our high rates of consent have been achieved by establishing frequent and open communications with participants that highlight extensive oversight procedures. Our approach to maintaining high consent rates via ethical oversight of genetic research and communication with study participants is summarized in this report and should be of help to other studies engaged in similar types of research. Published 2010 Wiley-Liss, Inc.

Objective This analysis was undertaken to determine the long-term intraindividual variability, determinants of change, and capacity of the inflammatory marker C-reactive protein (CRP) to predict metabolic traits and diabetes in a large community-based population. Methods Intraindividual CRP variability, predictors of CRP change, and metabolic events were evaluated in the Framingham Heart Study Offspring cohort using data from the same 2409 participants with CRP measured by the same methodology at each of 3 examination cycles, spanning 20 years. Results Between the first and second examinations (averaging 16 years apart), 23% to 47% of men and 27% to 49% of women remained within the same quintile of CRP values. An additional 24% to 51% of men and 24% to 50% of women occupied an adjacent quintile. Intermediate-term CRP variability (over 4 years) was similar to long-term variability. Both long- and intermediate-term variability of CRP were significantly less than that of plasma cholesterol measured in these same groups. Linear regression models for CRP at the intermediate examination demonstrated that CRP at the initial examination contributed the largest proportion of the variability (partial R-square = 0.27) seen in the overall model after adjustment for other covariates known to affect CRP concentrations. Although logistic regression models demonstrated that CRP over the intermediate term did not predict new-onset metabolic syndrome at the final examination, CRP did predict an increase in glucose and new-onset diabetes. Conclusion The results of this longitudinal analysis suggest the intraindividual, long-term variability of CRP concentrations is relatively small and predictive of new diabetes over an intermediate-term of 4 years.

Cystatin C (CysC) is associated with cardiovascular disease (CVD) and chronic kidney disease (CKD). We examined the clinical correlates and heritability of CysC and determined if associations between CVD risk factors and CysC differed by CKD status. Among Framingham Heart Study offspring (examined from 1998-2001, n = 3,241, mean age 61 years, 54% women), the 95(th) percentile cut-point was developed for CysC in a healthy subset (n = 779) after excluding participants with diabetes, hypertension, low high-density lipoproteins, obesity, smoking, high triglycerides, prevalent CVD, and CKD (as defined by glomerular filtration rate60 mL/min per 1.73 m(2)). Multivariable logistic regression was used to evaluate the association between CVD risk factors and high CysC (CysCor =95(th) percentile cut-point). In a family-based subset (n = 1,188), we estimated CysC heritability using the variance-components method. The cut-point for high CysC was 1.07 mg/L. Age, hypertension treatment, low diastolic blood pressure, body mass index, low high-density lipoprotein cholesterol, and smoking were associated with high CysC in multivariable models. These factors and estimated glomerular filtration rate (egFR) explained 39.2% of CysC variability (R(2)). Excluding CKD did not materially change associations. Multivariable-adjusted heritability for CysC was 0.35 (p0.001). In conclusion, high CysC is associated with CVD risk factors even in the absence of CKD. The strong associations between CysC and CVD risk factors may partially explain why CysC is a strong predictor of incident CVD.

Background— Plasma levels of matrix metalloproteinase-9 (MMP-9), a key determinant of extracellular matrix degradation, are increased in heart failure and in acute coronary syndromes. We investigated cross-sectional relations of plasma MMP-9 to vascular risk factors and echocardiographic left ventricular (LV) measurements. Methods and Results— We studied 699 Framingham Study participants (mean age, 57 years; 58% women), free of heart failure and previous myocardial infarction, who underwent routine echocardiography. We examined sex-specific distributions of LV internal dimensions (LVEDD) and wall thickness (LVWT) and sampled persons with both LVEDD and LVWT below the sex-specific median (referent, n=299), with increased LVEDD (LVEDD ≥90th percentile, n=204) and increased LVWT (LVWT ≥90th percentile, n=221) in a 3:2:2 ratio. Plasma MMP-9 was detectable in 138 persons (20%). In multivariable models, increasing heart rate (OR per SD, 1.41; 95% CI, 1.17 to 1.71) and antihypertensive treatment (OR, 1.63; 95% CI, 1.06 to 2.50) were key clinical correlates of detectable plasma MMP-9. In multivariable-adjusted models, detectable plasma MMP-9 was associated with increased LVEDD (OR, 2.84; 95% CI, 1.13 to 7.11), increased LVWT (OR, 2.54; 95% CI, 1.00 to 6.46), and higher LV mass ( P =0.06) in men but not in women (OR for increased LVEDD, 1.37; 95% CI, 0.54 to 3.46; for increased LVWT, 0.99; 95% CI, 0.39 to 2.52; P =0.59 for LV mass). Conclusions— In our community-based sample, detectable plasma MMP-9 levels were associated with increased LV diastolic dimensions and increased wall thickness in men. These observations indicate that plasma MMP-9 level may be a marker for cardiac extracellular matrix degradation, a process involved in LV remodeling.

