Maturitas 81 (2015) 311–316
Contents lists available at ScienceDirect
Maturitas journal homepage: www.elsevier.com/locate/maturitas
Subclinical cardiovascular disease in postmenopausal women with low/medium cardiovascular risk by the Framingham risk score Maria Augusta Maturana a,b , Roberta Fernandez Franz a , Marcela Metzdorf a , Thais Rasia da Silva a , Poli Mara Spritzer a,c,∗ a
Gynecological Endocrinology Unit, Division of Endocrinology, Hospital de Clínicas de Porto Alegre, Porto Alegre, RS, Brazil University Foundation of Cardiology, Institute of Cardiology of Rio Grande do Sul, Porto Alegre, Brazil c Laboratory of Molecular Endocrinology, Department of Physiology, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brazil b
a r t i c l e
i n f o
Article history: Received 26 October 2014 Received in revised form 3 March 2015 Accepted 13 March 2015 Keywords: Menopause Estrogen Subclinical cardiovascular disease Carotid intima–media thickness Coronary artery calciﬁcation
a b s t r a c t Objectives: To evaluate the prevalence of subclinical cardiovascular disease (CVD) and its association with clinical and hormone variables in postmenopausal women from Southern Brazil. Study design: Cross-sectional study. Main outcome measures: Coronary artery calciﬁcation (CAC) assessed by electron-beam computed tomography. Carotid intima–media thickness (IMT) and atheromatous plaques assessed using B-mode ultrasound. IMT was measured at three segments. Subclinical CVD was deﬁned as the presence of plaque and/or IMT >0.9 mm. Results: Ninety-seven postmenopausal women (mean age 55 ± 5 years, median duration of menopause 5.8 [3–10] years) were studied. A low/medium Framingham risk score (FRS) was present in 97.9% of participants; 35.1% had subclinical CVD on carotid ultrasound, and 24.7% had the presence of plaque. Seven women had a CAC score ≥100, and two had a score ≥200. CAC score (p < 0.001) and FRS (p = 0.013) were higher in patients with subclinical atherosclerosis. Positive correlations were found between IMT and age (rs = 0.293 p = 0.004), duration of menopause (rs = 0.237, p = 0.020), and CAC score (rs = 0.468, p < 0.001). Common carotid IMT (IMT-CC) was negatively associated with estradiol levels (ˇ = −0.237, p = 0.018) and positively with age (ˇ = 0.210, p = 0.033), and BMI (ˇ = 0.260, p = 0.010). However, correlations with estradiol and age did not remain signiﬁcant when adjusted for systolic blood pressure and LDL-cholesterol levels. Conclusion: A high prevalence of subclinical atherosclerosis was detected in this sample of postmenopausal women with low/medium CV risk by the FRS. The association between IMT-CC and age or endogenous estrogen levels was dependent of blood pressure and LDL-cholesterol in these postmenopausal women from Southern Brazil. © 2015 Elsevier Ireland Ltd. All rights reserved.
1. Introduction Menopause has been associated with increased risk of atherosclerosis, and postmenopausal women experience an upward transition of cardiovascular (CV) risk, possibly in association with changing hormonal status and aging [1,2]. While clinical manifestations of atherosclerosis arise only in middle age, the process of atherosclerosis and the development of vascular changes
∗ Corresponding author at: Division of Endocrinology, Hospital de Clínicas de Porto Alegre Rua Ramiro Barcelos, 2350, 90035-003 Porto Alegre, RS, Brazil. Tel.: +55 51 3359 8027; fax: +55 51 3359 8777. E-mail address: [email protected]
(P.M. Spritzer). http://dx.doi.org/10.1016/j.maturitas.2015.03.012 0378-5122/© 2015 Elsevier Ireland Ltd. All rights reserved.
