Measuring VLDL1-Triglyceride and VLDL2-Triglyceride Kinetics in Men: Effects of Dietary Control on Day-to-Day Variability.

The aim of this study was to assess the impact of dietary control on VLDL1- and VLDL2-TG kinetics and associated metabolic parameters. Twelve overweight/obese men were randomized to a provided 3 day isocaloric diet with fixed macronutrient composition (diet group, n=6) or to their regular unrestricted diet (non-diet group, n=6). VLDL1- and VLDL2-TG turnovers were measured twice 2-4 weeks apart, using primed-constant infusion of ex vivo labeled [1-14C]VLDL1-TG and [9,10-3H]VLDL2-TG. Isocaloric diet intervention lowered the difference in the mean of both VLDL2-TG secretion and clearance rate (p<0.01), and the coefficient of variation (CV) of VLDL2-TG clearance rate (p<0.05). The difference in mean and CV of the other kinetic estimates (VLDL1-TG secretion, clearance and oxidation rate) were lowered, but not significantly. The CV's of total triglyceride, VLDL1-TG, and VLDL2-TG concentrations were significantly lowered by diet intervention compared to regular diet; total triglyceride (31%-5%, p<0.01), VLDL1-TG (42%-9%, p<0.01), and VLDL2-TG (36%-10%, p<0.01). In conclusion, VLDL1- and VLDL2-TG kinetics show great day-to-day variability, which may be diminished by diet intervention. Therefore, standardized macronutrient intake prior to study days improves the probability of demonstrating significant outcomes of cross-sectional and intervention studies of VLDL1-TG and VLDL2-TG kinetics and metabolism.

Basal and insulin-regulated VLDL1 and VLDL2 kinetics in men with type 2 diabetes.

AIMS/HYPOTHESIS: Hypertriacylglycerolaemia is a hallmark of diabetic dyslipidaemia with increased concentrations of triacylglycerol (TG)-rich VLDL1 particles. However, whether VLDL1 secretion or removal is abnormal in type 2 diabetes remains unclear. The aim of this study was to compare basal and insulin-mediated VLDL1- and VLDL2-TG kinetics in men with type 2 diabetes and healthy men using a novel direct VLDL1- and VLDL2-TG labelling method. METHODS: Twelve men with type 2 diabetes and 12 healthy men matched for age and BMI were recruited. VLDL1- and VLDL2-TG turnover were measured during a 4 h basal period and a 3.5 h hyperinsulinaemic clamp period using a primed-constant infusion of ex vivo labelled VLDL1-TG and VLDL2-TG. RESULTS: Basal VLDL1-TG and VLDL2-TG secretion rates were similar in men with diabetes and healthy men. During hyperinsulinaemia, VLDL1-TG secretion rates were suppressed significantly in both groups, whereas no suppression of VLDL2-TG secretion rate was observed. VLDL1-TG to VLDL2-TG transfer rate was not significantly different from zero in both groups, while VLDL1-TG fatty acid oxidation rate was substantial, with a contribution to total energy expenditure of approximately 15% during postabsorptive conditions. VLDL1 and VLDL2 particle size (TG/apolipoprotein B [apoB] ratio) and apoB-100 concentration were unaltered by hyperinsulinaemia in men with type 2 diabetes, but significantly reduced in healthy men. CONCLUSIONS/INTERPRETATION: Insulin inhibits VLDL1-TG secretion rate similarly in age- and BMI-matched men with type 2 diabetes and healthy men, while VLDL2-TG secretion is unaltered by hyperinsulinaemia. However, VLDL1- and VLDL2-apoB levels are not lowered by hyperinsulinaemia in men with type 2 diabetes, which is indicative of a diminished hepatic response to insulin. TRIAL REGISTRATION: ClinicalTrials.gov NCT01564550.

VEGF-A mRNA measurement in meningiomas using a new simplified approach: branched DNA and chemiluminescence.