Assays for natriuretic peptides have received considerable attention as potential screening tests for congestive heart failure and left ventricular dysfunction. However, information regarding the impact of age, sex, and other physiologic characteristics on natriuretic peptide levels is limited. We examined a healthy reference sample of 911 subjects (mean age 55 years, 62% women) from the Framingham Heart Study who were free of hypertension, valvular disease, diabetes, atrial fibrillation, obesity, coronary heart disease, congestive heart failure, and renal failure, and who had normal left ventricular systolic function. Plasma brain natriuretic peptide and N-terminal atrial natriuretic peptide levels were measured, and multivariable regression used to assess correlates of natriuretic peptide levels. The strongest predictors of higher natriuretic peptide levels were older age and female sex. Other multivariable predictors included lower diastolic blood pressure (higher pulse pressure), lower body mass index, and higher left atrial size. Reference limits were then formulated based on the empirical distribution of natriuretic peptide levels by gender both across all ages and partitioned by age. Age-pooled reference limits compared with age-specific limits classified a higher proportion of healthy elderly subjects (17% vs 2.5%), but a lower proportion of healthy young subjects (1% vs 2.5%) as "abnormal." We conclude that interpretation of natriuretic peptide levels should take into consideration gender and possibly age. The reference limits derived from this large, healthy community-based sample will aid in the identification of elevated natriuretic peptide levels in clinical practice.

Background — Recent data suggest that the Pl A2 allele of the platelet glycoprotein IIIa receptor may be a genetic risk factor for cardiovascular disease. We previously reported that the Pl A2 allele was associated with increased platelet aggregability, as indicated by lower epinephrine threshold concentrations. Paradoxically, however, it has been reported that Pl A2 -positive platelets have reduced fibrinogen binding. Because fibrinogen mediates platelet aggregability, we hypothesized that plasma fibrinogen levels may interact with Pl A genotype in modulating platelet aggregability. Methods and Results — Glycoprotein IIIa Pl A genotype, fibrinogen level, and platelet aggregability were ascertained in 1340 subjects enrolled into the Framingham Offspring Study. Platelet aggregability was evaluated by the Born method. Higher fibrinogen levels were associated with increased epinephrine-induced aggregation ( P =0.002) and a trend for ADP-induced aggregation ( P =0.07). The fibrinogen effect was genotype specific, however, in that the increase in platelet aggregability with higher fibrinogen was present for the Pl A1/A1 genotype ( P =0.0005 and P =0.03 for epinephrine- and ADP-induced aggregation, respectively) but not for the Pl A2 -positive genotype ( P >0.90). Conclusion — Higher fibrinogen levels were associated with increased platelet aggregability. However, the association between fibrinogen and platelet aggregability was genotype specific. This interaction may be responsible for the conflicting findings regarding Pl A genotype and platelet aggregability. Further study of this gene-environment interaction may provide insight into cardiovascular disease risk.

Background —Hypertension is an established risk factor for acute coronary events. Because fibrinolytic and hemostatic factors are also associated with cardiovascular disease, we examined the relations of systolic and diastolic blood pressures (SBP and DBP) to levels of plasminogen activator inhibitor antigen, tissue plasminogen activator antigen, fibrinogen, factor VII, von Willebrand factor, fibrinogen, and plasma viscosity in subjects of the Framingham Offspring Study. Methods and Results —We studied 1193 men and 1459 women after the exclusion of subjects with known cardiovascular disease and those receiving anticoagulant or antihypertensive therapy. Linear regression models were used to evaluate SBP and DBP as predictors of fibrinolytic and hemostatic factor levels in separate sex models, with adjustment for age, body mass index, smoking, diabetes, total cholesterol, HDL, triglycerides, alcohol intake, and estrogen use (in women). In both sexes, levels of plasminogen activator inhibitor and tissue plasminogen activator antigen were positively related to SBP and DBP ( P P =0.008) and DBP ( P =0.001) in women only. There was no association between SBP or DBP and fibrinogen, factor VII, or von Willebrand factor in either sex. Conclusions —These data suggest that impaired fibrinolysis may play an important role in the pathogenesis of cardiovascular disease in hypertensive patients.