begin earlier in life, and have been associated with an increase in CV risk factors [3–5]. The decline in endogenous estrogen production during the menopausal transition has been associated with CV risk factors and higher prevalence of subclinical cardiovascular disease (CVD), such as atherosclerosis, in postmenopausal women [6–9]. Some non-invasive procedures are able to detect and measure different stages of atherosclerosis, even in its subclinical form. B-mode ultrasound is commonly used to assess brachial artery ﬂow-mediated dilation (FMD), carotid intima-media thickness (IMT), and carotid plaques, while the coronary artery calciﬁcation (CAC) score can be derived through electron-beam or multislice computed tomography (CT) scanning. Adverse FMD results, increased IMT, and the presence of carotid plaques or CAC have
M.A. Maturana et al. / Maturitas 81 (2015) 311–316
been associated with higher CV risk that is independent of other conventional risk factors [10–12]. Within this context, the aim of this study was to estimate the prevalence of subclinical CVD and its associations with endogenous estradiol levels and demographic, anthropometric, and metabolic variables in postmenopausal women. 2. Methods 2.1. Patients This cross-sectional study was carried out on a sample of women presenting with climacteric symptoms at the Gynecological Endocrinology Unit of Hospital de Clínicas de Porto Alegre, Brazil. Volunteers were also consecutively recruited through advertisements placed in a local newspaper and radio station. The inclusion criteria were as follows: (1) menopause, deﬁned as a last menstrual period at least 1 year before the beginning of the study plus follicle-stimulating hormone (FSH) levels above 35 IU/L; (2) age between 45 and 65 years; and (3) no use of hormone therapy in the past 3 months. The exclusion criteria were diabetes, current smoking, or prior diagnosis of CVD. The study was approved by the local Research Ethics Committee, and written informed consent was obtained from each participant. 2.2. Study protocol Anthropometric measurements included body weight, height, waist circumference (WC) (measured at the midpoint between the lower rib margin and the iliac crest), and body mass index (BMI, calculated as the latest measured weight in kilograms divided by the height in meters squared). Blood pressure was measured with participants in the seated position, with feet on the ﬂoor, and the arm supported at heart level after a 10-min rest. Two measurements were obtained, 10 min apart, using an automatic blood pressure monitor (HEM-742INT; Omron, Rio de Janeiro, Brazil). Hypertension was deﬁned as a systolic blood pressure ≥ 140 mmHg, diastolic blood pressure ≥90 mmHg, or current use of antihypertensive drugs . The presence of CV risk factors and of the metabolic syndrome were deﬁned as recommended by the Joint Interim Statement of the International Diabetes Federation Task Force on Epidemiology and Prevention; National Heart, Lung, and Blood Institute; American Heart Association; World Heart Federation; International Atherosclerosis Society; and International Association for the Study of Obesity . Increased WC was deﬁned by the cutoff ≥88 cm. The Framingham General Cardiovascular Risk Score (10-year risk) (FRS) was computed, using lipids, through the online interactive risk score calculator available on the Framingham Heart Study website . 2.3. Biochemical and hormone assays Serum FSH and estradiol (E2) levels, as well as glucose and lipid proﬁle (triglycerides, total cholesterol, and high-density lipoprotein [HDL] cholesterol) were determined in a 12-h fasting blood sample. All samples were obtained between 8 and 10 a.m. Glucose and lipid proﬁle were determined by colorimetric–enzymatic methods (Bayer 1800 Advia System, Mannheim, Germany), with a coefﬁcient of variation <3.4%. Low-density lipoprotein (LDL) cholesterol was estimated indirectly using the Friedewald formula . FSH was measured with chemiluminescent immunoassays (Centaur XP, Roche Diagnostics, Mannheim, Germany) with a sensitivity of 0.3 IU/L and intra- and interassay coefﬁcients of variation of 2.9% and 2.7%, respectively. Estradiol was measured by ECLIA (Roche Diagnostics, Mannheim, Germany), with an assay sensitivity of
