In addition to the increased probability of developing obesity in adulthood, excessive body fat accrual during the developmental years is linked with earlier onset of chronic diseases (1) and, because of greater exposure, more adverse metabolic health outcomes (2–4). This trend is particularly salient in African American (AA) girls, who are disproportionately burdened by obesity as well as risk factors for the future progression to type 2 diabetes mellitus and cardiovascular diseases when compared with other racial/ethnic groups (5). Identification of strategies to limit excess adipose tissue acquisition and prevent metabolic perturbations early in the life course, particularly among AA girls, is essential.
Adipose tissue deposition/accumulation is influenced by multiple metabolic pathways in which nutrition plays a central role (2,6,7). Thus, dietary modification strategies are commonly undertaken to control weight and fat accumulation in both overweight/obese adults and children (6–10). Carbohydrate (CHO), the main dietary component affecting insulin action, affects the postprandial metabolic response contributing to weight and body composition changes. For example, in comparison to a low-CHO meal, the response to a meal high in CHO results in an augmented metabolic cascade of events (increased serum glucose and insulin, and subsequent increased lipogenesis and glycogenesis) (11). The relatively more rapid absorption of glucose associated with this type of meal challenges homeostatic mechanisms, complicating the transition from the postprandial to the postabsorptive state. For example, hypoglycemia is more likely to occur following consumption of a high-CHO meal, triggering counterregulatory hormone pathways, thereby influencing satiety cues (12). In addition, the proportion of CHO influences fuel oxidation and energy expenditure, reflecting a shift from fat to glucose oxidation and concomitant downregulation of energy expended at rest (12,13). Thus, high CHO intake can perturb metabolic pathways involved in fuel metabolism and may permit unfavorable body tissue compartmentalization and be conducive to impaired insulin homeostasis (14).
The response to a high-CHO diet may be further amplified in an individual with inherently high insulin secretion (eg, AA girls), particularly during sensitive times of growth and development (eg, puberty), further increasing the potential for excess fat deposition (15,16). We previously reported higher insulin secretion in AA girls and greater adipose tissue deposition (17). Accordingly, a dietary approach that minimizes the insulin-related altered/exaggerated cascade of events following meal ingestion, implemented during a critical period for adipose tissue deposition, may be beneficial in improving metabolic outcomes and facilitating weight loss. The present study was conducted in obese peripubertal AA girls to evaluate the efficacy of a moderately restricted versus a standard CHO diet on weight/fat loss and biochemical markers of insulin homeostasis and metabolic parameters.
Participants were 26 overweight/obese (ranging from 92nd body mass index [BMI] percentile and above) AA girls ages 9 to 14 years. Obesity was defined as >95th age-specific BMI percentile, with all but 1 girl (92nd percentile) falling in that range. The girls were pubertal stage II to V as assessed by a pediatrician according to the criteria of Marshall and Tanner (18,19). Exclusion criteria were medical diagnoses and/or taking medications known to affect body composition, metabolism, cardiac function, and the like. Girls were recruited to participate in a weight loss study through newspaper advertisement, distribution of flyers (posted at various community partnerships), and word of mouth. The nature, purpose, and possible risks of the study were carefully explained to the girls and parent(s), and informed assent and consent, respectively, were obtained. Full datasets were available on 26 of 36 girls recruited. Reasons for noncompletion included difficulty complying with diet (8), lack of transportation preventing food pickup (20), relocation, and inability to perform blood draw (20). The protocol was approved by the institutional review board for human subjects at the University of Alabama at Birmingham (UAB). All of the measurements were performed at the Participant and Clinical Interactions Resources (PCIR) and the Department of Nutrition Sciences at UAB between 2008 and 2009.