Abstract —The platelet glycoprotein IIb/IIIa (GP IIb/IIIa) plays a pivotal role in platelet aggregation. Recent data suggest that the Pl A2 polymorphism of GPIIIa may be associated with an increased risk for cardiovascular disease. However, it is unknown if there is any association between this polymorphism and platelet reactivity. We determined GP IIIa genotype and platelet reactivity phenotype data in 1422 subjects from the Framingham Offspring Study. Genotyping was performed using PCR-based restriction fragment length polymorphism analysis. Platelet aggregability was evaluated by the Born method. The threshold concentrations of epinephrine and ADP were determined. Allele frequencies of Pl A1 and Pl A2 were 0.84 and 0.16, respectively. The presence of 1 or 2 Pl A2 alleles was associated with increased platelet aggregability as indicated by incrementally lower threshold concentrations for epinephrine and ADP. For epinephrine, the mean concentrations were 0.9 μmol/L (0.9 to 1.0) for homozygous Pl A1 , 0.7 mmol/L (0.7 to 0.9) for the heterozygous Pl A1 / Pl A2 , and 0.6 μmol/L (0.4 to 1.0) for homozygous Pl A2 individuals, P =0.009. The increase in aggregability induced by epinephrine remained highly significant ( P =0.007) after adjustment for covariates. For ADP-induced aggregation, the respective mean concentrations were 3.1 μmol/L (3.0 to 3.2), 3.0 μmol/L (2.9 to 3.2), and 2.8 μmol/L (2.4 to 3.3); P =0.19 after adjustment for covariates. Our findings indicate that molecular variants of the gene encoding GP IIIa play a role in platelet reactivity in vitro. Our observations are compatible with and provide an explanation for the reported association of the Pl A2 allotype with increased risk for cardiovascular disease.

Background Given the importance of thrombosis in causation of acute coronary syndromes, it is possible that the beneficial effect of low lipid levels on the risk of coronary events is achieved by lowering thrombotic potential of the blood. Hypobetalipoproteinemia is characterized by plasma concentrations of apolipoprotein B and LDL cholesterol that are one third of those observed in the general population. The aim of this study was to utilize subjects with hypobetalipoproteinemia to examine the relation between thrombotic potential and low levels of LDL cholesterol. Methods and Results Hemostatic risk factors were measured in 1878 individuals (1003 women and 875 men) participating in cycle 5 of the Framingham Offspring Study. The subjects were divided into five groups on the basis of LDL cholesterol level. Subjects with hypobetalipoproteinemia (LDL cholesterol Conclusions Decreasing levels of LDL cholesterol are associated with decreasing levels of hemostatic risk factors. Subjects with hypobetalipoproteinemia have the lowest levels of hemostatic risk factors and may be protected against thrombotic complications of atherosclerotic cardiovascular disease because of reduced thrombotic potential. One mechanism by which lipid-lowering therapy may decrease clinical cardiac events is through a reduction in thrombotic tendency.

Limited data are available on the determinants of homocysteinemia or the association between plasma homocysteine (Hcy) levels and prevalent cardiovascular disease (CVD) in maintenance dialysis patients. We assessed etiology of renal failure, residual renal function and dialysis adequacy-related variables, and vitamin status, as determinants of fasting total plasma homocysteine (Hcy) in 75 maintenance dialysis patients. We also assessed the potential interactive effect on plasma Hcy of folate status and a common mutation (ala to val; homozygous val-val frequency approximately 10%) in methylenetetrahydrofolate reductase (MTHFR), a folate-dependent enzyme crucial for the remethylation of homocysteine (Hcy) to methionine. Lastly, we evaluated whether the Hcy levels differed amongst these patients in the presence or absence of prevalent CVD, after adjustment for the traditional CVD risk factors. Fasting total plasma Hcy, folate, pyridoxal 5'-phosphate (PLP; active B6), B12, creatinine, glucose, total and HDL cholesterol levels, and presence of the ala to val MTHFR mutation were determined, and clinical CVD and CVD risk factor prevalence were ascertained. General linear modelling/analysis of covariance revealed: (1) folate status and serum creatinine were the only significant independent predictors of fasting Hcy; (2) there was a significant interaction between presence of the val mutation and folate status, i.e., among patients with plasma folate below the median (< 29.2 ng/ml), geometric mean Hcy levels were 33% greater (29.0 vs. 21.8 microM, P = 0.012) in the pooled homozygotes (val-val) and heterozygotes (ala-val) for the ala to val mutation, vs. normals (ala-ala); (3) there was no association between prevalent CVD and plasma Hcy. Given potentially intractable survivorship effects, prospective cohort studies will be required to clarify the relationship between plasma Hcy or any putative CVD risk factor, and incident CVD in dialysis patients. If a positive association between plasma Hcy and incident CVD can be established in maintenance dialysis patients, the current data provide a rationale for additional folic acid supplementation in this patient population.

