We employed a 12 week randomized controlled study design that compared the impact of the HPLC diet versus a LF diet in producing weight loss in severely obese adolescents. Both groups were informally prescribed an exercise program that included at least 30 minutes of daily moderately vigorous physical activity. Subjects were admitted to the Pediatric Clinical Research Center (CRC) for baseline testing and initiation of the diet, followed at 2 week intervals in the out-patient CRC clinic, and readmitted for 2 days at week 13 for final assessments. Subjects were re-contacted at 24 and 36 weeks after enrollment, and selected follow-up measurements were obtained. Between the end of the intervention period and these visits, no contact was made with the subjects to approximate a typical time limited weight loss intervention.
The study protocol was approved by the Colorado Multiple Institutional Review Board (COMIRB) and the Pediatric CRC. All participants provided assent and their parents or guardians gave informed consent. A local Data Safety Monitoring Board established through the CRC reviewed study data and progress at quarterly intervals.
Severely overweight adolescents between the ages of 12 and 18 were recruited through referral to a weight management clinic at The Children's Hospital, through word of mouth, and through advertisements in local newspapers. Inclusion criteria included primary obesity and a body weight estimated to be ≥ 175% of ideal body weight. Ideal body weight was defined as weight at the 50th percentile for a subject's height age (age at which height is 50th percentile) or weight at 50th percentile BMI for age. Exclusion criteria included current diagnosis of Type II Diabetes Mellitus; gall bladder, liver or renal disorders; known eating disorders; severe hypercholesterolemia (total cholesterol >300 mg/dl); endocrine disorders such as hypothyroidism or polycystic ovary syndrome; pregnancy; genetic disorder, such as Prader-Willi syndrome; mental retardation; severe depression; or use of any chronic medication that could impact appetite. Patients with poor family support that might have potentially precluded compliance with the study requirements were also excluded.
Subjects on the HPLC diet were instructed by the CRC bionutritionists to aim for a sustained very low carbohydrate intake (≤ 20 g/day) and for a concomitant high lean protein intake, which was estimated to provide 2.0-2.5 g protein/kg ideal body weight per day. Fat and energy intakes were not restricted; the only monitored restriction was carbohydrate intake. Subjects were instructed on appropriate food choices for the diet, and each subject was provided a diet education booklet, including a “food pyramid” tailored to the HPLC. Daily multivitamin-mineral and calcium supplements (500 mg/day elemental calcium) were recommended, as was a non-caloric fluid intake of ≥ 48 oz/day.
The low fat diet control group was instructed on a diet with a daily energy intake goal of 70% of resting energy expenditure estimated from the Harris-Benedict equation (12
), and with less than or equal to 30% of calories from fat. The subjects received a diet education booklet, based on the USDA Food Guide Pyramid. Multivitamin-mineral supplement and fluid recommendations similar to those for the HPLC group were also provided.
Both groups were encouraged to have at least 30 minutes/day of vigorous physical activity. Handouts were provided with ideas to encourage physical activity and an activity log was maintained. Quantitative data on physical activity were not collected.
Height and weight measurements were obtained with a wall mounted stadiometer and a 500 lb capacity scale, respectively. Measurements and age were entered into Epi Info, (Epi Info Version 3.5.1, Centers for Disease Control and Prevention, Atlanta, GA, 2008), through which BMI percentiles and Z-scores for age and sex were calculated.
Three-day diet records were scheduled at random times throughout the intervention period for a given subject. The number of days of records actually obtained varied among subjects, ranging from a minimum of 3 and up to 14 days, with an average of 8.1 days. Nutrient analyses were calculated by CRC bionutritionists using the Nutrient Data Systems for Research (NDS-R, V4.05, Minneapolis, MN: University of Minnesota).
At the time of the completion of diet records, subjects also recorded subjective feelings of hunger and fullness nine times throughout the day: before and after meals, and between meals. A 10 cm visual analog scale, with 0 being “not at all hungry” and 10 being “extremely hungry,” was used to rate hunger and satiety (13
Body composition measurements were obtained at baseline and at the 13 week in-patient visit by dual energy X-ray absorptiometry (DEXA; Lunar DPX-IQ, Madison, Wisconsin) on those subjects less than or equal to 136 kg, the machine's maximum capacity. All DEXA studies were performed by the Radiology department at The Children's Hospital.
Biochemical tests included fasting lipid profile, 2-hr oral glucose tolerance test, and β-hydroxybutyrate, which were analyzed through the CRC core laboratories. The homeostasis model assessment of insulin resistance (HOMA-IR) (14
) was calculated by dividing the product of the fasting insulin level (mU/mL) and glucose level (in mg/dL) by 402. The cut-off of 3.16 recommended for adolescents (15
) was applied to interpretation of the HOMA-IR data.
Studies undertaken for safety and potential adverse effects included serum electrolytes, blood urea nitrogen, creatinine, serum calcium, phosphorus, and magnesium; liver function tests (alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, gamma-glutamyl transpeptidase; and urine pregnancy test (β-HGG) for the female subjects. All surveillance laboratory tests were performed by the clinical laboratory at The Children's Hospital. At baseline, 6, and 13 weeks, an electrocardiogram (ECG) and a 24-hr Holter monitor were obtained and reviewed within 24 hr by a pediatric cardiologist at The Children's Hospital. Abdominal ultrasounds, performed by the Radiology department at The Children's Hospital, were obtained at baseline and 13 weeks.
A two-tailed, two-sample t-test with 0.05 Type I error was used to calculate sample size. Power calculations were based on an assumption that there would be a decrease of 6 kg in weight with the HPLC diet, and 1 kg with the LF diet. These assumptions were based upon the findings reported by Willi et al (5
). With an expected standard deviation of 4.5 kg, a sample size of 14 would provide 80% power to detect a 5 kg difference in weight loss between the groups. It was planned to randomize 50 subjects to allow for an expected drop-out rate of 40% by the end of the follow-up period.
Study data were entered into a secure, password protected Access (Microsoft Office, 2003) database constructed by the CRC bioinformatics core. Statistical analyses included two-sample t-tests to assess differences between the two dietary groups in baseline characteristics, 13 weeks, and the changes from 13 weeks to baseline, and average daily dietary intakes for continuous variables. Due to approximately half the subjects not completing the study, for a variety of reasons, the assumption of data missing at random could not be made. T-tests were therefore used to analyze the data rather than repeated measures or a mixed model approach. A chi-square test or Fisher exact test, if necessary, was used to determine differences between the two groups in categorical variables. For the variables total cholesterol (TC), triglycerides, and fasting insulin, the p-values were based on the logarithmic scale of the original values.