Participants were block randomized to 1 of 2 diets that they remained on for the duration of the study: reduced CHO (42% of energy from CHO; SPEC) or standard CHO (55% of energy from CHO; STAN). The 16-week intervention included 2 phases: a 5-week eucaloric (weight stable) phase and an 11-week hypocaloric (weight loss, approximately 1000-calorie reduction) phase. All food was provided for both phases of the study, with overall energy amount determined according to resting energy requirements (resting energy expenditure [REEs]; assessed via indirect calorimetry) multiplied by an activity factor of 1.2 (averaging 2010 kcal/day for eucaloric and 1025 kcal/day for hypocaloric). A physical activity questionnaire (PAQ-A (21)) was used to determine an appropriate activity factor. Based on reported low level of physical activity, an activity factor of 1.2 was used for all of the participants during each phase (22). At baseline, participants attended 2 visits. During the first visit, a physical examination by the study pediatrician was performed, questionnaires on typical diet and regular physical activity were given, and a whole-body dual-energy x-ray absorptiometry (DXA) scan was performed. During the second visit, participants in the fasted state reported to the PCIR at 06:30 for metabolic testing. After 30 minutes of rest, participants underwent indirect calorimetry and a liquid mixed meal test (LMMT). Following the LMMT, participants were provided meals according to prescribed diet and began the 5-week eucaloric phase. Throughout this phase, the energy prescription was evaluated and participants were weighed twice per week to ensure weight stability. Weight changes exceeding 2 kg from baseline resulted in caloric modification to maintain weight. At the midpoint of the eucaloric phase, a solid meal test using the assigned breakfast was given to determine the metabolic response to a meal on the prescribed diet. At the end of the 5-week eucaloric phase, participants repeated the DXA scan, indirect calorimetry, and LMMT. The liquid meal was administered to evaluate pancreatic response and changes over time to dietary modification, whereas the solid meal was used to evaluate the acute response to the actual meal. The LMMT and solid meal test were conducted only during the eucaloric phase. Given that high CHO intakes can alter metabolic pathways leading to impaired insulin and glucose profiles (16,23), it was important to evaluate the postprandial effects of the meal tests in the absence of weight loss to truly disentangle the effects of diet versus energy imbalance on body composition. During the 11-week hypocaloric phase, participants remained on the prescribed diet but received a 1000 kcal/day deficit. Participants were weighed twice per week to ensure compliance with the diet. Protocol design was such that a weight change reflecting >2 kg would result in energy modification, although this action was not necessary for any participant. Upon completion of the 11-week hypocaloric phase, an additional DXA scan was performed. Girls were encouraged to maintain their present level of physical activity for the duration of the study.
Assessment of Typical Diet and Physical Activity Patterns
A 4-day food record was used to assess typical dietary patterns, particularly breakfast consumption and meal patterning. The PAQ-A was used to obtain a measure of regular daily physical activity.
Table 1 outlines the nutritional composition of the SPEC diet and the STAN diet. The SPEC comprised 42% of energy from CHO and 40% of energy from fat, whereas the STAN comprised 55% of energy from CHO and 27% of energy from fat. Both diets contained ∼18% of energy from protein; however, both comprised <10% saturated fat. A moderate reduction was chosen to include an acceptable diet to the study population while maintaining a fat content <40% of the energy. Based on preliminary analyses and recent comparisons to National Health and Nutrition Examination Survey data (24), the reduction in CHO of the SPEC diet represented an approximate 25% decrement in amount of CHO consumed by this population. Although diets were designed to be similar in CHO (eg, fiber, fructose content) and fat quality (eg, cholesterol), in an effort to attain macronutrient profile while providing cultural- and age-appropriate meals, there were differences in fructose, fiber, and cholesterol (P < 0.01).
Participants were instructed to remain on the assigned diet throughout the duration of the 16-week intervention period: SPEC, specialized for the unique metabolic characteristics (high-insulin secretion/low-insulin sensitivity) of the sample population, or STAN, reflecting typical dietary composition among the same population. All of the meals were based on an energy level of 1600 kcal, and snacks in increments of 100 kcal were added to the menu as necessary for participants who required excess. All of the meals included culturally and age-appropriate foods (no supplements or formulas) and were prepared and packaged in the research kitchen of the UAB PCIR. Noncaloric fluid intake of ≥64 oz/day was recommended.
