Digna Cabral

Traditional Risk Factors Are Not Major Contributors to the Variance in Carotid Intima-Media Thickness

Background and Purpose— Carotid intima-media thickness (cIMT) was a widely accepted ultrasound marker of subclinical atherosclerosis in the past. Although traditional risk factors may explain ≈50% of the variance in plaque burden, they may not explain such a high proportion of the variance in IMT, especially when measured in plaque-freel ocations. We aimed this study to identify individuals with cIMT unexplained by traditional risk factors for future environmental and genetic research. Methods— As part of the Northern Manhattan Study, 1790 stroke-free individuals (mean age, 69±9 years; 60% women; 61% Hispanic; 19% black; 18% white) were assessed for cIMT using B-mode carotid ultrasound. Multiple linear regression models were evaluated: (1) incorporating prespecified traditional risk factors; and (2) including less traditional factors, such as inflammation biomarkers, adiponectin, homocysteine, and kidney function. Standardized cIMT residual scores were constructed to select individuals with unexplained cIMT. Results— Mean total cIMT was 0.92±0.09 mm. The traditional model explained 11% of the variance in cIMT. Age (7%), male sex (3%), glucose (<1%), pack-years of smoking (<1%), and low-density lipoprotein cholesterol (<1%) were significant contributing factors. The model, including inflammatory biomarkers, explained 16% of the variance in cIMT. Adiponectin was the only additional significant contributor to the variance in cIMT. We identified 358 individuals (20%) with cIMT unexplained by the investigated risk factors. Conclusions— Vascular risk factors explain only a small proportion of variance in cIMT. Identification of novel genetic and environmental factors underlying unexplained subclinical atherosclerosis is of utmost importance for future effective prevention of vascular disease.

Traditional cardiovascular risk factors explain the minority of the variability in carotid plaque

Background and Purpose— Subclinical atherosclerotic plaque is an important marker of increased vascular risk. Identifying factors underlying the variability in burden of atherosclerotic carotid plaque unexplained by traditional vascular risk factors may help target novel preventive strategies. Methods— As a part of the carotid substudy of the Northern Manhattan Study (NOMAS), 1790 stroke-free individuals (mean age, 69±9; 60% women; 61% Hispanic, 19% black, 18% white) were assessed for total plaque area (TPA) burden using 2-dimensional carotid ultrasound imaging. Multiple linear regression models were constructed. Model 1 used prespecified traditional risk factors: age, sex, low-density lipoprotein cholesterol, diabetes mellitus, pack-years of smoking, blood pressure, and treatment for blood pressure; and Model 2, an addition of socioeconomic and less traditional risk factors. The contributions of the components of the Framingham heart risk score and the NOMAS Global Vascular Risk Score to the TPA were explored. Results— Prevalence of carotid plaque was 58%. Mean TPA was 13±19 mm2. Model 1 explained 19.5% of the variance in TPA burden (R2=0.195). Model 2 explained 21.9% of TPA burden. Similarly, the Framingham heart risk score explained 18.8% and NOMAS global vascular risk score 21.5% of the TPA variance. Conclusions— The variation in preclinical carotid plaque burden is largely unexplained by traditional and less traditional vascular risk factors, suggesting that other unaccounted environmental and genetic factors play an important role in the determination of atherosclerotic plaque. Identification of these factors may lead to new approaches to prevent stroke and cardiovascular disease.

Association of the sirtuin and mitochondrial uncoupling protein genes with carotid plaque

