Senior Member, IEEE Leandro J Cymberknop

Eugenia Ipar
email: eipar@frba.utn.edu.ar

Nicolás A Aguirre
email: naguire@frba.utn.edu.ar

Senior Member, IEEE Ricardo L Armentano

Age-Stratified Coronary Artery Calcium Levels Quantification Based on Arterial Biomechanical Parameters*

Keywords: coronary artery calcium, pulse wave velocity, linear regression

Coronary Artery Calcium (CAC) constitutes a highly specific feature of 'coronary' atherosclerosis. The aim of this study is to explore the combination of arterial biomechanical parameters, used for general atherosclerosis evaluation, in CAC score assessment, stratified by year of age. Three hundred and fifty individuals participated in the study, with no history of cardiovascular disease. CAC score jointly with ascending aortic diameter (AAD) were determined through multislice computer tomography. Carotid intima-media thickness (cIMT) and presence of carotid and femoral plaques (PAB) were also assessed using B-mode ultrasound. Aortic stiffness was calculated by means of Pulse Wave Velocity (aPWV). Data were grouped by age, thus becoming a stratification factor. Univariate and multiple linear regression (MLR) analysis were applied for CAC score quantification. Univariate analysis evidenced a linear association between logtransformed levels of CACs stratified by age (LCACs) and aPWV (R 2 =0.43, P<0.05), cIMT (R 2 =0.52, P<0.05), AAD (R 2 =0.53, P<0.05), PAB (R 2 =0.46, P<0.05) and SBP (R 2 =0.10, P<0.05). An adjusted determination coefficient (adj. R 2 ) of 0.89 was obtained for MLR, where AAD, cIMT, PAB, SBP and BMI were significantly correlated with LCACs (P<0.05). This analysis provides a holistic description in terms of the contribution of atherosclerosis/arteriosclerosis markers (such as cIMT and aortic diameters) that may suggest the existence of increased levels of age-stratified CAC.

I. INTRODUCTION

Atherosclerosis defines a pathological arterial condition, intrinsically associated with heart disease and stroke. Particularly, Coronary Artery Calcium (CAC) constitutes a highly specific feature of 'coronary' atherosclerosis, and its development is now understood as an 'active' pathogenic process. Since traditional risk scores are useful for initial method stratification, the characterization of coronary calcification by multislice computer tomography provides a substantial improvement, being significantly associated with the occurrence of major cardiovascular events, cardiac mortality, and all-cause mortality [START_REF] Greenland | ACCF/AHA 2007 clinical expert consensus document on coronary artery calcium scoring by computed tomography in global cardiovascular risk assessment and in evaluation of patients with chest pain: a report of the American College of Cardiology Foundation Clinical Expert Consensus Task Force (ACCF/AHA writing committee to update the 2000 expert consensus document on electron beam computed tomography) developed in collaboration with the Society of Atherosclerosis Imaging and Prevention and the Society of Cardiovascular Computed Tomography[END_REF], [START_REF] Neves | Coronary artery calcium score: current status[END_REF].

It is well known that the predictive value of algorithms based on conventional risk factors is manifested as insufficient, where individuals of 'intermediate risk' may benefit from further risk stratification [START_REF] Kullo | Arterial ultrasonography and tonometry as adjuncts to cardiovascular risk stratification[END_REF]. Previous studies have demonstrated that the increase in the CAC score (CACs) could be utilized to evaluate the progression of atherosclerotic plaques and the future cardiovascular risk [START_REF] Budoff | Progression of coronary artery calcium predicts all-cause mortality[END_REF]. Moreover, values of CACs higher than 400 in low/intermediate risk individuals are recommended for coronary angiography, acting as a threshold to facilitate diagnosis [START_REF] Neves | Coronary artery calcium score: current status[END_REF]. Unfortunately, tests involving radiation exposure such as CAC increase health costs, thus they are not currently recommended as cardiovascular disease (CVD) screening tools in American or European guidelines [START_REF] Greenland | ACCF/AHA 2007 clinical expert consensus document on coronary artery calcium scoring by computed tomography in global cardiovascular risk assessment and in evaluation of patients with chest pain: a report of the American College of Cardiology Foundation Clinical Expert Consensus Task Force (ACCF/AHA writing committee to update the 2000 expert consensus document on electron beam computed tomography) developed in collaboration with the Society of Atherosclerosis Imaging and Prevention and the Society of Cardiovascular Computed Tomography[END_REF].