The same registered dietitian obtained anthropometric measurements on all of the participants. Participants were weighed (Scale-tronix 6702W; Scale-tronix, Carol Stream, IL) to the nearest 0.1 kg in minimal clothing without shoes. Height was also recorded without shoes using a digital stadiometer (Heightronic 235; Measurement Concepts, Snoqualmie, WA). BMI percentile was calculated using Centers for Disease Control and Prevention growth charts based on these measurements (http://apps.nccd.cdc.gov/dnpabmi).
The Tanner staging method has been demonstrated to be a reliable indicator of pubertal development. Direct observation for the assessment of pubertal stage by a pediatrician, the criterion standard for differentiating among the 5 stages of maturity (25,26), was used. The staging based on the criteria of Marshall and Tanner (19,27) is according to both breast and pubic hair development. One composite number is assigned for Tanner staging, representing the higher of the 2 values defined by breast and pubic hair development (28).
Liquid Meal Test/Insulin Secretion
To determine insulin response to a standardized meal, insulin sensitivity and secretion were derived via frequent blood sampling following ingestion of an LMMT (Carnation Instant Breakfast prepared with whole milk). To perform the test, a flexible intravenous catheter was placed in the antecubital space of the left arm. The dose for the liquid meal test was obtained according to amount of lean body mass (LBM) of the participant (1.75 g CHO/kg LBM) and provided a macronutrient distribution (24% fat, 58% CHO, 18% protein) comparable with the STAN diet. Subjects were required to consume the meal within 5 minutes. Blood was drawn at baseline (3 samples for 15 minutes) before the initiation of meal consumption at time 0. Subsequent blood samples were drawn every 5 minutes from time 0 to 30 minutes, every 10 minutes from time 40 to 180 minutes, and at 210 and 240 minutes. Using the obtained measures, the insulin sensitivity index (SI) was calculated by the oral glucose model (20).
Solid Meal Test
To examine the response of the subjects to an assigned diet meal, during week 3 of the eucaloric phase, the assigned diet breakfast was consumed at the PCIR. The breakfast meal was that which was assigned the day of the test and therefore was not exactly the same across participants; however, it was representative of the nutrient distribution of the assigned diet and provided a similar amount of energy (∼500). Metabolites were analyzed before, during, and after intake. To perform the test, a flexible intravenous catheter was placed in the antecubital space of the left arm. At time 0, subjects began meal consumption, completing within 20 minutes. Blood samples were collected at baseline, and at times 15, 60, 90, 120, 180, and 240 minutes. Samples were analyzed to determine whether, as predicted, SPEC would result in a more stable blood glucose:insulin profile (60-min insulin) than STAN.
Body Composition and Fat Distribution
Whole-body composition was measured by DXA using a GE Lunar Prodigy densitometer (GE LUNAR Radiation, Madison, WI). Participants were scanned in light clothing while lying flat on their backs with their arms at their sides. Because of size limitations, girls not fitting within the scanning box were right-sided hemi-scanned with the left side estimated as per instrument protocol.
Resting Energy Expenditure
REE was determined in the fasted state upon arrival in the morning of the outpatient visit (Deltatrac, Sensormedics Corp, Yorba Linda, CA) in the PCIR. The instrument was calibrated before each test against standard gases. Before beginning testing, girls rested in attempt to return to “basal” state. During testing, all of the subjects were instructed to lie still while remaining awake. A canopy hood was used to collect expired air for 20 minutes after a 10-minute equilibration period, and oxygen consumption and carbon dioxide production were measured continuously during this time. After completion, REE was calculated using the equation of Weir (29).
