Clinical and laboratory measurements

The questionnaires were completed by a trained nurse and the participants were requested to answer the questions carefully. Height and weight were measured according to a standard protocol by a team of trained nurses. Pre-pregnancy body mass index (BMI) was calculated using self-reported weight (kg) at 3 months before conception and divided by maternal height in meters squared (kg/m²). Brachial blood pressure was measured by trained nurses 3 times in the right arm after a 10-min rest period, using a manual mercury sphygmomanometer. A fasting blood sample was taken from all participants after 12 h fasting. Serum and plasma samples were separated from blood within 1 to 3 h following blood collection. Serum was separated by centrifugation at 500 × g for 10 min, and plasma was separated at 350 × g for 15 min at room temperature and then stored in −80 C until analysis. The concentration of FBS, urea and creatinine were measured by commercial kits (Pars Azmoon, Iran). Amino acid profiles were measured by HPLC method (Agilent Technologies).