Pre-clinical atherosclerosis evaluated by carotid artery intima- media thickness and the risk factors in children

Atherosclerosis (AS) is one of the cardiovascular diseases that is known to be the leading reason associated with premature death nowadays, and previous studies^1,2 have shown that there are minimal vascular pathologic changes in high risk children, infants and even during fetal development. Therefore, early detection of pre-clinical atherosclerosis, finding out the cardiovascular risk factors and adopting preventive measures are of great value in preventing AS in adulthood. Carotid artery intima-media thickness (cIMT) is recognized as an important predictive measure of clinical atherosclerosis events. Studies in adults showed that the measurement of the cIMT represented an excellent marker of pre-clinical atherosclerosis.^3,4 Similarly, studies in children with hypercholesterolemia demonstrated an increased cIMT as compared with control children.^5,6 Using high resolution B-mode ultrasound to examine the cIMT was demonstrated as useful in finding the early carotid structure changes.^7-11 However, information on the association of cIMT with different risk factors measured in childhood is limited. Hence, this study was designed to find out the risk factors of AS through measuring the cIMT in children and to help find ways to prevent AS as early as possible.

Subjects
Totally 79 children 5 to 18 years of age, mean (9.7±3.0) years, were enrolled in the study. The subjects were divided into two groups according to their ages: group I consisted of 42 children with ages from 5 to 9 years and group II consisted of 37 children with ages from 10 to 18 years. Twenty-three out of the 79 subjects had positive family histories of hypertension, obesity or dislipidemia while the remaining 56 subjects did not have such family histories. All of 79 children did not have history of cardiovascular diseases.

Protocol
The body mass index (BMI) = weight (kg)/height² (m²). A 2 ml blood sample for each subject was collected by vein puncture after 12 hours of fasting and plasma concentrations of total cholesterol (TC), triglycerides (TG), high density lipoprotein (HDL) and low density lipoprotein (LDL) cholesterol concentrations were detected. Also, the atherosclerotic index (AI) was calculated based on the equation: AI =LDL-C/HDL-C.

Measurement of serum lipid components
The blood samples were collected and TC, TG, HDL-C and LDL-C were determined by Automatic Biochemical Analyzer (Hitachi, Japan). The 75th and 90th percentiles of TC and LDL-C levels were adopted as the borderline and high levels of hypercholesterolemia. The 90th percentile of TG served as the cut-off for hypertriglyceridemia. And the 5th percentile of HDL-C acted as the lower limit of dyslipidemia in the children.

Measurement of cIMT　
High resolution carotid ultrasonographic studies were performed with the Philips HDI 5000 with a 5-12 MHz transducer. Before beginning the ultrasound imaging, subjects laid quietly on the bed. The investigators scanning protocol was for the right and left carotid arteries. The study protocol involved scanning of the far wall of the right and left common carotid arteries in the distal 1.0-1.5 cm. In each examination different scanning angles (anterior, later-posterior) were adopted to record the greatest intima-media thickness (Fig). For the purpose of measurement, the examiners were kept blinded to the subject. This research refers to a normal cIMT reference of healthy children in 2004.¹² We defined the cIMT as thickened when it was ≥ 0.45 mm in children younger than 10 years old or it was ≥ 0.55 mm in children who were ≥ 10 years old and ≤ 18 years old.

Statistical analysis
All analyses were performed using the SPSS 10.0 statistical software. Values were expressed as mean±standard deviation (SD). Comparisons were done between the two groups using Student's t tests and chi-square analysis. BMI, blood pressure, levels of lipids, family history and the cIMT were calculated by conditional Logistic regression analysis. P<0.05 was considered statistically significant.

General examination
There was no positive sign on physical examination of the 79 children; and no one had xanthoma on the skin. Purple striae were found in 4 out of 79 children (5.5%) whose BMI was (34.89±3.50) kg/m². But, for those without purple striae, BMI was (19.37±5.08) kg/m². Serum TC, TG, HDL-C and LDL-C levels were (4.41±1.26) mmol/L, (1.04±0.50) mmol/L, (1.26±0.43) mmol/L and (2.68±1.14) mmol/L in the 79 subjects, respectively. And AI was 2.30±1.14 in the above cases. The echocardiographic study of the 79 children showed a normal cardiac structure and function. Out of 23 subjects who had positive family history of hypertension, obesity or dyslipidemia, 17 presented with an increased cIMT. Compared with that of children without those histories, the incidence of the increased cIMT was significantly high (χ²=4.364, P<0.05).