Assay of Metabolites
Glucose was measured in 12 μL sera with the glucose oxidase method using a SIRRUS analyzer (interassay CV 2.56%; Stanbio Laboratory, Boerne, TX). Insulin was analyzed using a Tosoh AIA 1800 Automated Immunoassay Analyzer (Tosoh Bioscience, South San Francisco, CA). Assay sensitivity is 15.42 pmol/L, mean intraassay coefficient of variability is 4.69%, and interassay coefficient of variability is 6.0%. Triglycerides (TGs) were assessed with the glycerylphosphate method. High-density lipoprotein (HDL)-cholesterol was analyzed using a 2-reagent system involving stabilization of low-density lipoprotein (LDL), very-low-density lipoprotein (VLDL), and chylomicrons using cyclodextrin and dextran sulfate, and subsequent enzymatic-colorometric detection of HDL. Adiponectin and leptin were measured by radioimmunoassay as described (30,31).
Based on the data from a similar population, linear mixed models (LMMs) using SAS PROC MIXED were fit to estimate the difference during the 4-month period while accounting for dependency and missing data. Age and BMI percentile at the start of the program were used as covariates. The standard errors for the difference were multiplied by the square root of their degrees of freedom to estimate the standard deviation of difference scores. For subsequent power analyses, the standard deviation of difference scores was assumed to remain constant over the k = 3 repeated measures of the proposed study. These results indicated that we would need at least N = 22 total subjects to have 80% statistical power at α = 0.01. Statistical significance was determined as an alpha ≥0.05.
Differences in descriptive statistics were examined using t tests. The contributions of diet to metabolic parameters, body composition, and weight outcomes were evaluated using ANOVA to allow for inclusion of covariates. All of the models were adjusted for pubertal stage and height, and for models evaluating HDL, TG concentration was added as a covariate.
To conform to the assumptions of linear regression, all of the statistical models were evaluated for residual normality and logarithmic transformations were performed when appropriate. All of the data were analyzed using SAS 9.1 software (SAS Institute, Cary, NC).
Table 2 illustrates baseline sample characteristics, metabolites, and body composition measurements of participants in the total sample and by diet group. Participants were similar across all of the variables except fat percent, which was statistically higher for leg fat and total percent fat (P < 0.05 for both) but no statistical difference in trunk fat (P > 0.14), among girls on STAN.
Figure 1 illustrates the insulin (A) and glucose (B) response to the test meal given during week 3. Girls on SPEC had a lower insulin (P < 0.05) concentration at 60 minutes following ingestion of the meal. The difference in insulin response remained significant for up to 3 hours after the meal challenge (P < 0.01, for all time points). Girls on the STAN diet had higher glucose levels at 60, 90, and 120 minutes (P < 0.01).
Table 3 illustrates metabolic parameters assessed at baseline and after 5 weeks on the eucaloric diets. Changes in metabolic parameters during the 5-week eucaloric phase by diet group, within diet group (time), and diet × time group interactions are presented. After 5 weeks a time effect was observed; girls on SPEC had higher fasting insulin (6.7 mg/dL increase; P = 0.02) relative to their baseline values, but the insulin concentration did not differ by diet group (P = 0.9). The elevated TG (12.9 mg/dL increase) on the STAN diet and concomitant decrease on SPEC (14.9 mg/dL) were such that by the end of 5 weeks the girls on the SPEC diet had significantly lower TG relative to the girls on the STAN diet (P < 0.001). Girls on both diets had decreased LDL (STAN = 9.7; SPEC = 4.8) and increased adiponectin levels (0.8 = STAN; 0.04 = SPEC) for 5 weeks, although the changes did not reach statistical significance (P = 0.07, 0.06, respectively). Because there were differences in fat and CHO quality between the diets, the glycemic index and load, fat, fiber, and cholesterol were included as independent variables to assess their contribution. However, significant contribution (P < 0.10) was not observed for any of these variables (data not shown).