Both age and exposure to cardiovascular risk factors denote structural and functional changes in large arteries [START_REF] Cecelja | Association of pulse wave velocity and intima-media thickness with cardiovascular risk factors in young adults[END_REF]. Vascular diseases such as atherosclerosis produce damaging effects which use to be initially localized in specific vascular segments, such as carotid and/or femoral arteries. Consequently, the evaluation of the degree of atherosclerosis (i.e. vascular damage) can be performed by measurements such local/systemic arterial stiffness, increase of arterial wall layers' thickness and presence/degree of atherosclerotic plaques as consequence of lipid deposition, among others [START_REF] Armentano | Quantitative Vascular Evaluation: From Laboratory Experiments to Point-of-Care Patient (Clinical Approach)[END_REF]. All these parameters are considered strong predictors of cardiovascular mortality and coronary heart disease (CHD) events. Moreover, central arterial stiffness was related to subclinical coronary atherosclerotic burden in asymptomatic adults [START_REF] Kullo | Arterial ultrasonography and tonometry as adjuncts to cardiovascular risk stratification[END_REF].

The aim of the present work was to explore the use of arterial biomechanical parameters for CACs quantification in a community of asymptomatic adults, considering 'age' as a stratification factor. To the best of our knowledge, the simultaneous combination of age-stratified parameters for atherosclerosis development evaluation was not previously reported in CAC level assessments.

II. MATERIALS AND METHODS

A. Study Population

Three hundred and fifty (350) individuals with no prior cardiac diseases, aged between 40 and 70 years were included in the final study population. Age, sex, body mass index (BMI) and heart rate (HR) were determined as baseline *Research supported by UNDP URU/06/004 PEDECIBA-CSIC (Uruguay) and ICUTIBA 4852 and 7647 (Argentina) research projects. demographic profiles. Systolic and Diastolic blood pressures (SBP and DBP, respectively) were measured in the right arm as the mean of at least three measurements by sphygmomanometer procedure in the supine position after 10 min of rest. Cardiovascular risk factors such as presence of hypertension (HT), current smoking (CS) and diabetes (DM) mellitus jointly with Ten-year Framingham risk score (FRS, % probability of CHD event in the next 10 years) were evaluated. Individuals with symptoms of known history of cardiovascular disease, including CHD, stroke, and peripheral vascular disease were excluded from the cohort. The research protocol was carried out in accordance with the Code of Ethics of the World Medical Association (Declaration of Helsinki) and all subjects gave their written informed consent before inclusion into the study.

B. Arterial Biomechanical Parameters Assessment 1) Coronary Artery Calcium and Ascending Aortic Diameter Quantification

All participants underwent CAC scoring at baseline. The quantification of atherosclerotic burden in coronary arteries was determined by acquiring electrocardiographic-triggered non-contrast computed tomography imaging (IMATRON, 100ms scan time, a 3 mm slice thickening, and an ECG triggering) limited to the thorax region during an apnea of approximately five seconds. The threshold for a calcified lesion was set at a density of 130 Hounsfield units with an area ≥1mm2. The CAC score was calculated from the total sum of the product between each calcified plaque area and the maximum density calcium in the lesion and expressed in Agaston Units (AU) [START_REF] Chironi | Impact of coronary artery calcium on cardiovascular risk categorization and lipid-lowering drug eligibility in asymptomatic hypercholesterolemic men[END_REF]. Moderate plaque deposits are associated with a score of 100 to 300, which implies a relatively high risk of an acute event or other heart disease over the next three to five years. A score greater than 300 shows a sign of very high to severe disease and heart attack risk [START_REF] Greenland | ACCF/AHA 2007 clinical expert consensus document on coronary artery calcium scoring by computed tomography in global cardiovascular risk assessment and in evaluation of patients with chest pain: a report of the American College of Cardiology Foundation Clinical Expert Consensus Task Force (ACCF/AHA writing committee to update the 2000 expert consensus document on electron beam computed tomography) developed in collaboration with the Society of Atherosclerosis Imaging and Prevention and the Society of Cardiovascular Computed Tomography[END_REF]. Ascending aortic diameter (AAD) was concomitantly identified and estimated from the obtained images in a dedicated workstation, considering the outer aortic wall perpendicular to the axis of rotation.

