Metabolic fuel utilization and subclinical atherosclerosis in overweight/obese subjects
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- Montalcini, T., Gazzaruso, C., Ferro, Y. et al. Endocrine (2013) 44: 380. doi:10.1007/s12020-012-9845-2
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The utilization of different macronutrients is relevant for the risk of obesity, diabetes, or the appearing of vascular complications. The Respiratory Quotient (RQ) is a parameter measuring the fuel utilizations; in fact, it can indicate the fat stores utilization or lipogenesis activation. Aim of this study was to investigate the link between the RQ and the subclinical carotid atherosclerosis presence in overweight/obese subjects. 132 subjects with body mass index at least 25, at conventional diet, underwent an Indirect Calorimetry for the measurement of the Resting Metabolic Rate as well as the RQ and an evaluation of carotid arteries with ultrasound. Biochemical analyses were also performed. The mean age was 48 ± 12 years. There was a positive relation between carotid intima-media thickness and RQ (p = 0.010), with the high value in the subgroup with high RQ (p = 0.045 vs. group with low RQ). The RQ, an index of fuel utilization, is positively associated to subclinical carotid atherosclerosis in overweight/obese individuals.
KeywordsObesityAtherosclerosisIndirect calorimetryRespiratory Quotient
Carotid intima media thickness
Resting metabolic rate
Body mass index
Systolic blood pressure
Diastolic blood pressure
Obesity is a condition in which excessive body fat accumulates to a degree that adversely affects health . This condition, that reach pandemic status in the twenty-first century, is in fact positively associated to chronic disorders such as hypertension, dyslipidemia, type 2 diabetes mellitus, coronary heart disease, and certain form of cancers [2–4].
For these reasons, and for the high number of bariatric surgical procedures over the past years , obesity is a condition having an high economic impact. Determining causation is a complex process. It is well known that a reduced energy expenditure may be one of the components behind weight gain in general population . The utilization of different macronutrients is, also, a crucial point concerning the risk of obesity . It is assumed that regulatory processes match the dietary fuel supply with energy requirements to maintain a stable body mass and adiposity . The utilization of macronutrients could be evaluated with the Indirect Calorimetry, by the measure of the ratio of CO2 production to O2 consumption, defined Respiratory Quotient (RQ). RQ depends on the mixture of energy substrates (carbohydrates, fats, and proteins) that is the combination metabolized during rest or exercise . In particular, the endogenous fat stores utilization is associated to a decreased RQ, whereas subjects tending to burn more glucose but less fat show to have high 24-h respiratory exchange ratio, i.e., increased RQ [10, 11]. Furthermore, it was found that individuals with high RQ were at high risk to gain weight during subsequent years [10–12]. However, it is not clear whether the high RQ is associated to a high cardiovascular risk.
Carotid intima media thickness (CIMT) increases with increasing numbers of risk factors, including obesity [13, 14]. Studies on CIMT showed that it is a surrogate marker indicating the risk of coronary atherosclerosis . As the impact of an increased RQ on carotid atherosclerosis is not yet investigated, aim of this study was to explore the eventual relationship between RQ and subclinical carotid atherosclerosis in overweight/obese subjects.
Subjects and methods
Participants were recruited from subjects underwent a nutritional screening tests in our Clinical Nutrition Unit among individuals enrolled in the study on the Adherence to Mediterranean Diet (protocol number 2011.48 approved by the ethic committee of Policlinico Universitario Azienda Ospedaliera Mater Domini, Catanzaro).
All participants underwent a collection of their medical history, to obtain information about the presence of cardiovascular risk factors (hyperlipidemia, hypertension, diabetes, smoking). Furthermore, smoking habit was evaluated.
In this study, 132 subjects adhering to a diet containing at least 50–55 % of energy from a variety of carbohydrate sources, as determined by a food questionnaire, not affected by established or symptomatic cardiovascular disease, or chronic illness, were consecutively recruited. The proportion of the total energy intake derived from protein, fat, and carbohydrate was calculated by the nutritional software MetaDieta 3.0.1, (Meteda srl, S. Benedetto del Tronto, Italy). Inclusion criteria were age >20 and no more than 75 years, and BMI > 25 and no more than 40 kg/m 2 (until grade 2 obesity, using criteria of other study ). All patients provided informed consent to participate in the study.