Background The CD40 receptor and its ligand (CD40L) are known to modulate both inflammation and thrombosis—2 processes important for the development and clinical expression of atherosclerosis. Circulating soluble CD40L (sCD40L) concentration predicts cardiovascular risk in selected patient samples. The purpose of this study was to determine the predictors of sCD40L in a large, community-based sample. Methods We determined both serum and plasma sCD40L concentration in 3,259 participants (54% women) from the Framingham Offspring Study. Results In multivariable models, advancing age was the only consistent (inverse) correlate of both serum and plasma sCD40L concentration. Overall, the variability explained by clinical covariates was very low for both measurements of sCD40L; with values of only 1.4% and 2.7% for serum and plasma, respectively. We observed that genetic factors accounted for a modest (12% serum; 14% plasma) amount of the adjusted variability in sCD40L. Conclusions Circulating sCD40L concentration was poorly associated with known cardiovascular disease (CVD) risk factors and was modestly heritable. Determining if either serum or plasma sCD40L are predictive of CVD risk in the community will require longitudinal follow-up.

Background : Chronic subclinical inflammation is a prominent feature of atherosclerotic disease. The genetic background for this pro-inflammatory state is not well-established. Circulating biomarker concentrations have become attractive candidates to measure disease activity and prognosis. Methods : We examined 2356 single-nucleotide polymorphisms (SNPs) in 235 inflammatory pathway genes in association with 11 circulating inflammatory biomarkers in about 1800 Framingham Offspring cohort participants [CD40 ligand, CRP, intercellular adhesion molecule-1, interleukin-6 (IL6), urinary isoprostanes, monocyte chemoattractant protein-1 (MCP1), myeloperoxidase, P-selectin, tumor necrosis factor alpha, tumor necrosis factor receptor-2, fibrinogen]. We created residuals of log transformed biomarker concentrations adjusting for 16 potential confounders. Only SNPs with call rate ≥0.98 and HWE p>0.01, which had at least 5 minor allele carriers entered analyses. False discovery rate (FDR) and q-value methods were applied. Results : We observed similar results with FDR and q-value methods. A total of 45 associations were significant at a cutoff q value of 0.25. The top SNPs were observed in the SELP gene in association with P-selectin concentrations (rs6136 [nonsynonymous], p= 5.17*10 −39 , rs3753305 [intronic], p= 6.17*10 −9 ) and the ICAM1 gene in relation to ICAM-1 concentrations (rs1799969 [coding-nonsynonymous], p= 1.32*10 −8 ). Lowest p-values for trans-acting SNPs were observed for APCS (rs1374486 [function unknown], p= 1.01*10 −7 , and rs6695377 [function unknown], p= 1.85*10 −7 ) with MCP-1 concentrations and for IL6R (rs8192284 [coding-nonsynonimous,intronic], p= 3.36*10 −5 ) with IL6 concentrations. In addition, we could replicate reported findings for rs1799969, and rs5498 in the ICAM-1 gene in relation to ICAM-1 concentrations as well as associations of SNPs rs2857654, rs1024611, and rs2857657 in the CCL-2 gene with MCP-1 concentrations. Conclusions : The results of this candidate gene approach support the relevance of genetic variation for circulating inflammatory biomarker traits. Some former findings were confirmed and novel potential candidates are reported. Our findings merit replication in other cohorts.

We sought to investigate the hypothesis that smoking is accompanied by systemic inflammation.We examined the relation of smoking to 11 systemic inflammatory markers in Framingham Study participants (n=2944, mean age 60 years, 55% women, 12% ethnic minorities) examined from 1998-2001. The cohort was divided into never (n=1149), former (n=1424), and current smokers with last cigarette6h (n=134) oror =6h (n=237) prior to phlebotomy. In multivariable-adjusted models there were significant overall between-smoking group differences (defined as p0.0045 to account for multiple testing) for every inflammatory marker tested, except for serum CD40 ligand (CD40L), myeloperoxidase (MPO) and tumor necrosis factor receptor-2 (TNFR2). With multivariable-adjustment, pair-wise comparisons with never smokers revealed that former smokers had significantly lower concentrations of plasma CD40L (p0.0001) and higher concentrations of (CRP) C-reactive protein (p=0.002).As opposed to never smokers, those with acute cigarette smoke exposure (or =6h) had significantly higher concentrations of all markers (p0.0001) except serum CD40L, MPO, and TNFR2; plasma CD40L were significantly lower. Compared with never smokers, cigarette smokers have significantly elevated concentrations of most circulating inflammatory markers, consistent with the hypothesis that smoking is associated with a systemic inflammatory state.