Objective Sirtuins (SIRTs) and mitochondrial uncoupling proteins (UCPs) have been implicated in cardiovascular diseases through the control of reactive oxygen species production. This study sought to investigate the association between genetic variants in the SIRT and UCP genes and carotid plaque. Methods In a group of 1018 stroke-free subjects from the Northern Manhattan Study with high-definition carotid ultrasonography and genotyping, we investigated the associations of 85 single nucleotide polymorphisms (SNPs) in the 11 SIRT and UCP genes with the presence and number of carotid plaques, and evaluated interactions of SNPs with sex, smoking, diabetes and hypertension as well as interactions between SNPs significantly associated with carotid plaque. Results Overall, 60% of subjects had carotid plaques. After adjustment for demographic and vascular risk factors, T-carriers of the SIRT6 SNP rs107251 had an increased risk for carotid plaque (odds ratio, OR = 1.71, 95% CI = 1.23–2.37, Bonferroni-corrected p = 0.03) and for a number of plaques (rate ratio, RR = 1.31, 1.18–1.45, Bonferroni-corrected p = 1.4×10−5), whereas T-carriers of the UCP5 SNP rs5977238 had an decreased risk for carotid plaque (OR = 0.49, 95% CI = 0.32–0.74, Bonferroni-corrected p = 0.02) and plaque number (RR = 0.64, 95% CI = 0.52–0.78, Bonferroni-corrected p = 4.9×10−4). Some interactions with a nominal p≤0.01 were found between sex and SNPs in the UCP1 and UCP3 gene; between smoking, diabetes, hypertension and SNPs in UCP5 and SIRT5; and between SNPs in the UCP5 gene and the UCP1, SIRT1, SIRT3, SIRT5, and SIRT6 genes in association with plaque phenotypes. Conclusion We observed significant associations between genetic variants in the SIRT6 and UCP5 genes and atherosclerotic plaque. We also found potential effect modifications by sex, smoking and vascular risk factors of the SIRT/UCP genes in the associations with atherosclerotic plaque. Further studies are needed to validate our observations.

Carotid Plaque and Candidate Genes Related to Inflammation and Endothelial Function in Hispanics From Northern Manhattan

Background and Purpose— The genetic influence on carotid atherosclerotic plaque is mostly unknown. This study examines the association between carotid plaque and single nucleotide polymorphisms in selected genes implicated in inflammation and endothelial function. Methods— A total of 43 genes (197 single nucleotide polymorphisms) involved in inflammation and endothelial function were interrogated in 287 Dominicans from the Northern Manhattan Study (mean age, 64±7 years; 58% women) who had undergone high-resolution B-mode ultrasound for examination of carotid plaque. Using an additive genetic model, multiple logistic regression analyses were conducted, a within-gene haplotype analysis was performed, and interactions between genes were examined. Results were validated in an independent set of 301 Dominicans. Results— Carotid plaque was present in 143 (47%) participants. Nine genes had at least 1 single nucleotide polymorphism associated (P≤0.01) with carotid plaque phenotypes: TNF, NOS2A, IL6R, TNFSF4, PPARA, IL1A, TLR4, ITGA2, and HABP2. Single nucleotide polymorphisms in TNFSF4, PPARA, TLR4, ITGA2, and HABP2 were also implicated with the same carotid phenotype in the validation analysis. Haplotype analysis revealed an additional gene of interest, VCAM1. Conclusions— We report novel associations between variations in 10 genes involved in inflammation and endothelial function and carotid plaque phenotypes in a Dominican sample, with replication for 5 genes in an independent Dominican sample.

Ethnic Differences in Carotid Artery Diameter and Stiffness: The Northern Manhattan Study

OBJECTIVE Race/ethnic differences in carotid arterial function and structure exist among those with cerebrovascular disease, but whether differences persist among healthy populations is unknown. Our objective was to investigate differences in carotid artery diameter and stiffness between race/ethnic groups, and examine whether these race/ethnic differences were age-dependent. METHODS Carotid diameters were assessed by B-mode ultrasound among 1536 participants from the Northern Manhattan Study (NOMAS), and carotid stiffness metrics were calculated. We used multivariable linear regression models to determine the relationship between race/ethnicity and both carotid arterial stiffness and carotid diastolic diameter. RESULTS Mean participant age was 70 ± 9 years (Hispanics = 68 ± 8, blacks = 72 ± 9, and whites = 74 ± 9, p < 0.0001). Mean DDIAM was 6.2 ± 1.0mm (Hispanics = 6.2 ± 0.9 mm, blacks = 6.3 ± 1.0 mm, and whites = 6.3 ± 1.0 mm, p < 0.005) and mean STIFF was 8.7 ± 6.3 (Hispanics = 8.5 ± 5.7, blacks = 9.2 ± 6.2 and whites = 8.9 ± 6.9, p < 0.02). In a model that adjusted for sociodemographics and vascular risk factors including hypertension, diabetes, dislipidemia, renal function, physical acticity and a history of known coronary artery diseases; age was positively associated with greater DDIAM in Hispanics (p < 0.0001) but not among blacks or whites. Older age was associated with greater stiffness among Hispanics (p < 0.0001) and blacks (p < 0.003), but not among whites. CONCLUSIONS We found race/ethnic differences in the association between age and arterial stiffness and diameter, including age-dependent arterial dilation observed in Hispanics that was not observed among blacks or whites.