Physical examination, anthropometry, and indirect calorimetry
The subjects were evaluated in the morning after an overnight fast. Height was measured to nearest centimeter, and weight to nearest 100 g on an electronic scale. A measuring tape was used for waist circumferences (WC). WC was measured right below the ribs. The body mass index (BMI) was calculated as weight (in kilogram) divided by square of height (in meter). An individual was considered obese if his or her BMI was over 30 kg/m2. Smoking was considered present if he or she had current smoking habit. Blood pressure (BP) (including systolic blood pressure—SBP and diastolic blood pressure—DBP) was obtained in both arms in supine patients, after 5 min of quiet rest, with a mercury sphygmomanometer. A minimum of three BP readings was taken and mean values were used for the statistical analysis.
Resting metabolic rate (RMR) and RQ were measured by indirect calorimetry (Viasys Healthcare, Hoechberg, Germany) after a 12-h fast. Participants also remained inactive (no exercise) from previous day until test. The dinner before all experiments included type of foods and drinks consumed usually. Metabolic test lasted for 30 min using a transparent, ventilated canopy-hood system and after daily calibration with a reference gas mixture (95 % O2, 5 % CO2). The measurements were taken in peaceful and relaxing environment and at a constant temperature and humidity(20–23 °C). The first and final 5 min of each set were routinely discarded and the mean value of RMR relating to the remaining 20 min was used for the calculations, once the steady-state conditions were obtained. The coefficient of variation (CV) was <10 %. If steady-state could not be maintained that long, to measure mean value of RMR the 10-min with a CV < 10 % were used. RMR was calculated from O2 and CO2 volumes. Substrate oxidation was calculated from CO2 production and O2 consumption rates, and this ratio was expressed as RQ.
Venous blood was collected after overnight fasting, into vacutainer tubes (Becton & Dickinson) and centrifuged within 4 h. Serum glucose, total cholesterol, HDL cholesterol, and triglycerides were measured by standard laboratory techniques. Quality control was assessed daily for all determinations.
Ultrasound for subclinical carotid atherosclerosis assessment
The subjects underwent B-mode ultrasonography of the extracranial carotid arteries by use of a duplex system (a high resolution ultrasound instrument ATL, HDI 5000 with a 5- to 12-MHz linear array multifrequency transducer). All the examinations were performed by the same ultrasonographer blinded to clinical information. All patients rested in the supine position for at least 10 min before the study and were kept in this position during the procedure. ECG-leads were attached to the ultrasound recorder for on-line continuous heart rate monitoring. The right and left common (CCA) and internal carotid arteries (including bifurcations) were evaluated with the head of the subjects turned away from the sonographer and the neck extended with mild rotation. The IMT, defined as the distance between the intimal–luminal interface and the medial–adventitial interface, was measured as previously described . Briefly, in posterior approach and with the sound beam set perpendicular to the arterial surface, 1 cm from the bifurcation, three longitudinal measurements of IMT were completed on the right and left common carotid arteries far-wall, at sites free of any discrete plaques. The mean of the three right and left longitudinal measurements was then calculated. Then, we calculated and used for statistical analysis the mean CIMT between right and left CCA. Plaque, detected in longitudinal and transverse planes with anterior, lateral, and posterior approaches, was defined as an echogenic focal structure encroaching into the vessel lumen of at least 50 % of the surrounding IMT value. Stenosis was defined as a peak systolic velocity >120 cm/s and occlusion was defined as absence of Doppler signal. According to these criteria, subjects were considered as normal if no lesion was detected, or having carotid atherosclerosis when a plaque, stenosis, or occlusion was detected in at least one segment of common, bifurcation, or internal carotid artery. The coefficient variation of the methods was 3.3 %. To evaluate artery diameters, images were magnified, whereas depth and gain settings were set to optimize the image of the vessel wall, in particular, the media–adventitia interface (“m” line). The end-diastolic diameter of the vessel, defined as the distance between near-wall and far-wall junctions of the media and adventitia, was measured over four cardiac cycles with the use of digital callipers and the average was then calculated.
Data are reported as mean ± SD. A t test for independent samples was performed to compare CIMT means between genders. Stepwise multivariate linear regression analysis was used to test for confounding variables, defined as variables that correlated to CIMT at the univariate analysis. An ANOVA was performed to test the differences between tertiles of RQ. Significant differences were assumed to be present at p < 0.05. The simple size was 134 subjects based on the following parameters: a correlation coefficient (r2) between RQ and CIMT corrected for age of at least 0.25, an alpha equal to 0.01 and a beta equal to 0.1. All comparisons were performed using the statistical package SPSS 17.0 for Windows, Chicago (USA).
