Postprandial Glucose Kinetics After Carbohydrate Restriction

This study will be a randomized, controlled, open-label, two-period crossover dietary intervention designed to compare the effects of a low-carbohydrate diet with those of an energy-matched Mediterranean diet on fasting and post-load glucose metabolism. The study will enroll adults with overweight or obesity and impaired glucose regulation who do not have diabetes.

The main objective will be to determine whether four weeks of carbohydrate restriction will modify the rate of appearance of orally ingested glucose, endogenous glucose production, and glucose clearance compared with a Mediterranean diet prescribed with a similar energy deficit. The study will also assess whether the two dietary interventions will have different effects on glucose tolerance, beta-cell function, insulin clearance, whole-body and tissue-specific insulin sensitivity, gastrointestinal hormones, non-glucose substrates, and the fasting lipid profile.

Participants will be randomly assigned in a 1:1 ratio to one of two intervention sequences. Participants assigned to the first sequence will receive the low-carbohydrate diet followed by the Mediterranean diet. Participants assigned to the second sequence will receive the Mediterranean diet followed by the low-carbohydrate diet. Each dietary intervention will last four weeks. The two intervention periods will be separated by a four-week washout period, during which participants will resume their habitual diet. Because of the nature of the dietary interventions, participants and study personnel responsible for dietary counseling will not be blinded to treatment allocation.

Both diets will be mildly hypocaloric and will provide an estimated energy restriction of approximately 200-400 kcal/day relative to basal metabolic requirements calculated using the Harris-Benedict equation. The low-carbohydrate diet will be designed to provide approximately 20% of total energy from carbohydrates, 55% from fats, and 25% from proteins. The Mediterranean diet will be designed to provide approximately 50% of total energy from carbohydrates, 30% from fats, and 20% from proteins.

Both dietary interventions will emphasize minimally processed foods, vegetables, fruits, legumes, fish, eggs, whole grains when compatible with the assigned macronutrient composition, and extra-virgin olive oil. Consumption of red and processed meat, ultra-processed foods, saturated fats, free sugars, alcohol, and salt will be limited. A registered dietitian will provide each participant with an individualized dietary plan specifying food choices, portion sizes, raw food weights, and possible substitutions with comparable energy and macronutrient content. The dietary plans will be adapted to individual food preferences when possible to facilitate adherence.

Participants will complete three-day quantitative food records during each week of both intervention periods. The records will include the types and quantities of foods and beverages consumed, condiments, and cooking methods. Dietary records will be analyzed using the Italian Food Composition Tables. Dietary adherence, body weight, and other anthropometric variables will be monitored during scheduled study visits. Adherence to the low-carbohydrate intervention will be considered inadequate when the estimated carbohydrate intake during that intervention is 40% or more of total daily energy intake.

At the end of each dietary intervention, participants will undergo a frequently sampled 75-g oral glucose tolerance test using two stable glucose isotope tracers. The metabolic assessments will be performed after a 12-hour overnight fast and after consumption of a standardized evening meal on the preceding day.

A primed continuous intravenous infusion of [6,6-²H₂]glucose will begin 120 minutes before ingestion of the oral glucose load and will continue throughout the test. At time zero, participants will ingest 75 g of glucose enriched with [U-¹³C]glucose within five minutes. Blood samples will be collected before and at predefined time points for 180 minutes after glucose ingestion.

Plasma tracer enrichment and glucose concentrations will be used to estimate the systemic rate of appearance of orally ingested glucose, endogenous glucose production, and glucose clearance. Fasting values, peak values, time to peak, and areas under the curve will be calculated, as appropriate. Glucose fluxes will also be examined during the early and later phases of the oral glucose tolerance test.

Plasma glucose, insulin, C-peptide, glucagon, total glucagon-like peptide-1, glucose-dependent insulinotropic polypeptide, non-esterified fatty acids, and beta-hydroxybutyrate will be measured during the test. Fasting glycated hemoglobin, lipid concentrations, and other routine biochemical variables will also be assessed.

Insulin secretion rates will be estimated from plasma C-peptide concentrations using deconvolution analysis. Mathematical modeling of glucose and insulin secretion will be used to quantify beta-cell glucose sensitivity, beta-cell rate sensitivity, insulin secretion at a fixed glucose concentration, and the potentiation of insulin secretion during the oral glucose tolerance test. Fasting and post-load insulin clearance will be estimated from the relationship between insulin secretion rates and plasma insulin concentrations.

Whole-body insulin sensitivity will be assessed using glucose clearance normalized to circulating insulin concentrations and by the Matsuda index. Hepatic insulin resistance will be estimated from the relationship between endogenous glucose production and insulin secretion. Adipose tissue insulin resistance will be estimated from fasting non-esterified fatty acid and insulin concentrations.

The two dietary interventions will be compared using paired statistical methods that account for the crossover design. Continuous variables will be evaluated for normality and will be compared using paired parametric or nonparametric tests, as appropriate. All statistical tests will be two-sided.