Correlation of BMI, blood pressure and serum lipids with cIMT
A multiple regression analysis was performed where BMI, SBP, DBP, serum TC, TG, HDL-C, LDL-C and AI were selected as independent variables and cIMT as the dependent variable. And Logistic regression analysis for the two groups was conducted. In group I, it was found that DBP and AI correlated well with the cIMT and the regression equation was created as follows: cIMT=-18.208+0.171×DBP+3.615×AI. In this model, DBP and AI were suggested to be the risk factors to the thickened cIMT, where the odds radio (OR) for DBP was 1.187 with a 95% confidence interval of (1.012, 1.392) (P=0.035) and OR for AI was 37.165 with a 95% confidence interval of (4.025, 343.133) (P=0.001). In group II, an equation cIMT=-32.454+0.316×DBP+3.425×AI was created. In this model, DBP and AI were found to be the risk factors for the thickening of cIMT. OR for DBP was 1.371 with a 95% confidence interval of (1.083, 1.736) (P=0.009), and OR for AI was 30.724 with a 95% confidence interval (1.933, 448.4) (P=0.015).

DISCUSSION

Atherosclerosis is a slow and progressive disease that starts even in childhood. The pathological abnormalities often involve coronary arteries and carotid arteries and usually manifest clinically much later in life. In 1986, Pignoli¹³ et al found that there was no significant difference between cIMT and thickness measured by ultrasound imaging. High resolution B-mode ultrasonography is a non-invasive method allowing us to assess in vivo abnormalities in vascular structure and function. In particular, carotid intima-media thickness is commonly used as a measure for detecting pre-clinical atherosclerosis, especially for those who have risk factors, such as the offspring's of patients who had hypercholesterolemia, diabetes or cardiovascular diseases.^7-10 Cuomo et al¹⁰ found that arterial abnormalities in individuals with a familial predisposition to coronary artery disease occur early and can be easily detected in children. This research was aimed to find out the risk factors of subclinical atherosclerosis in Beijing children through using high resolution B-mode ultrasonograph to measure the cIMT.

Pathological changes in AS accrue under the endothelial cells. Fatty streaks and yellow spots and plaques appeared on the intima of the aorta, which makes it possible to find the thickened cIMT through ultrasound. cIMT has been associated with many factors. There are differences between ethnic groups and areas. The previous study¹⁴ indicated that the IMT of the common carotid artery in blacks were thicker than that of white; and males in eastern Europe were also found to have thicker cITM than males in the west.¹⁵ The normal cIMT of Brazilian children from 10 to 18 years old¹⁶ which was different from that of Beijing children.¹²

Considering the reasons mentioned above, cIMT may be related to many factors, such as the gender, height of the body, weight, and endocrine function. The risk factors of AS such as dyslipidemias, obesity, hypertension, diabetes, and insulin resistance were proved by the previous studies. In Pathobiological Determinants of Atherosclerosis in youth (PDAY), the investigators studied 2876 people from 15 to 34 years old who died in accidents. Their common artery pathological biochemical studies indicated that the severity of fatty streaks had a significant positive correlation with the LDL-C level and VLDL-C level but a negative correlation with the HDL-C level.^17-19

There was also a 26-year follow-up of participants in the Framingham Heart Study showing that obesity was an independent risk factor causing AS with a quick progression.²⁰ High risk factors for children for AS were defined as obesity, diabetes, hypertension and those with a positive family history of cardiovascular disease (especially those with premature cardiovascular disease). However, up to now, there was a lack of understanding of the risk factors for pre-clinical atherosclerosis in Chinese children.

This present study demonstrates that cIMT was closely related to obesity, dyslipidemia, hypertension, and even with a positive family history. In our study, out of 23 subjects who had a positive family history of hypertension, obesity or dyslipidemia, 17 presented with an increased cIMT. Compared with children without those histories, the incidence of the increased cIMT was significantly high (χ²=4.364, P<0.05). A comparison of the cIMT between left and right carotid arteries showed that there were no significant differences between left and right carotid arteries in cIMT (t=-1.319, P>0.05) and there was a close positive correlation between cIMT in the left and right carotid arteries (r=0.983, P<0.01). In group I, serum TC, TG, LDL-C and AI were significantly higher in the cIMT thickened children than those with normal cIMT. In group II, BMI, SBP, DBP and serum TC, LDL-C and AI were significantly higher in those with thickened cIMT than in those with normal cIMT. Multiple Logistic regression analysis showed that cIMT had a quantitative relationship with DBP and AI, suggesting that they were possible risk factors for pre-clinical AS.

Ai et al¹² found that there was no significant difference in cIMT between boys and girls in 2004. They found that cIMT in children younger than 10 years of age differed significantly from those beyond that age. Therefore, in the present study we divided our study population into 2 groups according to the ages.

In conclusion, this study, by measuring cIMT and analyzing the risk factors between children with thickened cIMT and those with normal cIMT, demonstrated that diastolic blood pressure and AI were the risk factors in pre-clinical arthrosclerosis. Regularly monitoring the high-risk children for the development of and speed of progression of vascular lesions would be of great value in avoiding and preventing cardiovascular disease.

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