Study design and sample
The study was carried out in São Caetano do Sul, state of São Paulo, as part of the International Study of Childhood Obesity, Lifestyle and the Environment (ISCOLE) which includes 12 countries, with data from children between 9 and 11 years of age. Details of the ISCOLE study protocol have been previously published [13]. The municipality of São Caetano do Sul, located in the state of São Paulo, Brazil, has an area of 15.3 km2 and is located in a subtropical climate [10, 14]. The population of 10-year-old children in the municipality in 2013 was 1,557 (52.1% boys). The municipality has a service economy and the best Human Development Index in Brazil [14, 15].
The study protocol was introduced to school members and parents. After the respective permissions, it was applied in all selected schools. All children aged between 9 and 11 years old and in the 5th grade of elementary school were eligible to participate in the project. All schools were inserted in random order within a list and were selected according to a random draw in each stratum, considering a proportion of 80% (public) to 20% (private). Sixteen public and four private schools were selected in order to obtain a total sample of 500 children (50% of each sex) with 25–30 children from each school. Data collection was carried out between March 2012 and April 2013 and all assessments were carried out during a full week per school. Children were eligible to participate in the study if they: (a) were 9–11 years of age; (b) regularly enrolled in a school in the Sao Caetano do Sul system; and (c) did not have clinical or functional limitations preventing daily physical activity. Details of the sample size calculation and inclusion and exclusion criteria have been previously published [13, 14].
This study was conducted according to the guidelines laid down in the Declaration of Helsinki and all procedures involving human subjects/patients and must have been approved by the Pennington Biomedical Research Center Institutional Review Board and Ethics Committee of the Federal University of São Paulo (number: 332.529). Informed consent was obtained from parents/legal guardian(s) of the children.
A total sample of 584 (297 girls) children was invited to participate in the study and met the inclusion criteria [13, 14]. Participants with missing data (n = 176) were excluded. Thus, the final sample included 402 children (202 girls) with complete data. Details have been published elsewhere [13, 14].
Breastfeeding, parents’ body mass index and birth weight
A family health history questionnaire was completed by the parents or legal guardians of children between 9 and 11 years of age [13, 14]. The questionnaire contained information on breastfeeding, parental weight and height, and children’s birth weight. Breastfeeding (month) was assessed by the age at which the child completely stopped being breastfed and was analyzed continuously. Data on parental height and weight were collected via questionnaire and consequently the father’s and mother’s body mass index was calculated (kg/m2) and analyzed separately. Children’s birth weight (kg) was reported by parents or guardians and was analyzed continuously. Further details on the questionnaire can be found in a previous studies [13, 14].
Obesity indicators
The variables analyzed were: height, body weight, percentage of body fat (%BF) and waist circumference (WC) [13]. Height was measured to the nearest 0.1 cm using a portable Seca 213 stadiometer (Hamburg, Germany). Body weight and %BF were measured using a Tanita SC-240 scale, portable body composition analyzer (Arlington Heights, IL) after children removed heavy items from their pockets, shoes and socks [16]. Two measurements were obtained and the mean was used in the analysis (the third measurement was obtained if the first two measurements had a difference greater than 0.5 kg or 2.0% of distance for body mass). BMI was calculated from height and body weight (kg/m2), and subsequently converted to z-scores based on the World Health Organization (WHO) growth reference data. The nutritional status was classified as: underweight: <-2SD; eutrophic: -2 SD to 1 SD; overweight > + 1 SD to 2 SD; and obese: >+2 SD [17].
WC measurements were made on exposed skin at the end of normal expiration using a non-elastic anthropometric tape midway between the lower margin of the last rib and the iliac crest [13].
Sedentary time and moderate-to-vigorous physical activity
The Actigraph GT3X accelerometer (ActiGraph, Ft. Walton Beach, USA) was used to objectively monitor sedentary time and moderate-to-vigorous physical activity. The instrument was worn at the waist using an elastic belt, in the mid axillary line on the right side. Participants were encouraged to use the accelerometer 24 h/day for at least 7 days, including two weekend days. Children were instructed to remove the accelerometer only for water activities. To increase compliance, study staff instructed children on how to wear the accelerometer during the initial in-school assessment, and conducted an in-person compliance check 2–4 days after initialization. Further, participants were contacted twice during data collection via telephone (one weekday evening and one weekday) to ensure they were wearing the device, and to address any questions. Accelerometer monitoring procedures were identical to those described in previous studies [13, 18].
The minimum amount of accelerometer data that was considered acceptable for analysis was four days (including at least one weekend day), with at least 10 h/day of wear-time, after removal of sleep time [19, 20]. Blocks of 20 consecutive minutes of zero counts were considered as non-wear-time of the device and discarded from the analysis.
The investigation team verified that the data were complete using version 5.6 of the Actilife software (ActiGraph, Pensacola, FL). Data were collected at a sampling rate of 80 Hz, downloaded using 1 s epoch, which were subsequently aggregated for periods of 15 s [21]. The cut-points capture the sporadic nature of children’s activity and provide the best classification accuracy among the currently available cut-points for physical activity in children [20]. Specifically, sedentary time was defined as time accumulated at ≤ 25 activity counts/15 seconds, ≥ 26–573 activity counts/15 seconds for light physical activity, ≥ 574–1002 activity counts/15 seconds for moderate physical activity, ≥ 1003 activity counts/15 seconds for vigorous physical activity, and ≥ 574 activity counts/15 seconds for moderate-to-vigorous physical activity [21]. Time (min/day) spent sedentary, in light, and moderate-to-vigorous physical activity was calculated and used in analysis.
Sociodemographic variables
A family demographic questionnaire was completed by the parents or legal guardians. Full details of the questionnaires are provided elsewhere [13, 14]. Parents were asked about the children’s sex, age, and race/ethnicity (white/caucasian, black, mixed, or other). Total annual family income was used as an indicator at the household level and was categorized into four categories based on data distribution. These categories represent increasing levels of annual income (Brazilian currency), so that those with the lowest income were organized in the first category and those with the highest income in the last: <R$19,620 (level 1); 19,621 to 32,700 (level 2); 32,701 to 58,860 (level 3) and > R$58,861 (level 4). The mother’s educational level was divided into three categories: incomplete high school, complete high school/incomplete higher education and complete higher education.
Statistical analysis
The Kolmogorov-Smirnov test was used to assess data normality. Data is reported using means with 95% confidence interval (95%CI) and frequency as well as percentage (%) for categorical variables. For the comparison of categorical variables, we used the chi-square test and for continuous variables, we used the analysis of variance with one factor. Significant differences by nutritional status categories were analyzed by overlapping 95%CI, with a significant difference being considered when there was no overlap of the 95%CI; and no difference was considered when one of the 95%CI was partially included by the other [22].
Associations between variables were initially evaluated via Spearman correlation. Linear regression models were performed to estimate β-coefficient and 95%CI for the association between breastfeeding, parental BMI and birth weight (independent variables) with obesity indicators during late childhood (dependent variables). Separate models were determined for each independent variable adjusted for potential confounders mutually adjusted to each other. Model 1 was adjusted for school, sex, age, race/ethnicity and annual household income. Model 2 was adjusted for school, sex, age, race/ethnicity, annual household income, sedentary time and moderate-to-vigorous physical activity. The level of statistical significance was set as p < 0.05. Analyzes were performed using the Statistical Package for the Social Sciences V22 software (SPSS Inc., IBM Corp., Armonk, New York, NY, USA) [23].