Mediterranean diet and carotid atherosclerosis in the Northern Manhattan Study

OBJECTIVE Adherence to a Mediterranean-style diet (MeDi) may protect against clinical vascular events by reducing atherosclerosis, but data is limited. This is the first observational study of the association between MeDi adherence and carotid plaque thickness and area. METHODS The study included 1374 participants of the population-based Northern Manhattan Study with diet assessed and carotid intima-media thickness (cIMT) and plaque measured using B-mode ultrasound (mean age 66 ± 9 years, 60% female, 60% Hispanic, 18% White, 19% Black). A MeDi adherence score (range = 0-9, 9 representing maximal adherence) was examined continuously and in quintiles (3/4/5/6-9 vs. 0-2). RESULTS Mean cIMT = 0.9 ± 0.1 mm and 57% had plaque (median plaque thickness = 1.5 mm, 75th percentile = 2.2; median plaque area = 4.2 mm(2), 75th percentile = 15.8). There was no association between MeDi and cIMT or plaque presence. MeDi adherence was inversely associated with the 75th percentile of plaque thickness and median of plaque area in quantile regression analyses. These associations persisted after controlling for demographics, smoking, physical activity, and total energy consumption (effect of a 1-point increase in MeDi score on the 75th percentile of plaque thickness = -0.049 mm, p = 0.03; median of plaque area = -0.371 mm(2), p = 0.03), and when additionally controlling for vascular disease biomarkers, medication use, BMI, and previous cardiac disease. The protective associations appeared strongest for those with a MeDi score of 5 (4th quintile) vs. 0-2 (bottom quintile). Differential effects of a MeDi on plaque thickness and area across race/ethnic groups was suggested. CONCLUSIONS Moderate and strict adherence to a MeDi may protect against a higher burden of carotid atherosclerotic plaque, which may mediate the protection against clinical vascular events. Efforts to improve adherence to a MeDi are critical to reducing the burden of atherosclerotic disease.

Association of the sirtuin and mitochondrial uncoupling protein genes with carotid intima-media thickness.