Background: Alcohol intake has been associated with lower platelet activity; however, few large-scale studies have included women, and to our knowledge, the relationship of alcohol intake with measures of platelet activation has not been studied. Methods: We performed a cross-sectional analysis of adults free of cardiovascular disease enrolled in the Framingham Offspring Study. Study physicians assessed alcohol consumption with a standardized questionnaire. We measured platelet activation in response to 1 and 5 μm of adenosine diphosphate (ADP) with a P-selectin assay among 1037 participants and platelet aggregability in response to ADP, epinephrine, and collagen among 2013 participants. Results: Alcohol consumption was inversely associated with P-selectin expression in response to 1 μm ADP (p= 0.007) and 5 μm ADP (p= 0.02) among men but not women. Alcohol consumption was also inversely associated with platelet aggregation induced by ADP among both women (p= 0.04) and men (p trend = 0.008) and by epinephrine among men (p= 0.03) Conclusions: Alcohol consumption is inversely associated with both platelet activation and aggregation, particularly in men. Additional research is needed to determine whether these findings contribute to the contrasting associations of alcohol consumption with risk of thrombotic and hemorrhagic cardiovascular events.

A greater life expectancy has led to an increasing proportion of elderly patients. Increasing age is an important risk factor for cardiovascular disease, but the mechanism of risk is not well understood. Because thrombosis plays a key role in plaque development and the onset of acute coronary syndromes, the age-related increase in cardiovascular risk may be a result of a prothrombotic imbalance. The study aim was to examine the relation between age and thrombotic potential in the Framingham Offspring Cohort. Hemostatic factors previously associated with cardiovascular risk were measured in 3,230 patients (55% women) without evidence of cardiovascular disease who were participating in cycle 5 of the Framingham Offspring Study. The subjects were divided by age into decades. Advancing age was associated with a significant increase in fibrinogen and von Willebrand factor levels and measures of impaired fibrinolytic potential (plasminogen activator inhibitor and tissue plasminogen activator antigens). For men, the mean fibrinogen levels were 21% higher in thoseor =70 years versus those aged40 years (326 vs 268 mg/dl, p0.001 for linear trend). The mean fibrinogen levels were 15% higher in older than in younger women (330 vs 286 mg/dl, p0.001). The significant relations persisted after multivariate adjustment. In conclusion, advancing age is associated with elevated levels of hemostatic factors indicative of a prothrombotic state. Because these factors are also associated with endothelial dysfunction, these findings are consistent with an injurious effect of age on the endothelium. Measures to reduce thrombotic potential may be of particular value in the elderly, because they counter the prothrombotic state that develops with aging.

SummaryAlthough obesity is associated with increased cardiovascular risk, the mechanism has not been fully explained. Since thrombosis is a critical component of cardiovascular disease, we examined the relationship between obesity and hemostatic factors. We studied 3230 subjects (55% females, mean age 54 years) without a history of cardiovascular disease in cycle 5 of the Framingham Offspring Study. Obesity was assessed by body mass index and waist-to-hip ratio. Fasting blood samples were obtained for fibrinogen, plasminogen activator inhibitor (PAI-1) antigen, tissue plasminogen activator (tPA) antigen, factor VII antigen, von Willebrand factor (VWF), and plasma viscosity. Body mass index was directly associated with fibrinogen, factor VII, PAI-1 and tPA antigen in both men and women (p

OBJECTIVES We sought to determine the clinical factors and heritability associated with inflammation measured as circulating levels of soluble-intercellular adhesion molecule-1 (sICAM-1) in a community-based cohort. BACKGROUND Several prospective studies indicate that circulating sICAM-1 is predictive of future cardiovascular events. However, in some studies this predictive value is lost after multivariable adjustment for traditional cardiovascular disease (CVD) risk factors. We addressed the heritability of sICAM-1 and its relation to CVD risk factors in a community-based cohort. METHODS We examined 3,295 subjects from the Framingham Heart Study and measured sICAM-1 levels. We then used linear and stepwise multivariable regression to determine predictors or sICAM-1 levels. RESULTS In age- and gender-adjusted regression models, increased sICAM-1 levels were positively associated with age, total/high-density lipoprotein cholesterol, systolic blood pressure, body mass index (BMI), blood glucose, diabetes, smoking, and prevalent CVD. In stepwise multivariable regression models, sICAM-1 levels remained associated with age, female gender, total/high-density lipoprotein cholesterol ratio, BMI, blood glucose, smoking, and prevalent CVD. The residual heritability of sICAM-1 was 24%. CONCLUSIONS In addition to prevalent CVD, established CVD risk factors and non-traditional ones such as BMI were associated with systemic inflammation as determined by sICAM-1 levels. There also is significant heritability of sICAM-1, which suggests a genetic component to systemic inflammation.