Diet groups did not differ in amount of weight lost at the end of the intervention, although there was a trend toward greater weight loss on the SPEC diet relative to the STAN diet during the eucaloric phase (P = 0.09). This marginal difference in weight loss appeared to have been attributed to loss from the LBM compartment (P = 0.11; data not shown) as opposed to fat compartments. Girls on SPEC had less leg fat (P = 0.04) and marginally less percent fat (P = 0.06) at the end of the eucaloric phase relative to girls on STAN. A significant difference in fat compartments between the 2 diets was not detected at the end of the hypocaloric phase or at the end of the intervention, suggesting the plausibility of greater leg and percent fat loss on the STAN diet, although not detected as a significant difference. It is unclear if the greater loss is attributable to greater amount present before the intervention. Table 4 illustrates the change during time in weight and fat parameters from baseline. Although diet groups also did not differ in total or compartmental fat loss during the intervention, there were significant changes in weight and fat loss on both diets.
High-CHO diets have been associated with increased fat deposition (16,32,33). During the pubertal transition, the additional “insult” of transient insulin resistance may augment fat mass accrual, especially among AA girls, who in general are more insulin resistant and more apt to gain fat during this period (17,34). The physiological premise of the present study was that a dietary approach minimizing associated perturbations may improve long-term health outcomes. We hypothesized that a moderately reduced CHO diet would improve body composition outcomes through attenuation of the insulin response, thereby increasing the capacity to control weight/fat accumulation in this population. There were no group differences in weight loss or body composition changes at the end of the intervention. A differential response in terms of insulin/glucose homeostasis was observed between diet groups, because those in the reduced CHO group experienced lower insulin response at 60 through 120 minutes and better glucose maintenance during a meal challenge. Individuals receiving the reduced CHO diet had decreased TG concentrations to a greater degree than those who received a standard CHO diet. Our results suggest that dietary CHO reduction favorably influences lipid parameters but did not result in greater weight or fat loss relative to a standard diet in peripubertal AA girls.
Low-fat, high-CHO diets have been consistently shown to induce hypertriglyceridemia (35–37). Moderate CHO reduction resulted in decreased TG, whereas the standard CHO diet (55% CHO; 27% fat), admittedly not a classic “high-CHO diet” (eg, >75% CHO), resulted in marginally increased TG. Our findings suggest that even a modest reduction in CHO relative to “typical” intake has the capacity to reduce serum TG. Interestingly, both diets demonstrated a decrease in LDL and an increase in adiponectin (although not significant), suggestive of improvement of at least some aspects of metabolism. Both diets were designed to be of higher quality in terms of macronutrient profile than what has been reported to be practiced by a similar pediatric cohort in the same geographic region (38,39). We speculate that metabolic improvements in both groups are at least partially attributed to this as well as the implementation of a more regular and healthier eating pattern than previously practiced (eg, breakfast consumption and smaller, more frequent meal consumption). The contribution of macronutrient quality cannot be ignored, however. Indeed, CHO quality (eg, fiber, fructose) and fat quality (eg, cholesterol) affect de novo fat synthesis. The beneficial effect of the SPEC diet on lipid profile (although with higher dietary cholesterol) supports the adverse effect of fructose and/or beneficial effect of fiber on endogenous cholesterol synthesis (35–37). Future studies on the extent to which the varying dietary quality exerts effects on metabolic parameters, particularly during formative years, are warranted.
A significant dietary effect was not observed in weight loss or body composition; however, it is possible that there were baseline differences in body composition that may have introduced confounding. For example, the difference in trunk fat, although not statistically different between the groups, could have mechanistically influenced the metabolic ability to use substrates as well as mediate differences in metabolic and reproductive hormones (40). Although visceral adiposity was not assessed, waist circumference was not statistically different (P = 0.30) between the STAN (102.4 cm) and SPEC (96.2 cm) groups. We must be aware that a statistically significant difference is not the same as a difference of biological significance; that is, lack of a statistically significant group difference in fat depots may not be transferred to indifference in metabolic risk. Although the present pediatric obesity epidemic literature is scant, Randle et al (41) demonstrated that use of 1 nutrient inhibited the use of another directly and without hormonal mediation several decades ago. This remains an auspicious avenue for investigation.