2) Pulse Wave Velocity (PWV)

Aortic stiffness was determined over the entire population through the measurement of the propagation speed of the arterial pulse wave. Carotid-to-femoral pulse wave velocity (aPWV, considered the gold standard method) was obtained by simultaneous acquisition of pulse pressure morphology in the carotid and femoral sites (using tonometric transducers), and calculated as distance over pulse transit time, according to a procedure previously reported [START_REF] Armentano | Quantitative Vascular Evaluation: From Laboratory Experiments to Point-of-Care Patient (Clinical Approach)[END_REF].

3) Intima Media Thickness (IMT) and Atheroma Plaques

Measurements were performed at carotid and femoral arteries by real-time B-mode ultrasound (Ultramark 4, Advanced Technology Laboratory) with a 7.5 MHz probe, according to a procedure previously described [START_REF] Armentano | Quantitative Vascular Evaluation: From Laboratory Experiments to Point-of-Care Patient (Clinical Approach)[END_REF]. Two conditions were evaluated at the arterial site: 1) Atheroma plaque formation, corresponding to a focal thickening making prominence in the lumen of the vessel and 2) Diffuse thickening of the arterial intima and media (cIMT measurement). Each site (carotid and femoral) was considered as diseased if one or more plaque was found regardless the location. Peripheral atherosclerosis burden (PAB) was expressed as the number of diseased sites from 0 to 2.

C. Statistical Analysis

Descriptive statistics were expressed as mean ± SD, while qualitative variables were expressed as number of participants and percentages. Due to distribution of CACs was positively skewed, the scores were log-transformed after adding 1 (LCACs=log(CACs+1)) [START_REF] Kullo | Aortic pulse wave velocity is associated with the presence and quantity of coronary artery calcium: a community-based study[END_REF], [START_REF] Chironi | Early thoracic aorta enlargement in asymptomatic individuals at risk for cardiovascular disease: determinant factors and clinical implication[END_REF]. Particularly in this work, subjects were grouped by age, being this parameter considered as an external parametric value. Then, mean values of each subgroup were obtained. In first place, univariate linear regression analysis was used to assess the individual association of LCACs (outcome variable) to arterial biomechanical parameters (cIMT, AAD, aPWV and PAB), including SBP and BMI (exposure variables). Subsequently, a multiple linear regression analysis (MLR, no intercept) was performed to assess the simultaneous interaction of all parameters with LCACs. The number of individuals included in analysis, coefficients, 95% confidence interval, P-values and adjusted coefficient of determination (Adj. R 2 ) are presented in the results. Statistical significance was determined at P<0.05.

III. RESULTS

Table I describes the demographic, hemodynamic and biochemical characteristics of the studied population. Mean age was 56.5±9.1 years and 30% were women. Average values for BMI, HR, SBP and DBP were 26.1±4.3 kg/m 2 , 65.5±10.0 bpm, 124.3±13.3 mmHg and 73.3±9.4 mmHg respectively. The prevalence rates of risk factors were 48.9% for hypertension, and 21.3% for current smoking and 6.4 % for diabetes mellitus while 10-year FRS was of 9.2±5.5 %. II, jointly with the distributions of arterial lesions and logtransformed coronary artery calcification levels. MLR analysis results are detailed in Table III. An adjusted determination coefficient (adj. R 2 ) of 0.89 was obtained. As can be seen, cIMT, AAD, PAB, SBP and BMI were significantly correlated with age-stratified levels of LCACs (P<0.05). On the other hand, aPWV did not show statistical significance.

