Branched-Chain amino acids intake is negatively related to body adiposity in individuals at cardiometabolic risk

Objective To assess the relationship between branched-chain amino acids intake in the current diet and the metabolic and body adiposity markers in a population at cardiovascular risk. Methods This is a cross-sectional study with 282 adults and elderly people from the Cardiovascular Health Care Program of the Universidade Federal de Viçosa. Sociodemographic, anthropometric and body composition data, as well as metabolic biomarkers were collected using standardized protocols. Dietary intake of branched amino acids was assessed using a 24-hour recall.

Branched-Chain amino acids intake is negatively related to body adiposity in individuals at cardiometabolic risk

A B S T R A C T Objective
To assess the relationship between branched-chain amino acids intake in the current diet and the metabolic and body adiposity markers in a population at cardiovascular risk.

Methods
This is a cross-sectional study with 282 adults and elderly people from the Cardiovascular Health Care Program of the Universidade Federal de Viçosa. Sociodemographic, anthropometric and body composition data, as well as metabolic biomarkers were collected using standardized protocols. Dietary intake of branched amino acids was assessed using a 24-hour recall.

Results
Individuals with a higher branched-chain amino acids intake (≥2.6g/day, median value) had lower body fat (29.6 vs 32.2%; p=0.019), and higher serum ferritin (113.2 vs. 60.1mg/dL; p=0.006) and uric acid concentrations (4.4 vs. 4.0; p=0.023). In addition, a lower prevalence of overweight and excessive abdominal fat (p<0.05) was found in the individuals with higher branched-chain amino acids intake. They also had a higher daily intake of fiber, copper, zinc, magnesium, and iron, as well as a lower intake of total lipids.

Conclusion
In the present study, the intake of branched amino acids is negatively related to total and central adiposity, but more studies are needed to fully elucidate this possible relationship. (

I N T R O D U C T I O N
Excess weight is the main risk factor for the development of Cardiovascular Diseases (CVD), being one of the biggest public health problems worldwide. It should be noted that eating habits are an important modifiable risk factor for such changes [1][2][3][4][5].
A recent meta-analysis [13] reported that a higher dietary intake of BCAA was inversely related to the prevalence of overweight and obesity in adults. The possible mechanisms for the effect of BCAA on body weight are still poorly understood, however it is known that BCAA, especially leucine, can act in ways to control cellular metabolism, providing a decrease in food intake and body weight [14][15][16]. However, few studies have assessed BCAA intake, especially in the Brazilian population and, to date, few studies have assessed the relationship of these nutrients with metabolic and adiposity markers in a population at cardiovascular risk [10]. Therefore, the aim of this article is to assess the relationship between BCAA intake, included in the current diet, and metabolic and adiposity biomarkers in a population at cardiovascular risk.