Although composition of the diet is an important factor in the determination of glucose and fat oxidation, individuals prone to obesity have an overall lower fat oxidation rate as compared with nonobese individuals (40,42). Contributing to disturbances related to obesity is the degree of metabolic inflexibility likely existent in this population because of the demands of puberty (8). For example, in overweight 10-year-old girls in the throes of puberty, high-fat feeding induced lower thermogenesis and higher fat deposition than isocaloric low-fat feeding (43). The higher fat accompanying the reduced CHO diet may have perturbed metabolic parameters related to fat oxidation, which may supersede other beneficial effects related to fat loss. It is also possible that excess weight gain results in a degree of irreversible exacerbation of metabolic changes during the pubertal transition. Sunehag et al recently reported no effect of modifications in CHO intake on glucose and lipid metabolism in healthy nonobese prepubertal children (44), suggesting that alteration in fat oxidation may be the result of an already perturbed metabolic environment. Furthermore, increased fasting insulin experienced by those in the reduced CHO group may have diverted fatty acids toward storage versus oxidation as well as altering fatty acid use by muscle. Reducing CHO intake can rapidly decrease the body's glycogen content, which may be responsible for elicitation of compensatory response (change in tissue partitioning/fuel use/water storage) because of the body's inability to store excess CHO. This may explain the observed reduction in LBM during the eucaloric phase. Observed LBM loss also may have been reflective of fluid shift in response to altered fuel use (ie, decreased glycogen storage, resulting in decreased concomitant water storage). Interestingly, during the hypocaloric phase, we did not observe additional LBM reduction in individuals on SPEC, and LBM did not differ between groups by the end of the intervention. It is important to recognize, however, that at baseline, girls on STAN had significantly more total fat relative to girls on SPEC, raising the possibility that lipid metabolism of girls on STAN or SPEC may differ in use of glucose and fat oxidation and differences observed may be related to baseline fat mass. There is paucity in the literature centered on metabolic flexibility in the pediatric population. Indeed, obesity is associated with increased fatty acid concentration and poor suppression of lipolysis with increased insulin levels (17,23,40). Intriguingly, existing studies focus on glucose availability, with little attention paid to lipid or protein availability, irrespective of the inefficient switch to these substrates in the insulin-resistant state (40). Furthermore, a differential effect among AA girls has not been evaluated. Although such hypotheses are in need of further confirmation, our findings highlight the importance of longer-term tailored, rather than one-size-fits-all, dietary strategies for weight management.
A major strength of the present study was the robust measurement used to assess metabolism and physiology. Providing all food to participants provided additional strength to the study design of this intervention. Although we cannot ensure 100% compliance, biweekly weigh-ins helped increase adherence to the diet. Nevertheless, we were limited in this pilot study by small sample size and the inclusion of only 1 racial/ethnic group. Identification of metabolic changes following weight loss also would have been beneficial, because it would have been ideal to have no differences in fat mass between the 2 diet groups. When groups are randomized, however, there is also a risk that the groups may differ at baseline. Although substantial evidence exists reporting the effect of fat and CHO quality on lipid profile, the variance and modest sample size may have precluded our observance of a significant contribution. Future studies should include metabolic measures following the hypocaloric phase, a larger sample size, multiple races/ethnicities, as well as individuals with a wider range of body habitus. In addition, beyond gross macronutrient effects, the evaluation of quality of macronutrients represents an auspicious area of evaluation.
In summary, we observed comparable weight and fat loss in both diet groups and a significantly greater reduction in TG following the SPEC diet compared with the STAN diet during the 16-week intervention. The fact that weight and fat loss for both groups were significantly reduced after 16 weeks argues that caloric intake independent of macronutrient consumption can be beneficial for controlling excess weight and fat accumulation in a population at risk for adverse health consequences. Future research is needed to determine long-term effectiveness of CHO reduction (with an added emphasis on CHO quality) on weight/fat loss on obesity-related metabolic parameters.
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