Arterial biomechanical parameters are shown in Table

IV. DISCUSSION

In this study, arterial biomechanical parameters used in atherosclerosis characterization were considered for CACs quantification, a surrogate of coronary atherosclerosis burden. Our findings showed that cIMT, AAD, PAB, BMI and SBP behave as significant predictors of CACs levels in a MLR analysis corresponding to a population without previous story of CVD or stroke. Particularly, and unlike similar studies, age was used as a parametric value for the stratification of the population under study, not being considered as a predictor in MLR CACs evaluation. Previous publications have demonstrated that the presence of atherosclerotic lesion in specific artery sites (plaque) as well as in coronary arteries (calcification) is associated with an increased estimated risk of coronary events [START_REF] Simon | Extracoronary atherosclerotic plaque at multiple sites and total coronary calcification deposit in asymptomatic men: association with coronary risk profile[END_REF]. Additionally, associations between AAD and subclinical atherosclerosis burden reflected by CAC were also identified in [START_REF] Chironi | Early thoracic aorta enlargement in asymptomatic individuals at risk for cardiovascular disease: determinant factors and clinical implication[END_REF]. It has been also proved that carotid ultrasound-based IMT measurement, a surrogate for systemic atherosclerotic disease burden, is associated with prevalent and incident CHD and stroke [START_REF] Kullo | Arterial ultrasonography and tonometry as adjuncts to cardiovascular risk stratification[END_REF][START_REF] Nambi | Carotid intima-media thickness and presence or absence of plaque improves prediction of coronary heart disease risk: the ARIC (Atherosclerosis Risk In Communities) study[END_REF], where an increased carotid IMT was related to calcification in young and middle age adults [START_REF] Davis | Increased carotid intimal-medial thickness and coronary calcification are related in young and middle-aged adults: the Muscatine study[END_REF]. Regarding aPWV and its relationship with coronary artery calcification, a significant association between aPWV and presence and quantity of CAC was reported in [START_REF] Kullo | Aortic pulse wave velocity is associated with the presence and quantity of coronary artery calcium: a community-based study[END_REF], which was also demonstrated in this study through the age-stratification approach (Fig. 1). It is well known that aortic arterial stiffness increases with age, being also closely related to pathophysiologicals conditions such as elevated levels of blood pressure, plaque formation and arterial remodeling, among others [START_REF] Armentano | Quantitative Vascular Evaluation: From Laboratory Experiments to Point-of-Care Patient (Clinical Approach)[END_REF]. In [START_REF] Bielak | Age dependent associations between blood pressure and coronary artery calcification in asymptomatic adults[END_REF], aPWV behave as predictor of CHD and stroke, independent of conventional risk factors, measures of atherosclerosis and pulse pressure. The latter, which behaves as an indirect measure of arterial stiffness, has been also related to coronary atherosclerotic burden [START_REF] Mattace-Raso Vt | Arterial stiffness and risk of coronary heart disease and stroke: the Rotterdam study[END_REF]. Conversely, aPWV was not significantly correlated with LCACs quantification in MLR analysis, remaining as a marginally significant predictor. Considering the simultaneous presence of biomarkers of atherosclerosis development (i.e cIMT, AAD and PAB) and SBP, this condition could be overlapping the explicative contribution of aPWV to the entire the analysis.

Due to the isolated analysis of the CACs present some limitations in the evaluation of patients with suspected acute coronary syndrome, it is suggested to previously evaluate the probability of cardiovascular events to complement the interpretation of the CAC score [START_REF] Greenland | ACCF/AHA 2007 clinical expert consensus document on coronary artery calcium scoring by computed tomography in global cardiovascular risk assessment and in evaluation of patients with chest pain: a report of the American College of Cardiology Foundation Clinical Expert Consensus Task Force (ACCF/AHA writing committee to update the 2000 expert consensus document on electron beam computed tomography) developed in collaboration with the Society of Atherosclerosis Imaging and Prevention and the Society of Cardiovascular Computed Tomography[END_REF]. For this reason, the combination of atherosclerosis markers for CACs quantification contributes to a more integrated picture in CHD risk assessment. It has to be also noteworthy that CAC evaluation requires the use of ultrafast computed tomography, which is usually recommended in individuals with chest pain who are classified as being at low to intermediate risk. For CACs equal to zero AU, no other examination is required, but if the score is higher than a threshold, coronary angiography would be indicated [START_REF] Neves | Coronary artery calcium score: current status[END_REF]. Nevertheless, cost-effectiveness and feasibility of testing should be also considered. The use of simple combined measurements for the evaluation of atherosclerosis could contribute to the identification of increased levels of CAC, basically acting as a "filter" in screening tests, thus improving risk assessment and reducing costs.