Background — Moderate alcohol consumers have lower rates of cardiovascular disease than abstainers. One proposed mechanism is a beneficial effect on hemostatic parameters, but previous studies have provided conflicting results. Methods and Results — We measured levels of fibrinogen, plasma viscosity, von Willebrand factor, factor VII, plasminogen activator inhibitor antigen-1, and tissue plasminogen activator antigen in a cross-sectional analysis of 3223 adults free of cardiovascular disease enrolled in the Framingham Offspring Study. We assessed their alcohol consumption with a standardized questionnaire. Light-to-moderate alcohol consumption was associated with lower levels of fibrinogen, plasma viscosity, von Willebrand factor, and factor VII. This association was most pronounced for consumers of 3 to 7 drinks weekly for viscosity and 7 to 21 drinks weekly for the other hemostatic measures. Alcohol intake of 7 to 21 drinks weekly or more was associated with impaired fibrinolytic potential, reflected by higher levels of plasminogen activator inhibitor antigen-1 and tissue plasminogen activator antigen. Wine drinkers had lower plasminogen activator inhibitor antigen-1 levels than other drinkers, particularly at 3 to 21 drinks weekly, but beverage type did not otherwise consistently affect the results. Conclusions — Light-to-moderate alcohol consumption is associated with lower levels of coagulatory factors, but higher intake is associated with impaired fibrinolytic potential. These findings are consistent with the hypothesis that a balance between hemostatic and fibrinolytic activity may contribute to the complex relation of alcohol use with coronary heart disease.

Background —Platelet aggregation plays an important role in arterial thrombosis in coronary heart disease, stroke, and peripheral arterial disease. However, the contribution of genetic versus environmental influences on interindividual variation in platelet aggregability is poorly characterized. Methods and Results —We studied the heritability of platelet aggregation responses in 2413 participants in the Framingham Heart Study. The threshold concentrations of epinephrine and ADP required to produce biphasic platelet aggregation and collagen lag time were determined. Mixed-model linear regression was used to calculate correlation coefficients within sibships and within spouse pairs. Variance and covariance component methods were used to estimate the proportion of platelet aggregation attributable to measured covariates versus additive genetic effects. After accounting for environmental covariates, the adjusted sibling correlations for epinephrine, ADP, and collagen lag time were 0.24, 0.22, and 0.31, respectively ( P =0.0001 for each). In contrast, adjusted correlations for spouse-pairs were −0.01, 0.05, and −0.02, respectively (all P >0.30). The estimated heritabilities were 0.48, 0.44, and 0.62, respectively. Measured covariates accounted for only 4% to 7% of the overall variance in platelet aggregation, and heritable factors accounted for 20% to 30%. The platelet glycoprotein IIIa Pl A2 polymorphism and the fibrinogen Hind III β-148 polymorphism contributed Conclusions —In our large, population-based sample, heritable factors play a major role in determining platelet aggregation, and measured covariates play a lesser role. Future studies are warranted to identify the key genetic variants that regulate platelet function and to lay the groundwork for rational pharmacogenetic approaches.

Background —Fibrinogen has been identified as an independent risk factor for cardiovascular disease and associated with traditional cardiovascular risk factors. Also, the role of elevated fibrinogen in thrombosis suggests that it may be on the causal pathway for certain risk factors to exert their effect. These associations remain incompletely characterized. Moreover, the optimal fibrinogen assay for risk stratification is uncertain. Methods and Results —In 2632 subjects from cycle 5 of the Framingham Offspring Population, fibrinogen levels were determined with a newly developed immunoprecipitation test (American Biogenetic Sciences) and the functional Clauss method. With the immunoprecipitation method, there were significant linear trends across fibrinogen tertiles ( P P P =0.035 and P =0.018 for men and women, respectively) but not with the Clauss method. Conclusions —Fibrinogen was associated with traditional cardiovascular risk factors. Elevation of fibrinogen may provide a mechanism for risk factors to exert their effect. Also, fibrinogen levels were higher among subjects with cardiovascular disease compared with those without disease. The immunoprecipitation test showed a stronger association with cardiovascular disease than the Clauss method, suggesting that it may be a useful screening tool to identify individuals at increased thrombotic risk.

Abstract —Elevated factor VII levels have been associated with increased cardiovascular risk in some studies. The arginine/glutamine (Arg/Gln) polymorphism of the factor VII gene has been previously shown to modify factor VII levels. However, the presence of a gene/environment interaction on factor VII levels or a link with cardiovascular disease (CVD) remains uncertain. We studied subjects from the Framingham Heart Study to determine (1) the extent to which this genetic polymorphism affects factor VII levels; (2) whether interactions exist between this polymorphism and environmental factors on factor VII levels; and (3) the association between the polymorphism and CVD. Genotype data and factor VII antigen levels were available in 1816 subjects. Factor VII levels differed significantly among genotypes in an additive fashion: Gln homozygous, 82.7±2.5%; heterozygous, 92.2±0.7%; and Arg homozygous, 100.5±0.4% ( P P =0.02) between the genotype and total cholesterol on factor VII levels, such that the correlation coefficient and slope (factor VII level/total cholesterol) were greatest in Gln/Gln subjects. Among 3204 subjects characterized for genotype and CVD, there was no significant relationship between the genotype and CVD ( P =0.12). In the Framingham Heart Study, the Arg/Gln polymorphism was significantly associated with factor VII antigen levels. The strength of the association suggests that genetic variation plays an important role in determining factor VII levels. However, despite being associated with factor VII levels, the Arg/Gln polymorphism was not associated with prevalent CVD.

