The purpose of this study is to determine the impact of consuming cocoa on blood glucose levels, glucose metabolism, and other markers of pre-diabetes in overweight and/or obese individuals. Our hypothesis is that consumption of cocoa improves insulin sensitivity and glucose metabolism in subjects at risk for developing type-2 diabetes.
Study Type
INTERVENTIONAL
Allocation
RANDOMIZED
Purpose
PREVENTION
Masking
DOUBLE
Enrollment
16
Human Integrative Physiology Laboratory
Blacksburg, Virginia, United States
Change in insulin sensitivity
Insulin sensitivity will be determined using Bergman's minimal model (MINMOD Millennium software) via a frequently sampled intravenous glucose tolerance test (IVGTT). Fasting baseline blood samples will be taken prior to the dextrose injection (0.3 g/kg; 50% solution) at minute 0. Venous samples will be collected at minutes 1, 2, 3, 4, 5, 6, 7, 8, 10, 12, 14, 16, and 18. Insulin (0.025 U/kg) will be injected at minute 20. Venous sampling will continue at minutes 22, 23, 24, 25, 27, 30, 40, 50, 60, 70, 80, 90, 100, 120, 150, and 180. Glucose concentration will be immediately analyzed an automated glucose oxidase analyzer. Insulin will be later measured from serum using the Immulite 1000 immunoassay analyzer.
Time frame: Baseline and 4 weeks
Change in blood glucose response to a mixed meal
The test meal will be 2-8 oz. servings of a meal replacement beverage providing a mixed macronutrient profile, supplemented with either a cocoa beverage dry mix (10 g cocoa) or a calorie-matched placebo beverage mix (0 g cocoa). Blood samples will be taken at baseline and subsequently 1, 2, 3, and 4 hours after the first sip of the test beverage. Plasma glucose concentrations will be analyzed immediately using a glucose auto-analyzer (Yellow Springs Instruments).
Time frame: Baseline and 1 week
Change in hormone secretion response to a mixed meal
Serum insulin, GLP-1, GIP, and C-peptide concentrations will be determined using commercially available enzyme-linked immunosorbent assay (ELISA) kits from the blood samples drawn at baseline, 1, 2, 3, and 4 hours during the mixed meal challenge.
Time frame: Baseline and 1 week
Change in skeletal muscle metabolic flexibility
Subjects will consume a high fat meal containing 820 kcal; 52 g total carbohydrates, 24 g protein, and 58 g fat. During the high fat challenge session, two skeletal muscle biopsies will be conducted on alternate legs; one before the high fat meal and one 4 hours later. Biopsies of the vastus lateralis will be performed using a 5 mm modified Bergström needle. Collected tissue will be washed in 0.9% sterile saline to remove blood and tissue. Samples will be weighed and added to buffer and placed on ice for immediate analysis of substrate flexibility. For substrate flexibility, pyruvate oxidation will be used to assess the activity of pyruvate dehydrogenase. Calculated metabolic flexibility is expressed as a ratio of pyruvate oxidation to pyruvate oxidation + free fatty acids.
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Time frame: Baseline and 4 weeks
Change in blood endotoxin levels
During the high fat challenge (see Outcome 4), blood samples will be collected at baseline and 1, 2, 3, and 4 h after the first bite of the meal. Serum endotoxin will be measured in duplicate using Limulus Amebocyte Lysate Pyrogent ® 5000 assay kits.
Time frame: Baseline and 4 weeks
Change in gut permeability
The four-sugar \[40 g sucrose, 1 g mannitol, 1 g sucralose and 5 g lactulose\] probe test will be used to assess total gut permeability. After an overnight fast and urine evacuation, participants will be asked to consume the sugar-probe beverage within 5 minutes. They will be given two breakfast sandwiches to consume immediately. Participants will be instructed to collect all of their urine in a provided container from the time the beverage was consumed (0 h) until 5 h. A second urine collection container will be filled between 6-24 h. Twenty-four hours later, the volume of urine in each container will be measured and aliquots collected for later analysis. Gastroduodenal permeability is defined as sucrose/mannitol ratio (0-5 h). Small intestinal permeability is defined as the calculated lactulose/mannitol ratio for 0-5 samples. Colonic permeability is defined as both the 6-24 h lactulose/mannitol ratio and 6-24 h sucralose/mannitol ratio.
Time frame: Baseline and 4 weeks