5.0 pg/mL and intra- and interassay coefﬁcients of variation of 5.7% and 6.4%, respectively. 2.4. Measurement of carotid intima–media thickness Carotid IMT (C-IMT) was assessed bilaterally by B-mode ultrasonography (Xsario, Toshiba) by a single operator (an expert sonographer), using a standardized protocol. A 7.5-MHz, ﬁxedangle, multi-frequency linear array probe was used. The right and left carotid arteries were scanned to obtain a total of nine images, of the far wall of the common carotid (1 cm proximal to the carotid bulb), of the carotid bulb (1 cm proximal to the ﬂow divider), and of the proximal internal carotid arteries (1 cm distal from the ﬂow divider) [17,18]. In each segment, three measurements of maximum IMT were obtained. Subsequently, the average IMT of the three segments was calculated for each of the two carotid arteries . Subclinical atherosclerosis was deﬁned as IMT >0.9 mm and/or the presence of atherosclerotic plaque in any of the studied segments . Plaque was deﬁned by at least two of the following three criteria: C-IMT >1.5 mm; shape abnormalities such as protrusion into the lumen or loss of alignment with adjacent arterial wall boundary; the presence of brighter echoes than adjacent boundaries . 2.5. Coronary artery calciﬁcation assessment CAC was assessed by chest CT, using a 64-slice multidetector system (LightSpeed VCT, GE Healthcare). All participants were scanned by certiﬁed technologists. Sixty-four contiguous images were acquired, beginning at the carina and proceeding caudally during a single breath-hold at 40% or 65% of the R–R interval, depending on heart rate. A radiologist read all scans. The readerwork station interface identiﬁed and quantiﬁed CAC from images calibrated according to calcium phantom readings. Total coronary calcium was quantiﬁed as proposed by Agatston et al. . 2.6. Statistical analysis All descriptive data were expressed as mean ± standard deviation (SD) or median and interquartile range (IQR). The Student’s t test was used for comparisons between group means, and the Mann–Whitney U test for comparisons between median values. Spearman rank or Pearson correlation coefﬁcients were calculated between variables using a two-tailed signiﬁcance test for variables with a Gaussian or non-Gaussian distribution, respectively. The regression model was used to estimate independent associations between common carotid IMT (IMT-CC) and age, BMI, and estradiol levels. Adjustments were made according to systolic blood pressure and LDL-cholesterol levels. IMT-CC values were log-transformed for multiple regression analysis. All analyses were performed using the Statistical Package for the Social Sciences (SPSS) version 16 (SPSS Inc., Chicago, IL, USA). Findings were deemed signiﬁcant at p < 0.05. 3. Results The distribution of clinical, biochemical, and hormonal variables is shown in Table 1. The mean age of participants was 55.5 (±5) years, and the median (IQR) duration of menopause was 5.8 (3–10) years. A low/medium FRS was present in 97.9% of participants. Using the atherosclerotic cardiovascular disease (ASCVD) risk estimator 56.6% of participants presented a risk lower than 7.5% and all other had an intermediate risk. Hypertension was present in 38 participants (39.2%), all of whom were on antihypertensive drugs and 26 participants (26.8%) had metabolic syndrome. LDL cholesterol
M.A. Maturana et al. / Maturitas 81 (2015) 311–316
Table 1 Metabolic, anthropometric, and biochemical proﬁle of postmenopausal women (n = 97). Variable
Age (years) Duration of menopause (years)a BMI (kg/m2 ) Waist circumference (cm) Systolic blood pressure (mmHg) Diastolic blood pressure (mmHg) Total cholesterol (mg/dL) LDL-cholesterol (mg/dL) Triglycerides (mg/dL) Fasting blood glucose (mg/dL) Estradiol (pg/mL) Low FRS (<10%) Moderate FRS (10–20%) High FRS (>20%)
55.5 (5) 5.8 (3–10) 27.2 (5) 86.1 (10) 127 (18) 78 (10) 215.6(33) 138 (28) 116.7 (62) 93.1 (8) 22.4 (11)
76 (78.3) 19 (19.6) 2 (2.1)
BMI, body mass index; FRS, Framingham risk score. a Median (interquartile range).