Recent investigations have highlighted the importance of the matricellular protein osteopontin as a key mediator in the cardiovascular system, specifically in the processes of vascular remodelling, vascular and valvular calcification and left ventricular (LV) remodelling.1 To our knowledge, no prior study has investigated whether plasma osteopontin concentrations are related to cardiovascular disease risk factors, valve calcification, and LV dilatation and hypertrophy in a community-based sample. Accordingly, we investigated the clinical and echocardiographic correlates of plasma osteopontin in a community-based sample. The design and selection criteria of the Framingham Offspring study have been described previously.2 On the basis of the sex-specific distributions of echocardiographic LV measurements, we sampled participants from the sixth examination cycle (1995–8) with both LV end diastolic dimension (LVEDD) and wall thickness (LVWT) below the sex-specific median (referent, n = 129), with increased LVEDD (⩾ 90th sex-specific centile, n = 134) and increased LVWT (⩾ 90th sex-specific centile, n = 128) in a 1:1:1 ratio. Eighteen participants were included in both LV dilatation and increased LVWT groups, so the study sample consisted of 373 participants. Participants underwent a standardised medical history and physical examination. Fasting blood samples were drawn and frozen at −70°C without any freeze–thaw cycles until assay. Plasma osteopontin was measured in duplicate by using an enzyme-linked immunosorbent assay (Calbiochem, Inc) with an intra-assay coefficient of variation of 4.6%. …

Maintenance dialysis patients experience an exceedingly high incidence of arteriosclerotic cardiovascular disease (CVD) events that are poorly predicted by traditional CVD risk factor indices. We evaluated the prevalence of three non-traditional CVD risk factors, i.e. hyperhomocysteinemia, hyperfibrinogenemia, and lipoprotein (a) (Lp(a)) excess, and combined hyperhomocysteinemia, hyperfibrinogenemia, and Lp(a) excess, in maintenance dialysis patients. Fasting total plasma homocysteine (Hcy), fibrinogen, Lp(a), glucose, and total and HDL cholesterol levels, and traditional CVD risk factor (i.e. glucose tolerance, smoking, hypertension, dyslipidemia) prevalences were assessed in 71 dialysis patients and 71 age, sex, and race matched Framingham Study controls free of clinical renal disease, with normal serum creatinine (≤ 1.5 mg/dl). Mean plasma Hcy (23.7 vs. 9.9 μM, P = 0.0001), fibrinogen (457 vs. 309 mg/dl, P = 0.0001), and Lp(a) (30 vs. 17 mg/dl, P = 0.0070) levels were substantially increased in the dialysis patients. Matched odds ratios (with 95% confidence intervals), dialysis patients/controls, for hyperhomocysteinemia, hyperfibrinogenemia, and Lp(a) excess, alone or combined, were markedly greater in the dialysis patients, with no evidence of confounding by the traditional CVD risk factors: hyperhomocysteinemia, 105.0 (29.9–368.9); hyperfibrinogenemia, 16.6 (6.6–42.0); Lp(a) excess, 3.5 (1.5–8.4); all three combined 35.0 (5.7–199.8). Given in vitro evidence that Hcy, Lp(a), and fibrinogen interact to promote atherothrombosis, combined hyperhomocysteinemia, hyperfibrinogenemia, and Lp(a) excess may contribute to the high incidence of vascular disease sequelae experienced by dialysis patients, which is inadequately explained by traditional CVD risk factors. Controlled, prospective studies of well-characterized maintenance dialysis cohorts are urgently required to substantiate this hypothesis.

Hyperhomocysteinemia occurs frequently in end-stage renal disease (ESRD), but its prevalence in comparison with traditional cardiovascular disease (CVD) risk factors is unknown. Fasting total plasma homocysteine, potential determinants of plasma homocysteine (i.e., plasma B-vitamins and serine), total and HDL cholesterol, glucose, and creatinine, were determined in 24 ESRD patients on dialysis, and 24 age, gender, and race matched Framingham Offspring Study controls with normal renal function. Presence of clinical CVD and CVD risk factors was established by standardized methods. Mean plasma homocysteine was markedly higher in the ESRD patients versus controls (22.7 vs. 9.5 mumol/l). ESRD patients were 33 times more likely than controls to have hyperhomocysteinemia (> 15.8 mumol/l) (95% confidence interval, 5.7-189.6). Hyperhomocysteinemia persisted in the ESRD patients despite normal to supernormal B-vitamin status. Plasma serine levels below the tenth percentile of the control distribution were found in 75% of the ESRD patients. Oral serine supplementation caused a 37% increase in mean plasma serine, but had no effect on plasma homocysteine in four ESRD patients with supernormal plasma folate, low plasma serine, and hyperhomocysteinemia. Given its unusually high prevalence, improved management of hyperhomocysteinemia might reduce CVD sequelae in ESRD.