Sirtuins (SIRTs) are a family of nicotinamide adenine dinucleotide (NAD+)–dependent deacetylases involved in chromatin remodelling, cellular metabolism, and lifespan regulation.1 Uncoupling proteins (UCPs) are a family of inner mitochondrial membrane proteins capable of driving the adenosine triphosphate synthase pathway via regulation of the proton electrochemical gradient.2 SIRTs and UCPs have been implicated in atherosclerosis and cardiovascular disease (CVD) through the control of reactive oxygen species production.1,2 In our previous in vivo study, the up- or downregulation and the enzymatic activity of SIRT/UCP proteins have been related to the degree of tolerance to brain ischemia.3 Therefore, a different alteration in functionality of these proteins may have a different impact on CVD and subclinical atherosclerosis. We have recently demonstrated a link between variants in the SIRTs and UCPs genes and risk of carotid plaque (CP) and reported the effect-modifications of these relationships by sex, smoking, diabetes, and hypertension in the Northern Manhattan Study (NOMAS).4 Genetic variants in the SIRTs and UCPs genes may have an important role in development of atherosclerosis, independent of, or synergistic with, common vascular risk factors. Given that CP and carotid intima-media thickness (cIMT) are mainly biologically and genetically distinct phenotypes of atherosclerosis,5 in the present study, we extend our investigations to the associations of genetic variations in the SIRTs and UCP genes with cIMT. We analyzed 1018 participants with cIMT and genotype data from NOMAS.6 All participants provided written informed consents. The study was approved by the Institutional Review Boards of Columbia University and the University of Miami. cIMT was assessed by B-mode ultrasound according to standardized scanning and reading protocols.7 A total of 85 single-nucleotide polymorphism (SNPs) in the 11 UCP and SIRT genes were available from the AffyMetrixGenome-Wide Human SNP array 6.0. Detailed genotyping procedures were described previously.4 Generalized linear modelling was performed to evaluate the association of each SNP with cIMT after adjustment for age, sex, eversmoking, and the top 2 principle components by EIGENSTRAT to adjust for population stratification.8 The effect modifications by sex and ever smoking were explored by including interaction terms to the regression models. Stratified analyses by sex and smoking were also performed. The mean age of the participants was 70 ± 9 years, 61% were women, 67% Caribbean Hispanic, 17% Black, and 15% White. The mean cIMT was 0.96 ± 0.10 mm for men and 0.93 ± 0.09 mm for women. The prevalence of hypertension was 62%, diabetes 18%, hypercholesterolemia 27%, and ever smoking 52%. In univariate analysis, age, sex, and smoking were significantly associated with cIMT. Three SNPs showed an association with cIMT with a nominal P <0.05, after adjustment for significant covariates and the top 2 principal components of ancestry (Table I): elevated cIMT for TT-carriers at rs1430583 (β = 0.036, P = 0.006) or CC-carriers at rs6818140 (β = 0.036, P = 0.007) in UCP1, and lower cIMT for CC-carriers allele at rs12363280 (β =−0.041, P =0.046) in SIRT3. Significant effect-modification was observed by sex and smoking (Table II). Men homozygous for the minor alleles at rs6818140 (C) in UCP1 had increased cIMT, whereas women homozygous for the T allele (minor allele) at rs3825075 in SIRT3 had lower cIMT. Significantly elevated cIMT was found in smokers homozygous for any of the minor alleles at rs6818140 (C) in UCP1, rs7109266 (A) in UCP2, or rs4802998 (G) in SIRT2. Table I SNPs associated with cIMT Table II UCP and SIRT SNPs showing interaction with sex and smoking The present study demonstrates a sex-specific effect of genetic variants in UCP1 and SIRT3 on cIMT. Sex hormones have been shown to modulate UCP1 expression.9 Recent reports have described the SIRT3 sex differences in its expression after physical activity.10 These findings may in part explain sex-dependent associations of UCP1 and SIRT3 genetic variants with cIMT. The present study also shows an effect-modification of UCP1, UCP2, and SIRT2 with cIMT by smoking. A direct association between smoking and UCPs was found in mice where exposure to cigarette smoke increased UCPs in the brown fat.11 The regulation of the UCP1 transcription has been demonstrated in the development of atherosclerotic lesions through its effect on monocytes/macrophages, vascular endothelial cells, and vascular smooth muscle cells.12 UCP2 levels measured in peripheral blood lymphocytes were higher in smokers than that in nonsmokers, indicating a role of this protein in response to the oxidative stress induced by smoke.13 SIRT2 is a predominantly cytoplasmic protein implicated in oxidative stress, cell cycle regulation, and apoptosis,1 and its role in atherosclerosis may be exaggerated by smoking. Further studies including gene expression, proteomics, and deep sequencing are needed to clarify exact mechanisms of these interactions. Previously, using the same study population we demonstrated novel associations of genetic variants in UCP1, 3, 4, and 5 and SIRT3, 5, and 6 and CP.4 Several important effect modifications of these relationships were found by sex, smoking, hypertension, and diabetes. The lack of replication of these findings in the present study further confirms that plaque and cIMT are different subclinical phenotypes of atherosclerosis with different genetic mechanisms.5 Plaque is more strongly related to traditional risk factors and to coronary artery disease than is IMT.5 A combined approach using traditional vascular risk factors and validated genetic markers for atherosclerosis may offer a better risk-prediction of vascular diseases. In addition, it may be possible to identify alterations in pathways involved in CVD beyond the effect of traditional vascular risk factors. Strength of this study is the cohort population with adequate representation of minority ethnic groups, which are usually far underrepresented in studies of this type. Our results however need to be taken with caution because of several limitations.4 We included a relative small convenience sample from our genome-wide association studies with available SNPs in the 7 SIRT and 5 UCP genes (a total of 3055 SNPs, http://www.ncbi.nlm.nih.gov/snp) and did not adjust for multiple testing. Despite these limitations the present study reveals novel associations between the UCP and SIRT genes and cIMT with specific effect-modifications by sex and smoking. cIMT is a significant predictor of vascular events. Genetics may play an important role in the determination of atherosclerosis, detecting asymptomatic individuals at increased vascular risk and help developing new therapeutic approaches, which are specifically tailored to atherosclerotic pathways.