Characteristics of the population
Mean CIMT (mm)
Stepwise multivariate linear regression analysis
In this study, we showed a positive relation between CIMT and RQ in overweight and obese (grades 1 and 2) subjects, referred to our Clinical Nutrition Unit.
We also observed an association with age and the traditional cardiovascular risk factors, like SBP and WC, and the adjustment for them confirmed the association between CIMT and RQ (Table 2). Finally, we found that subjects with high RQ have highest CIMT value (Fig. 1). This is an original finding, never investigated to date. This study was not designed to explore the mechanisms responsible for the relationship between the subclinical atherosclerosis and the substrates-type utilization, but some hypothesis may be proposed. In fact in tissues and organs, fuel availability is integrated at the cellular level by fuel sensors that activate or inhibit specific metabolic pathways [18, 19]. In response to fuel oversupply, anabolic pathways are activated, as showed by the fact that high RQ indicates lipogenesis activation [10–12]. It is well known that an excess of lipid deposit can induce lipotoxicity, can impair insulin signaling through different mechanisms . In line with these studies, other authors showed that individuals able to increase fat oxidation quickly, in response to day-to-day changes in fat intake, will be less prone to insulin resistance [21, 22]. Furthermore, it is recognized that postprandial endogenous insulin can increase RQ . All these findings could lead us to speculate on a possible involvement of the endogenous insulin in the relation between RQ and CIMT .
Furthermore, it was showed that some genes, such as those involving uncoupling proteins, leptin and leptin receptor, b2-adrenergic and b3-receptors, peroxisome proliferator-activated receptors, melanocortin-4 receptor, pro-opiomelanocortin, are related to energy expenditure, and may be affected specifically by dietary intake and composition . It was also demonstrated that these genes or the proteins expressed are involved in the development of cardiovascular disease , contributing to explain partially our findings.
Interestingly, in our study there is no relationship between CIMT and BMI, while we found a positive relation between CIMT and WC. Thus, it is possible to hypothesize a detrimental effect mediated mainly by metabolic factors related to fuel utilization, on carotid arteries, rather than a simple impact of BMI [26, 27]. Furthermore, it showed a high RQ in subjects with and without obesity [28, 29] indicating, again, the main role of metabolic factors rather than BMI per se . In particular, these two parameters could be the expression of different phases in the natural history of atherosclerosis: WC could be the expression of a constant excess of caloric intake, gradually leading to the accumulation of fat in the abdominal region, increasing the atherosclerotic risk, while it is possible that RQ modification could be an early marker of subclinical atherosclerosis, before that the anthropometric variations become manifest [9, 12]. Consequently, if confirmed by prospective studies, RQ measurement could be considered, in future, a marker to identify subjects at high risk for the development of the atherosclerotic process. Furthermore, some investigations suggest that RQ measurement could be useful for the development of the appropriate nutritional therapies in the overweight condition [20, 22, 31–37]. As expected in our study we did not find any association between carotid plaques and RQ, as the young age of the enrolled population, having a relative low prevalence of risk factors. Furthermore, it was showed that the atherosclerotic plaques formation is a long-term process, linked to the levels of insulin in a complex way [22, 23]. The use of indirect calorimetry certainly composes the main strength of this work, providing an accurate measurement of RQ and REE. Indirect calorimetry is considered a simple and affordable tool for measuring energy expenditure and for quantifying the utilization of macronutrients . However, in the clinical practice, measurement of substrate oxidation is less reliable than that of energy expenditure . The adequacy of the respiratory gas exchanges should be verified, and the body compartments of the respiratory gases should be in steady-state conditions. It is also necessary to ascertain whether the subjects are in the postabsorptive state. Furthermore, in many conditions, given the importance of maintaining the energy balance in the long term, it could be important repeatedly to measure the REE by indirect calorimetry  rather than to perform a single evaluation as in our study. The small sample size and the lack of control group are other limitations. Of course our results need confirmation by prospective studies, but we think that these results could suggest to perform other investigations to better explore the implication of RQ on cardiovascular risk in overweight and obese subjects.
Conflict of interest
There is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.