levels >160 mg/dL were found in 20% and 21% presented triglycerides levels >150 mg/dl. Two women were on statins, and one was taking aspirin. Table 2 shows carotid IMT, presence of plaque, and CAC scores for the study participants. Overall, 35.1% of participants had subclinical atherosclerosis on carotid ultrasound evaluation and 24.7% had plaque present, all with a low degree of obstruction (<50% stenosis). The CAC score was zero in 57 subjects (Table 2). Only seven women had a CAC score ≥100 and two-presented ≥200. Among women with FRS <10% (low risk), 32% had subclinical CVD and 20% had plaque. Fig. 1 shows that FRS was higher in women with carotid IMT >0.9 mm and/or the presence of atherosclerotic plaque. Fig. 2 shows the correlations between carotid IMT and age, duration of menopause and CAC score, as well as between IMT-CC and endogenous estradiol levels. IMT-T was associated with age (Fig. 2A) (rs = 0.293, p = 0.004), duration of amenorrhea (Fig. 2B) (rs = 0.237, p = 0.020), and CAC score (Fig. 2C) (rs = 0.468, p < 0.001). IMT-CC was negatively associated with endogenous estradiol levels (Fig. 2D) (rs = −0.211, p = 0.038). A multivariate regression analysis was performed to assess independent associations of variables with common carotid IMT. Table 3 shows that BMI were independently associated with IMT CC. A positive correlation was found between age and IMT-CC and negative between IMT-CC and estradiol but these correlations did not remain signiﬁcant when adjusted for LDL-C levels and systolic blood pressure.
Table 2 Carotid intima–media thickness, presence of plaque, and coronary artery calciﬁcation in postmenopausal women (n = 97). Variable
IMT (mm) IMT-IC (mm) IMT-Bulb (mm) IMT-CC (mm) CAC score = 0 CAC score ≤50 CAC score >50 IMT >0.9 mm and/or plaque IMT >0.9 mm Plaque
0.73 (0.17) 0.72 (0.36) 0.82 (0.23) 0.65 (0.12)
57 (58.8) 88 (90.7) 9 (8.2) 34 (35.1) 36 (37.1) 24 (24.7)
IMT, carotid intima–media thickness; IMT-IC, internal carotid intima–media thickness; IMT-CC, common carotid intima–media thickness; CAC, coronary artery calciﬁcation.
Fig. 1. Framingham risk score stratiﬁed by the presence of carotid IMT >0.9 mm and/or atherosclerotic plaque.
Table 3 Multiple regression analysis of common carotid intima–media thickness vs. age, estradiol, and BMI. Independent variables
Age Estradiol BMI
0.210 −0.237 0.260 R2 = 0.145
0.033 0.018 0.010
0.199 −0.201 0.221 R2 = 0.133
0.072 0.062 0.047
BMI, body mass index. Dependent variable: log-transformed common carotid intima–media thickness (mean of six measurements obtained in the common carotid arteries). ˇa and pa : adjusted for systolic blood pressure and LDL-cholesterol.
4. Discussion The present study found a moderately high prevalence of carotid IMT changes, suggesting an early subclinical CVD in a sample of postmenopausal women with predominantly low FRS values. In this sample of women from Southern Brazil, 35.1% of participants had subclinical atherosclerosis, with a mean carotid IMT of 0.73 (±0.17) mm, and 24.7% of patients were found to have atherosclerotic plaque. Additionally, we found that IMT-CC was positively associated with age and negatively with endogenous estradiol levels but these associations were dependent of blood pressure and LDL-cholesterol levels. In contrast, BMI was independently associated with IMT-CC. The prevalence of subclinical atherosclerosis increases with age, and seems to be higher in men . Our results, some methodological differences notwithstanding, are similar to those reported by other authors. A Taiwan population study that included more than 2500 persons found a mean IMT of 0.68 (±0.16) mm in women aged 55–64 years . Participants screened for the Kronos Early Estrogen Prevention Study (KEEPS) presented an IMT of 0.726
M.A. Maturana et al. / Maturitas 81 (2015) 311–316
Fig. 2. Correlations of carotid IMT with age and duration of menopause and of IMT-CC with endogenous estradiol levels. (A) IMT-T and age; (B) IMT-T and duration of menopause; (C) IMT-T and CAC score; (D) IMT-CC and endogenous estradiol.