For decades, the preferred and almost sole method for measurement of gene expression has been RT-qPCR. The method is robust, inexpensive, and well-studied; however, PCR is also quite laborious and vulnerable to contamination. As part of an investigation of VEGF-A gene expression in meningiomas, an alternative and less laborious method for gene expression analysis based on branched DNA hybridization and chemiluminescence (Lumistar) was tested. Albeit the two methods differ, in principle, cellular mRNA-concentration is measured with both. Because they both determine gene expression via the measurement of mRNA-concentration, they were expected to be comparable. The aim of the present study was to compare Lumistar to the traditional RT-qPCR approach in a routine laboratory setting, where there is emphasis on rapid analysis response. Meningioma (n = 10) and control brain tissue (n = 5) samples were collected and VEGF-A and GAPDH mRNA were quantified using both RT-qPCR and Lumistar. Furthermore, two dilution series of two of the meningioma samples were prepared in order to make quantitative analyses. Both Lumistar and RT-qPCR-results were found to follow concentration dependent linear paths when diluted (p < 0.0001 and p < 0.01). Finally, Lumistar and RT-qPCR analyses were performed with the inclusion of a reference gene (GAPDH), where similar results were obtained with the two methods (R2 = 0.48; p = 0.01). It is intriguing that in spite of the vast difference in handling and assay principles, gene expression results are similar. The preferred method depends on the variability of the samples, budget, and time. Lumistar was less time consuming, while RT-qPCR was less expensive and best suited for data sets with large sample variability.

Short communication: Genetic variation in choice consistency for cows accessing automatic milking units.

Dairy cows milked in automatic milking systems (AMS) with more than 1 milking box may, as individuals, have a preference for specific milking boxes if allowed free choice. Estimates of quantitative genetic variation in behavioral traits of farmed animals have previously been reported, with estimates of heritability ranging widely. However, for the consistency of choice in dairy cows, almost no published estimates of heritability exist. The hypothesis for this study was that choice consistency is partly under additive genetic control and partly controlled by permanent environmental (animal) effects. The aims of this study were to obtain estimates of genetic and phenotypic parameters for choice consistency in dairy cows milked in AMS herds. Data were obtained from 5 commercial Danish herds (I-V) with 2 AMS milking boxes (A, B). Milking data were only from milkings where both the present and the previous milkings were coded as completed. This filter was used to fulfill a criterion of free-choice situation (713,772 milkings, 1,231 cows). The lactation was divided into 20 segments covering 15d each, from 5 to 305d in milk. Choice consistency scores were obtained as the fraction of milkings without change of box [i.e., 1.0 - µ(box change)] for each segment. Data were analyzed for one part of lactation at a time using a linear mixed model for first-parity cows alone and for all parities jointly. Choice consistency was found to be only weakly heritable (heritability=0.02 to 0.14) in first as well as in later parities, and having intermediate repeatability (repeatability coefficients=0.27 to 0.56). Heritability was especially low at early and late lactation states. These results indicate that consistency, which is itself an indication of repeated similar choices, is also repeatable as a trait observed over longer time periods. However, the genetic background seems to play a smaller role compared with that of the permanent animal effects, indicating that consistency could also be a learned behavior. We concluded that consistency in choices are quantifiable, but only under weak genetic control.

Isolated hyperglycaemia does not increase VLDL-triacylglycerol secretion in type 1 diabetic men.

AIMS/HYPOTHESIS: In type 1 diabetes, abnormalities of both glucose and lipoprotein metabolism are seen. The relationship between these factors is not understood, but studies indicate that hyperglycaemia may increase hepatic VLDL-triacylglycerol (VLDL-TG) secretion and reduce VLDL-TG fatty acid oxidation, which could lead to the development of dyslipidaemia. The aim of this study was to determine the isolated effect of hyperglycaemia on VLDL-TG and NEFA kinetics in men with type 1 diabetes. METHODS: VLDL-TG and palmitate kinetics were measured in eight men with type 1 diabetes using ex vivo labelled VLDL-TG and palmitate tracers. A 2.5 h basal period (plasma glucose 5 mmol/l) was followed by a 4 h hyperglycaemic period (plasma glucose 16 mmol/l). Steady-state VLDL-TG kinetics (VLDL-TG secretion, clearance and oxidation rates) were assessed by an isotope dilution technique using an intravenous primed-constant infusion of ex vivo labelled [1-(14)C]VLDL-TG in combination with sampling of blood and expired air. Palmitate turnover was measured using [9,10-(3)H]palmitate. RESULTS: The VLDL-TG secretion rate (36.0 ± 9.6 vs 30.8 ± 6.1 µmol/min, NS) and clearance rate (209 ± 30.4 vs 197 ± 41.7 ml/min, NS) were unchanged during the basal and hyperglycaemic periods, resulting in unchanged VLDL-TG concentrations (0.25 ± 0.11 µmol/l vs 0.28 ± 0.10 µmol/l, NS). In addition, VLDL-TG fatty acid oxidation and palmitate flux were not changed during hyperglycaemia. CONCLUSIONS/INTERPRETATION: Four hours of acute hyperglycaemia (16 mmol/l) without a concomitant increase in insulin does not alter VLDL-TG and NEFA kinetics in men with type 1 diabetes. CLINICAL TRIAL REGISTRY NUMBER: NCT01178957.

