Page 73 - Human Bile Acid Metabolism: a Postprandial Perspective

In order to counteract the insulin-desensitizing effects of prolonged fasting on glucose and lipid metabolism, the diet period was followed by three days on a eucaloric free diet [17]. Subjects were instructed to eat their estimated daily caloric need (estimated using the Harris Benedict equation).
Study design
The study was performed between May 2012 and February 2014 at the Department
of Endocrinology and Metabolism of the Academic Medical Center Amsterdam.
On study days, subjects were admitted at 07:30 h to the Metabolic Unit after an
overnight fast. A cannula was inserted into an antecubital vein for blood sampling. 4 This hand was kept in a heated hand box throughout the test to arterialize venous
blood. At 09:30 h, 3 blood samples were taken at 10-minute intervals for the determination of basal plasma glucose and insulin concentrations. At 10:00 h, subjects consumed a standard meal consisting of 50 g of parmesan cheese, 60 g of boiled egg and 75 g of glucose dissolved in 200 ml water (559 kcal; 30% from fat, 16% from protein, 54% from carbohydrates), after which blood samples were obtained at 0, 15, 30, 45, 60, 75, 90, 120, 150, 180 and 240 min after the meal. Blood was collected into chilled tubes containing either EDTA or Heparin as anticoagulant on ice and immediately centrifuged, and plasma was subsequently stored at 20 °C until analysis. For GLP-1 assays, a dipeptidyl peptidase inhibitor (Ile-Pro-Ile, Sigma-Aldrich, St. Louis, MO, USA) was added at 0.01 mg/ml and plasma was stored at 80 °C. Resting energy expenditure was measured for a 10 min-interval at baseline, 90 and 240 min after the meal by indirect calorimetry using a ventilated hood system (Vmax Encore 29; SensorMedics, Anaheim, CA). Energy expenditure was calculated as described by Frayn [18]. The abbreviated Weir equation was used to calculate 24-hour energy expenditure.
Laboratory analysis
Plasma glucose concentrations were analyzed bedside using the glucose oxidation method (EKF Diagnostics, Barleben/Magdeburg, Germany). Insulin was determined on an IMMULITE 2000 system (Siemens Healthcare Diagnostics, Breda, the Netherlands). GLP-1 concentrations were measured by ELISA using a commercially available assay (EMD Millipore, Billerica, MA, USA). BA were determined using a UPLC-tandem MS method to detect CA, CDCA, DCA and UDCA in their conjugated and unconjugated forms [19]. FGF19 was measured using an in-house developed ELISA as published previously [20].
Acute dietary weight loss
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