M E T H O D S
A cross-sectional study with 282 patients included in the Programa de Atenção à Saúde Cardiovascular da Universidade Federal de Viçosa (PROCARDIO-UFV, Cardiovascular Health Care Program of the Universidade Federal de Viçosa), who had their first medical appointment until July 2016, with complete data concerning their BCAA intake. The PROCARDIO-UFV performs nutritional intervention to promote cardiovascular health in the academic community of the Universidade Federal de Viçosa (UFV), registered in the Registro Brasileiro de Ensaios Clínicos (ReBEC, Brazilian Registry of Clinical Trials), code RBR-5n4y2g [17].
The criteria for inclusion in the program are: age ≥20 years; both sexes; being a student, worker, or a dependent family member of UFV workers; present cardiovascular diseases or the occurrence of cardiometabolic risk factors such as overweight or obesity (Body Mass Index (BMI) ≥25 or 27kg/m²) or/and dyslipidemia (Triglycerides (TG) ≥150mg/dL; Total Cholesterol (TC) ≥ 200 mg/dL or/and High Density Lipoprotein (HDL-c) <40 or <50mg/dL for men and women, respectively), or/and blood pressure ≥130/≥85mmHg or diagnosed arterial hypertension or/and fasting blood glucose ≥100mg/ dL or diagnosed diabetes mellitus. The data used were related to the first medical consultation at the PROCARDIO-UFV. Among the 417 users of the program, 282 were selected for having complete data on their BCAA intake.
The study was approved by the UFV's Human Research Ethics Committee (Protocol number 066/2012/CEPH), in accordance with Resolution 466/2012 of the Conselho Nacional de Saúde (CNS, National Health Council/Ministry of Health, Brazil) and with the principles of the Helsinki Declaration. All study participants read and signed the informed consent form.
The participants responded to a 24-hour food recall (R24h), reporting all the food and drinks consumed the day before (weekday or weekend) the medical consultation, as well as their quantities. In the present study, the daily intake of calories, carbohydrates, proteins, lipids, fibers, vitamins A, C, D and E, selenium, copper, manganese, magnesium, zinc, calcium, iron, and sodium were assessed using the DietPRO software, version 5.0i [18].
The determination of BCAA intake (leucine, isoleucine, and valine) was performed using the National Nutrient Database for Standard Reference (USDA, 2015), as such data are not available in the Brazilian tables [19]. The foods reported in the R24h and not listed in the USDA table had their estimated composition considering the foods that showed nutritional composition and similar cooking methods. The preparations were broken down into their constituent ingredients and, if there was no choice of composition for the prepared food, the composition of the raw foods was used. The intake of each amino acid was performed in an electronic spreadsheet (Microsoft Excel ® ), developed especially for this purpose.
Anthropometric measurements (body weight, height, hip, and waist circumference) were measured using a standardized, previously established protocol [20]. Waist-to-Hip (WHR) and Waist-to-Height (WHtR) ratios were calculated. Total Body Fat (BF%) was assessed by tetrapolar electrical bioimpedance (Biodynamics 310 model, Washington, USA), according to the manufacturer's protocol. Excess weight was classified according to a BMI greater than or equal to 25 (adults) and 27 (elderly) kg/m 2 [21,22]. Excess abdominal fat was assessed using waist circumference values equal to or greater than 80 cm and 90 cm for women and men, respectively [23]. The age, sex, educational level, relationship to the UFV, income, smoking, regular practice of physical activities, and intake of alcoholic beverages variables were collected by interviewing participants through a questionnaire.
The results were presented in absolute and relative frequencies, mean±standard deviation, and/or median (25 th -75 th percentile). The normality of each variable was assessed using the Kolmogorov-Smirnov test. To assess the possible association of BCAA intake and other variables of interest, the sample of the present study was categorized according to the median BCAA intake (2.6g/day). The use of the median as a cut-off point for statistical analysis has been used before [25,26]. All food intake variables were adjusted for total caloric intake using the residual method [27].
The Student t and Mann-Whitney-U tests were used to compare the two groups, when appropriate. Pearson's Chi-square, linear trend chi-square or Fischer's exact test were used when appropriate to verify associations between sociodemographic and body composition variables, and the median BCAA intake. Spearman's correlation was used to assess the relationship between BCAA intake and other nutrients with variables of interest. All statistical analyses were performed using the SSPS 21.0 ® program, considering the level of statistical significance as 5%.

R E S U L T S
Of the individuals who had a higher BCAA intake (≥2.6g/day), 28.4% (n=40) were male, 28.2% (n=40) were employees of the university, and 75.9% (n=107) were not smokers (Table 1). Regarding food intake, those with a BCAA intake ≥2.6g/day had a higher intake of fibers, copper, zinc, magnesium, and iron (Table 2).
Individuals with the highest BCAA intake (≥2.6g/day) had higher concentrations of ferritin and uric acid, in addition to lower body fat and HDL-c values (Table 3).
Moreover, there was a higher prevalence of overweight and excessive abdominal fat in individuals with a lower BCAA intake (Figure 1). In addition, BCAA intake was negatively correlated with body fat (Figure 2).