(±0.90) . A large Spanish study found a median IMT of 0.67 (0.59–0.76) mm in women with a mean age of 59 (±12) years . The prevalence of subclinical carotid atherosclerosis was higher in the Healthy Women Study (HWS) , which evaluated 200 healthy postmenopausal women aged 52–60 years and found a mean IMT of 0.76 (±0.11) mm; furthermore, 50% of the population had plaque. The higher prevalence of atherosclerotic plaque in the HWS may be at least partly explained by the high rate of smokers (64%) and by differences in methodological deﬁnition of plaque. The prevalence of plaque among pre- and postmenopausal women included in the HWS was 25% and 57%, respectively [23,24]. Woodard et al. , in a study that included African-American and white women during the menopause transition, found a lower
prevalence of plaque: 15.14%. Although the median age was 50, only 30.7% of participants were postmenopausal . Recently, Lambrinoudaki et al. , in a study that included 120 postmenopausal women (mean age 53 years), showed a combined IMT (median total measurements of bilateral carotids) of 0.724 (±0.127), and presence of plaque in 28% of participants. Taken together, these data suggest that menopause has a negative impact on vascular health, and the high prevalence of subclinical CVD in patients considered to be at low CV risk by conventional scores emphasizes the need for additional CVD evaluation. In turn, participants of our study presented a low prevalence of coronary calciﬁcation, being ≤50 in 90.7% of which 58.8% was zero. These results are similar to those reported for
M.A. Maturana et al. / Maturitas 81 (2015) 311–316
women being screened for the KEEPS, which was around 14.5%. . Physiological changes associated with menopause seem to contribute to an unfavorable CVD risk proﬁle. These changes include the cessation of ovarian estrogen secretion, changes in several CV risk factors such as body fat distribution [26,27], hypertension , and adverse changes in lipid proﬁle  and endothelial and autonomic function [30,31], as well as the metabolic syndrome and insulin resistance . Additionally, previous studies focusing on IMT have reported a relationship between the menopausal transition and increase in subclinical CVD rated [8,24,33]. In the longitudinal SWAN Study of women in the menopausal transition, the overall rate of change in IMT was 0.007 mm/year. Moreover, the progression rate of IMT increased substantially in the late perimenopausal stage, independently of age and race (0.017 mm/y) . The present study also suggests that IMT correlates positively with duration of menopause. In turn, premature ovary insufﬁciency (POI) and bilateral oophorectomy in young women have been associated with an increased incidence of CVD, myocardial infarction, and overall mortality. Observational studies suggest an interval of 5–10 years between loss of ovarian function and increased CVD risk [34,35]. These ﬁndings suggest that changes in the endogenous sex hormone milieu (notably endogenous estradiol), whether gradually during the menopausal transition or acutely in POI patients, are a major factor in the genesis of CVD in women. However, besides menopause, age also contributes to CVD in women as reported in a study by Vaidya et al. , in which cross sectional analyses of age-related CV mortality in different women’s birth cohorts have not found any abrupt acceleration at the menopause period. In the last decade, many studies have focused on the association between endogenous estradiol and subclinical CVD markers, with no consistent results. Creatsa et al. , in a study of 120 healthy postmenopausal women, found that estradiol levels were not related to brachial artery FMD or pulse wave velocity. Ouyang et al.  did not ﬁnd a signiﬁcant correlation of estradiol levels with either IMT-CC or CAC in postmenopausal women from the Multi-Ethnic Study of Atherosclerosis, which included 1947 postmenopausal women aged 45–84 years. Some studies have shown a favorable effect of endogenous estradiol on the vasculature [6,8,38]. Recently, El Khoudary et al. , in the SWAN Study, demonstrated the impact of endogenous sex hormones in the progression of subclinical atherosclerosis during the menopausal transition. In that study, lower estradiol and SHBG levels were associated with subclinical atherosclerosis progression, independently of blood pressure, BMI, and lipids. Pappa et al.  have also reported a beneﬁcial inﬂuence of endogenous estrogen on the vasculature and on IMT. Our current results are in line with these reports, showing that endogenous estrogen levels were inversely associated with IMTCC in a sample of recent postmenopausal women and that blood pressure and LDL-cholesterol levels may have a negative impact on these associations. In this sense, it is important to underline that around a third of our patients presented some degree of hypertension and/or dyslipidemia. In contrast, the lack of inﬂuence of endogenous estrogen on IMT in some other studies [36,37] might be, at least in part, explained by the fact that participants were older and had a longer duration of menopause. In fact, molecular mechanisms of estrogens on CV system seem to be related to endothelial function and the degree of atherosclerosis progression, which are intensiﬁed at and after the menopause . In this regard, endogenous estradiol levels might be related to lower subclinical CV markers only in relatively young and recent postmenopausal women, in which the detrimental effects of aging on vasculature has not yet been established [6,8].