In summary, this analysis provides a holistic description in terms of the contribution of atherosclerosis markers (such as cIMT and aortic diameters) that may suggest the existence of increased levels of age-stratified CAC. In this sense, the approach focuses on the evaluation of the systemic atherosclerotic/arteriosclerotic process, rather than on the development of the coronary process itself.

Figure 1 .

 1 Figure 1. Linear relationship between aortic pulse wave velocity (aPWV) and log transformation of coronary artery calcium score + 1 (CAC+1) (R 2 =0.46, P<0.05). Data were stratified by age.

Figure 2 .

 2 Figure 2. Biomechanical parameters vs. log transformation of coronary artery calcium score + 1 (CAC+1) behavior, with age considered as a parametric value. Top Panel: Ascending Aortic Diameter (AAD). Bottom panel: Intima Media Thickness (IMT)

TABLE I

 I

	. VALUES ARE EXPRESSED AS MEAN±STANDARD DEVIATION AND DEMOGRAPHICS AND CLINICAL CHARACTERISTICS. PERCENTAJE. BMI: BODY MASS INDEX, HR: HEART RATE, SBP: SYSTOLIC BLOOD PRESSURE, DBP: DIASTOLIC BLOOD PRESSURE, HT: HYPERTENSION, CS: CURRENT SMOKING, DM: DIABETES MELLITUS, FRS: FRAMINGHAM RISK SCORE.
	Parameter	Value
	Individuals (females) [n]	350 (105)
	Age [years]	56.5±9.1
	BMI [kg/m 2 ]	26.1±4.3
	HR [bmp]	65.5±10.0
	SBP [mmHg]	124.3±13.3
	DBP [mmHg]	73.3±9.4
	HT [%]	48.9
	CS [%]	21.3
	DM [%]	6.4
	10 year -FRS [%]	9.2±5.5
	Univariate analysis evidenced a linear association between LCACs and aPWV (R 2 =0.43, P<0.05), cIMT (R 2 =0.52, P<0.05), AAD (R 2 =0.53, P<0.05), PAB (R 2 =0.46, P<0.05) and SBP (R

2 =0.10, P<0.05). Data was stratified by age, thus becoming a stratification factor (Fig.

1

). The latter is used in Fig.

2

to represent age-stratified behavior (mean values for each age group) between AAD and CAC (top panel) and IMT and CAC (bottom panel).

TABLE II .

 II ARTERIAL RELATED PARAMETERS. VALUES ARE EXPRESSED AS MEAN±STANDARD DEVIATION. aPWV: AORTIC PULSE WAVE VELOCITY, AAD: ASCENDING AORTIC DIAMETER, CIMT: CAROTID IMTIMA-MEDIA THICKNESS, PAB: PERIPHERAL ATEROSCLEROSIS BURDEN, LCACS: LOG TRANSFORMED CORONARY ARTERY CALCIUM SCORE

	Parameter	Value
	aPWV [m/s]	10.6±2.7
	AAD [mm]	32.8±0.4
	cIMT [mm]	0.7±0.1
	PAB 0 [%]	24.4%
	PAB 1 [%]	32.2%
	PAB 2 [%]	43.4%
	LCACs [AU]	2.6±2.5

TABLE III .

 III MULTIPLE REGRESSION ANALYSIS FOR CAC SCORE QUANTIFICATION. aPWV: AORTIC PULSE WAVE VELOCITY, AAD: ASCENDING AORTIC DIAMETER, CIMT: CAROTID IMTIMA-MEDIA THICKNESS, PAB: PERIPHERAL ATEROSCLEROSIS BURDEN, BMI: BODY MASS INDEX, SBP: SYSTOLIC BLOOD PRESSURE

	Parameter	Coefficient	95% CI	p-value	Adj. R 2
	PAB	0.880	0.233, 1.527	0.009
	AAD	2.354	0.619, 4.088	0.009
	BMI SBP	-0.178 -0.051	-0.323, -0.033 -0.095, -0.007	0.017 0.024	0.891
	cIMT	3.865	0.260, 7.470	0.036
	aPWV	0.180	-0.066, 0.426	0.146