D I S C U S S I O N
Amino acids can play an important role in the development of CVD [10,[28][29][30][31][32][33][34][35]. This relationship can be justified by the food source of these nutrients (legumes, oilseeds, eggs, fish, meat, milk, and dairy products). In fact, the cardiometabolic effects related to animal protein food sources are probably better explained by the non-protein components than by the protein components of these foods [36][37][38].  Data presented mean ± SD (standard deviation) or median and quartiles (p25-p75), when appropriate.  The first relevant result of this study was the higher body fat (%) in those individuals with a lower BCAA intake. Other results also were towards the inverse relationship between BCAA intake and body adiposity, as well as the results of other cross-sectional studies, carried out in Brazil, China, Japan, the United Kingdom and the United States [10,14,15,39]. Probably, leucine is the most influential BCAA regarding energy balance [40]. When these amino acids are released into the gastrointestinal tract after the hydrolysis of dietary proteins, the production of anorexigenic hormones is stimulated and thus the production of orexigenic hormone is inhibited. In addition, leucine induces the activation of the pro-opiomelanocortin, hypothalamic neuropeptide, as well as negatively regulating neuropeptide Y signaling.
In the central nervous system, these signals can affect food intake, increasing satiation and satiety. Also, leucine can act directly on adipocyte, liver, and muscle cells, influencing the expression https://doi.org/10.1590/1678-9865202033e190208 8 AP ALMEIDA et al.
of fatty acid synthase (adipocytes) and lipid catabolism (adipocytes, liver and muscle cells), favoring, subsequently, the reduction of adiposity [40]. Leucine is also a potent activator of mTOR, a serine/threonine kinase involved in many cellular processes, which includes protein synthesis and cell growth. Central leucine administration can increase hypothalamic mTOR signaling and decrease food intake and body weight [14]. Another possible mechanism for the effect of BCAA on body weight is the improvement of glucose tolerance, since impaired glucose tolerance may be related as one of the possible causes of obesity [15].
As shown in Table 2, individuals with a higher BCAA intake also had a higher intake of fibers and minerals (iron, copper, zinc, and magnesium) (p<0.05), as well as a lower intake of total fats (p<0.05). We also observed a negative correlation between the percentage of body fat and the current intake of fibers, iron, zinc, and copper (r=-0.16 and p=0.02; r=-0.206 and p=0.003; r=-0,16 and p=0.03; r=-0.16 and p=0.02, respectively). Therefore, the lower body fat in these individuals with a higher BCAA intake may also be related to a better quality of food in general.
Still, in the present study, serum ferritin and uric acid concentrations were significantly higher in the individuals with higher BCAA intake. However, these concentrations, even though higher, are still within normal values. They do not represent an increase in the cardiometabolic risk for these individuals [20,41,42].
In addition, HDL-c concentrations were lower in those individuals with higher BCAA intake. The opposite result was observed in the study by Cocate et al., in which HDL-c concentrations were higher in the third tertile, when compared to the first tertile of BCAA intake [10]. BCAA can act as signaling molecules to control energy homeostasis involving the disposition of glucose and lipid metabolism. Any changes in their intake could also lead to changes in the lipid profile [1,43], but additional studies that assess this relationship are necessary.
The present study has some limitations. First, its cross-sectional design makes impossible to infer about a cause-effect relationship in the results presented. However, the authors used the statistical tests that are suitable for this type of study and performed the interpretations of the results with scientific relevance. Another factor is the use of only a 24-hour recall, as it provides information only about the individual's current and unusual intake. However, this food survey has been used in epidemiological studies that evaluated the relation between food intake and cardiometabolic risk factors with promising results and acceptance by the scientific community [44][45][46][47].

C O N C L U S I O N
The results of the present study indicate that a higher BCAA intake (>2.6g/day) has an inverse association with excess weight and body fat, in addition to a positive association with biomarkers (ferritin and uric acid) in individuals at cardiometabolic risk. Further studies are needed to assess the relationship between BCAA intake and chronic diseases.

C O N T R I B U T O R S
AP ALMEIDA contributed to research design, obtaining data, analysis and interpretation of data, writing the article and critical review of the manuscript for important intellectual content. FS FORTES contributed to research design, obtaining data, analysis and interpretation of data and writing the article. BKS SILVEIRA and NA REIS contributed to research design and data collection. HHM HERMSDORFF contributed to research design, analysis and interpretation of data and critical review of the manuscript for important intellectual content.