The strengths of our study include providing data on a less well-represented ethnic group, women from Southern Brazil. One limitation of the present study is the sample size, which may have limited the statistical power for some analyses, such as those related to metabolic and anthropometric variables and other reproductive factors, as previously reported [20,24,40]. However, even large studies have been unable to elucidate the broad range of IMT results observed . Other limitation is the cross-sectional design, which does not allow a cause-and-effect direction. Further longitudinal and/or experimental studies are needed to clarify other contributory mechanisms that could explain the range of variability of IMT values. In conclusion, the present study found a moderately high prevalence of subclinical atherosclerosis in a sample of postmenopausal women with low/medium CV risk as determined by the FRS, and suggests that BMI, age and endogenous estrogen, as well as cardiovascular risk factors, such as blood pressure and LDL-cholesterol levels may impact on the presence of subclinical CVD in recently postmenopausal women. Contributors Maria Augusta Maturana: participated in the study design, data collection and analysis, in drafting the article and approved the ﬁnal version. Roberta M.C. Moreira: participated in data collection and analysis and approved the ﬁnal version. Marcela Metzdorf: participated in data collection and analysis and approved the ﬁnal version. Thais Rasia da Silva: participated in data collection and analysis and approved the ﬁnal version. Poli Mara Spritzer: participated in the study design, data analysis, in drafting the article and approved the ﬁnal version. Competing interests The authors declare that they have no conﬂict of interest. Funding This work was supported by grants from Brazilian National Institute of Hormones and Women’s Health/Conselho Nacional de Desenvolvimento Cientíﬁco e Tecnológico (CNPq INCT 573747/2008-3), Brazil. The funding sources were not involved in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in the decision to submit the paper for publication. Ethics Ethical approval was provided by Androgens endogenous SHBG: association with variable metabolic, endothelial, autonomic activity and cardiovascular disease in subclinical postmenopausal. References  Colditz GA, Willet WC, Stampfer MJ, Rosner B, Speizer FE, Hennekens CH. Menopause and the risk of coronary heart disease in women. N Engl J Med 1987;316:1105–10.  Vaidya D, Becker DM, Bittner V, Mathias RA, Ouyang P. Ageing, menopause, and ischaemic heart disease mortality in England, Wales, and the United States: modelling study of national mortality data. BMJ 2011;343:d5170.  Davis PH, Dawson JD, Riley WA, Lauer RM. Carotid intimal–medial thickness is related to cardiovascular risk factors measured from childhood through middle age: the Muscatine Study. Circulation 2001;104:2815–9.  Koskinen J, Kähönen M, Viikari JSA, et al. Conventional cardiovascular risk factors and metabolic syndrome in predicting carotid intima–media thickness progression in young adults the cardiovascular risk in young Finns study. Circulation 2009;120:229–36.