Cerebral Hemodynamics in the Elderly A Transcranial Doppler Study in the Einstein Aging Study Cohort

We sought to describe the relationship between age, sex, and race/ethnicity with transcranial Doppler hemodynamic characteristics from major intracerebral arterial segments in a large elderly population with varying demographics.

Sirtuin/uncoupling protein gene variants and carotid plaque area and morphology.

Background Sirtuins and uncoupling proteins have been implicated in cardiovascular diseases by controlling oxidative stress. Aims We sought to investigate the association of sirtuins and uncoupling proteins single nucleotide polymorphisms with total carotid plaque area and morphology measured by ultrasonographic gray scale median. Methods We analyzed 1356 stroke-free subjects (60% women, mean age = 68 ± 9 years) from the Northern Manhattan Study. Multiple linear regression models were used to evaluate the association of 85 single nucleotide polymorphisms in 11 sirtuins/uncoupling protein genes with total plaque area and gray scale median after controlling for demographics, vascular risk factors (RFs), and population stratification. We investigated effect modifications of these relationship by gender and RFs and performed stratified analysis if the interaction effect had P < 0·005. Results Among individuals with present plaque (55%), the mean total plaque area was 20·3 ± 20·8 mm2 and gray scale median 90 ± 29. After adjustment, SIRT6 rs107251 was significantly associated with total plaque area (β = 0·30 per copy of T allele increase, Bonferroni-corrected P = 0·005). T allele carriers of rs1430583 in UCP1 showed a decreased gray scale median in women but not in men. The minor allele carriers of rs4980329 and rs12363280 in SIRT3 had higher gray scale median in men but not in women. Variants in UCP3 gene were significantly associated with higher mean gray scale median in individuals with dyslipidemia. Conclusion Our findings suggest that polymorphisms in SIRT6/UCP1 genes may be important for increased carotid plaque burden and echodensity, but translation of these findings to an individual risk of cerebrovascular events needs further investigation. Significant associations of rs1430583 in women, rs12363280 in men, and rs1685354 in those with dyslipidemia also deserve further investigations.

Subclinical Atherosclerosis among Young and Middle-Aged Adults Using Carotid Intima-Media Thickness Measurements

Objectives The presence of atherosclerotic plaque in the carotid arteries is a strong predictor of cardiovascular disease (CVD). Research and data on CVD risk have been derived primarily from individuals aged 55 years or older, and assessment of CVD risk among young and middle-aged adults seldom has been studied. The use of ultrasonography to measure carotid intima-media thickness (IMT) and carotid plaque appears to have utility to detect subclinical atherosclerosis in asymptomatic adults. This study evaluated the presence of carotid plaque using ultrasonography among healthy young and middle-aged adults. Methods Participants were men and women recruited in Miami, Florida, and were 18 to 50 years old with no history of CVD. Participants underwent a general physical examination and carotid artery ultrasonography to evaluate carotid IMT and carotid plaque. Results From a total of 173 participants with a mean age of 34 years (standard deviation 8.9), 21.0% (95% confidence interval [CI] 15.0–27.2) were identified as having carotid plaque. IMT values ranged from 0.49 to 1.03 mm, with a mean value of 0.70 mm (standard deviation 0.09). In multivariable logistic regression older age (adjusted odds ratio [AOR] 1.08, 95% CI 1.01–1.16, P = 0.024) and cigarette smoking (AOR 2.67, 95% CI 1.02–7.00, P = 0.045) were associated with plaque, after controlling for IMT (AOR 2.55, 95% CI 1.40–4.65, P = 0.002). Conclusions Traditional CVD risk factors such as those evaluated in this study may fail to provide adequate predictive value of carotid atherosclerosis in younger populations with no history of CVD, because the majority of traditional risk factors identified in previous research were not associated with carotid plaque in this young sample. Further research assessing nontraditional risk factors among asymptomatic individuals is required, and the evaluation of IMT as an intervention tool to detect CVD risk in these asymptomatic populations is warranted.