Background Although extensive evidence indicates that estrogen is responsible for the markedly decreased cardiovascular risk of premenopausal women, the mechanism through which estrogen might exert its protective effect has not been adequately explained. Since thrombosis is now recognized to play an important role in the onset of cardiovascular disease, we investigated the relation between estrogen status and fibrinolytic potential, a determinant of thrombotic risk. Methods and Results We determined levels of plasminogen activator inhibitor (PAI-1) antigen and tissue plasminogen activator (TPA) antigen in 1431 subjects from the Framingham Offspring Study. Fibrinolytic potential was compared between subjects with high estrogen status (premenopausal women and postmenopausal women receiving hormone replacement therapy) and low estrogen status (men and postmenopausal women not receiving hormone replacement therapy). In all comparisons, subjects with high estrogen status had greater fibrinolytic potential (lower PAI-1 levels) than subjects with low estrogen status. First, postmenopausal women receiving estrogen replacement therapy had lower levels of PAI-1 than those not receiving therapy (13.0±0.5 versus 19.5±1.0 ng/mL, P P P =.089). Third, premenopausal women had markedly lower levels of PAI-1 antigen than postmenopausal women not receiving estrogen therapy (14.8±0.6 versus 19.5±1.0 ng/mL, P Conclusions Each of these comparisons indicates that the cardioprotective effect of estrogen may be mediated, in part, by an increase in fibrinolytic potential. These findings might provide at least a partial explanation for the protection against cardiovascular disease experienced by premenopausal women, and the loss of that protection following menopause.

The recent findings that lipid lowering therapy reduces clinical events to a greater degree than it reduces stenosis severity suggest beneficial effects other than plaque regression. To determine if a favorable lipid profile is associated with decreased thrombotic tendency, we determined the relationship between two hemostatic risk factors, fibrinogen and plasminogen activator inhibitor (PAI-l), and LDL cholesterol (LDL-C) in 1,878 subjects free of cardiovascular disease in the Framingham Offspring Study. The subjects studied ranged from those with hypobetalipoproteinemia, a condition characterized by LDL-C ≤ 70 mg/dl, and a low incidence of thrombosis and coronary heart disease, to other LDL-C levels grouped by NCEP guidelines. LDL-C (mg/dl) l 70 70–99 100 129 g 130 * p-value Number subjects 41 389 653 795 Fibrinogen (mg/dl) 264 ± 54 287 ± 59 297 ± 57 303 ± 55 l 0.0001 PAI-l antigen (ng/ml) 14 ± 9 27 ± 18 22 ± 17 23 ± 46 l 0.0001 * p-value for linear effect of LDL-C These findings were similar in men and women. Multivariate adjustment for age, gender, body mass index, diabetes, smoking, alcohol intake, and use of antihypertensive medication did not alter the results. Conclusion Decreasing LDL-C levels are associated with decreasing levels of hemostatic risk factors. These findings support the hypothesis that lipid lowering therapy may reduce clinical events by reducing thrombotic tendency.

Hormone replacement therapy (HRT) is associated with a decreased risk of coronary heart disease (CHD) in postmenopausal women, but the mechanism of its protective effect is not fully characterized. To evaluate the relation ship between postmenopausal HRT and fibrinogen, an established hemostatic risk factor for CHD, we measured fibrinogen using the Clauss method in 858 women free of CHD in cycle 4 of the Framingham Offspring Study. The table shows mean ± SD fibrinogen levels for pre and postmeonopausal women according to use of HRT N Fibrinogen (mg/dl) %Δ p-value Premenopause 303 289 ± 53 Postmenopause (No HRT) 383 313 ± 58 ↑7.7% * l 0.001 * Postmenopause (HRT) 86 284 ± 44 ↓9.3% ** l 0.001 ** * Premenopause vs. Postmenopause (No HRT); ** Postmenopause (No HRT) vs. Postmenopause (HRT) (Age adjusted) These differences persisted upon multivariate adjustment for smoking, use of antihypertensive medication, diabetes mellitus, body mass index, alcohol intake, total cholesterol and HDL cholesterol. Postmenopausal HRT is associated with fibrinogen levels that are comparable to those found in premenopausal women. These data suggest that the protective effect of postmenopausal HRT may be mediated in part by favorable alterations in the hemostatic system.