M.A. Maturana et al. / Maturitas 81 (2015) 311–316
 Baldassarre D, Nyyssonen K, Rauramaa R, et al. Cross-sectional analysis of baseline data to identify the major determinants of carotid intima–media thickness in a European population: the IMPROVE study. Eur Heart J 2010;31: 614–22.  Pappa T, Alevizaki M. Endogenous sex steroids and cardio- and cerebro-vascular disease in postmenopausal period. Eur J Endocrinol 2012;167:145–56.  Stamatelopoulos KS, Armeni E, Georgiopoulos G, et al. Recently postmenopausal women have the same prevalence of subclinical carotid atherosclerosis as age and traditional risk factor matched men. Atherosclerosis 2012;221(2):508–13.  El Khoudary SR, Wildman RP, Matthews K, Thurston RC, Bromberger JT, SuttonTyrrell K. Progression rates of carotid intima-media thickness and adventitial diameter during the menopausal transition. Menopause 2013;20:8–14.  Lambrinoudaki I, Armeni E, Georgiopoulos G, et al. Subclinical atherosclerosis in menopausal women with low to medium calculated cardiovascular risk. Int J Cardiol 2013;164(1):70–6.  Nambi V, Chambless L, Folsom AR, et al. Carotid intima–media thickness and presence or absence of plaque improves prediction of coronary heart disease risk: the ARIC (Atherosclerosis Risk In Communities) study. J Am Coll Cardiol 2010;55:1600–7.  Peters SA, Ruijter HM, den, Bots ML, Moons KG. Improvements in risk stratiﬁcation for the occurrence of cardiovascular disease by imaging subclinical atherosclerosis: a systematic review. Heart 2012;98:177–84.  Folsom AR, Kronmal RA, Detrano RC, et al. Coronary artery calciﬁcation compared with carotid intima–media thickness in the prediction of cardiovascular disease incidence: the Multi-Ethnic Study of Atherosclerosis (MESA). Arch Intern Med 2008;168:1333–9.  Chobanian AV, Bakris GL, Black HR, et al. The Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure: the JNC 7 report. JAMA 2003;289:2560–72.  Alberti KG, Eckel RH, Grundy SM, et al. Harmonizing the metabolic syndrome: a joint interim statement of the International Diabetes Federation Task Force on Epidemiology and Prevention; National Heart, Lung, and Blood Institute; American Heart Association; World Heart Federation; International Atherosclerosis Society; and International Association for the Study of Obesity. Circulation 2009;120:1640–5.  Framingham, Heart Study from: www.framinghamheartstudy.org/riskfunctions/cardiovascular-disease/10-year-risk.php [viewed September 2014].  Friedewald WT, Levy RI, Fredrickson DS. Estimation of the concentration of low-density lipoprotein cholesterol in plasma, without use of the preparative ultracentrifuge. Clin Chem 1972;18:499–502.  Burke GL, Evans GW, Riley WA, et al. Arterial wall thickness is associated with prevalent cardiovascular disease in middle-aged adults. The Atherosclerosis Risk in Communities (ARIC) Study. Stroke 1995;26:386–91.  Roman MJ, Naqvi TZ, Gardin JM, Gerhard-Herman M, Jaff M, Mohler E. American Society of Echocardiography; Society for Vascular Medicine and Biology. American Society of Echocardiography report. Clinical application of noninvasive vascular ultrasound in cardiovascular risk stratiﬁcation: a report from the American Society of Echocardiography and the Society for Vascular Medicine and Biology. Vasc Med 2006;11(3):201–11.  Agatston AS, Janowitz WR, Hildner FJ, Zusmer NR, Viamonte Jr M, Detrano R. Quantiﬁcation of coronary artery calcium using ultrafast computed tomography. J Am Coll Cardiol 1990;15:827–32.  Su TC, Chien KL, Jeng JS, et al. Age- and gender-associated determinants of carotid intima–media thickness: a community-based study. J Atheroscler Thromb 2012;19:872–80.  Miller VM, Black DM, Brinton EA, et al. Using basic science to design a clinical trial: baseline characteristics of women enrolled in the Kronos Early Estrogen Prevention Study (KEEPS). J Cardiovasc Transl Res 2009;2:228–39.
 Grau M, Subirana I, Agis D, et al. Carotid intima–media thickness in the Spanish population: reference ranges and association with cardiovascular risk factors. Rev Esp Cardiol (Engl Ed) 2012;65:1086–93.  Lassila HC, Tyrrell KS, Matthews KA, Wolfson SK, Kuller LH. Prevalence and determinants of carotid atherosclerosis in healthy postmenopausal women. Stroke 1997;28(3):513–7.  Sutton-Tyrrell K, Lassila HC, Meilahn E, Bunker C, Matthews KA, Kuller LH. Carotid atherosclerosis in premenopausal and postmenopausal women and its association with risk factors measured after menopause. Stroke 1998;29:1116–21.  Woodard GA, Narla VV, Ye R, et al. Racial differences in the association between carotid plaque and aortic and coronary artery calciﬁcation among women transitioning through menopause. Menopause 2012;19:157–63.  Donato GB, Fuchs SC, Oppermann K, Bastos C, Spritzer PM. Association between menopause status and central adiposity measured at different cutoffs of waist circumference and waist-to-hip ratio. Menopause 2006;13:280–5.  Lovejoy JC, Champagne CM, de Jonge L, Xie H, Smith SR. Increased visceral fat and decreased energy expenditure during the menopausal transition. Int J Obes (Lond) 2008;32(6):949–58.  Zanchetti A, Facchetti R, Cesana GC, et al. Menopause-related blood pressure increase and its relationship to age and body mass index: the SIMONA epidemiological study. J Hypertens 2005;23:2269–76.  Matthews KA, Meilahn E, Kuller LH, Kelsey SF, Caggiula AW, Wing RR. Menopause and risk factors for coronary heart disease. N Engl J Med 1989;321:641–6.  Maturana MA, Rubira MC, Consolim-Colombo F, Irigoyen MC, Spritzer PM. Androgenicity and venous endothelial function in post-menopausal women. J Endocrinol Invest 2010;33:239–43.  Franz R, Maturana MA, Magalhães JA, Moraes RS, Spritzer PM. Central adiposity and decreased heart rate variability in postmenopause: a cross-sectional study. Climacteric 2013;16(5):576–83.  Janssen I, Powell LH, Crawford S, Lasley B, Sutton-Tyrrell K. Menopause and the metabolic syndrome: the Study of Women’s Health Across the Nation. Arch Intern Med 2008;168(14):1568–75.  Johnson BD, Dwyer KM, Stanczyk KZ, et al. The relationship of menopausal status and rapid menopausal transition with carotid intima–media thickness progression in women: a report from the Los Angeles Atherosclerosis Study. J Clin Endocrinol Metab 2010;95(9):4432–40.  Archer DF. Premature menopause increases cardiovascular risk. Climacteric 2009;12(Suppl 1):26–31.  Goldmeier S, De Angelis K, Rabello Casali K, et al. Cardiovascular autonomic dysfunction in primary ovarian insufﬁciency: clinical and experimental evidence. Am J Transl Res 2013;6(1):91–101.  Creatsa M, Armeni E, Stamatelopoulos K, et al. Circulating androgen levels are associated with subclinical atherosclerosis and arterial stiffness in healthy recently menopausal women. Metabolism 2012;61(2):193–201.  Ouyang P, Vaidya D, Dobs A, et al. Sex hormone levels and subclinical atherosclerosis in postmenopausal women: the Multi-Ethnic Study of Atherosclerosis. Atherosclerosis 2009;204(1):255–61.  Wildman RP, Colvin AB, Powell LH, et al. Association of endogenous sex hormones with the vasculature in menopausal women: the Study of Women’s Health Across the Nation (SWAN). Menopause 2008;15(3):414–21.  Novella S, Heras M, Hermenegildo C, Dantas AP. Effects of estrogen on vascular inﬂammation: a matter of timing. Arterioscler Thromb Vasc Biol 2012;32:2035–42.  Stöckl D, Peters A, Thorand B, et al. Reproductive factors, intima media thickness and carotid plaques in a cross-sectional study of postmenopausal women enrolled in the population-based KORA F4 study. BMC Women’s Health 2014;14:7.