Clinical and Metabolic Characterization of Women With Gestational Diabetes Mellitus Within the First Year Postpartum.

Context: Women with gestational diabetes mellitus (GDM) have an increased risk of long-term complications, including impaired glucose metabolism, type 2 diabetes (T2DM), cardiovascular disease, and obesity. In current clinical practice, a 1 size fits all approach to GDM is applied, although heterogeneity among women with GDM has been recognized. Objective: To give the most adequate preventive care and postpartum (PP) guidance, we aimed to make a metabolic characterization and identify subgroups of women with previous GDM within the first year PP. Methods: In this prospective cohort study, we collected data in gestational week 34-38, at 3 months, and 1 year PP on women with GDM who participated in a PP follow-up program in Central Region Denmark from April 2019 to December 2022. Results: In total, 1270 women were included in the program in late pregnancy. Of the 768 women participating in either the oral glucose tolerance test 3 months PP (n = 545) or the 1-year follow-up (n = 493) or both (n = 261), 608 (79.2%) were normoglycemic, 137 (17.8%) had prediabetes, 20 (2.6%) had T2DM, and 3 (.4%) had developed T1DM. More than 40% of the women gained weight in the first year PP compared with their pregestational weight. Conclusion: Our study shows that 20.8% of women with GDM who volunteered to participate in a clinical follow-up program developed prediabetes or diabetes (T1DM and T2DM) within the first year PP. The GDM diagnosis encompasses a heterogenetic group of women and a deeper characterization may provide an opportunity for a more personalized risk assessment to prevent the progression to T2DM.

Lean body mass, not FFA, predicts VLDL-TG secretion rate in healthy men.

OBJECTIVE: Triglyceride is a risk factor for cardiovascular disease. However, the impact of body composition and free fatty acid (FFA) levels on very-low-density-lipoprotein triglyceride (VLDL-TG) secretion remains controversial. The aim was to identify predictors of VLDL-TG secretion in a data set compiled from seven previously published studies. METHODS: VLDL-TG kinetics was studied in 96 healthy men covering a wide span in body composition. A primed-constant infusion of ex vivo labeled [1-(14)C]-triolein VLDL-TG was used. Body composition was determined by dual X-ray absorptiometry and computed tomography scanning. Energy expenditure was measured by indirect calorimetry. Palmitate flux was measured by a [9,10-(3)H]-palmitate infusion. RESULTS: VLDL-TG secretion rate correlated significantly with body mass index (BMI), lean body mass (LBM), total fat mass, resting energy expenditure (REE), and insulin. A trend toward an inverse relationship between VLDL-TG secretion rate and FFA concentration was observed. In mixed model linear regression analysis, VLDL-TG secretion rate was positively associated with LBM (P = 0.03), and VLDL-TG clearance rate was inversely related to total fat mass (P < 0.01). CONCLUSIONS: LBM is a predictor of VLDL-TG secretion in healthy men, whereas FFA availability is not associated with VLDL-TG secretion. The work suggests reporting VLDL-TG secretion rates normalized for LBM when comparing subjects with differences in body composition.

Visceral obesity is associated with increased soluble CD163 concentration in men with type 2 diabetes mellitus.

Monocyte/macrophage-specific soluble CD163 (sCD163) concentration is associated with insulin resistance and increases with deteriorating glycemic control independently of BMI. This led to the proposal of the hypothesis that obesity-associated white adipose tissue inflammation varies between individuals. The objective was to examine the effect of male overweight/obesity and type 2 diabetes mellitus (T2DM) on associations between adiposity parameters and sCD163. A total of 23 overweight/obese non-diabetic men, 16 overweight/obese men with T2DM, and a control group of 20 normal-weight healthy men were included. Body composition and regional body fat distribution were determined by whole-body dual X-ray absorptiometry scan and abdominal computed tomography (CT) scan. Serum sCD163 concentrations were determined by ELISA. Associations between adiposity parameters and sCD163 were investigated using multiple linear regression analysis. In the normal-weight healthy men, there was no significant association between adiposity parameters and sCD163, whereas in the overweight/obese non-diabetic men, measures of general and regional adiposity were positively associated with sCD163. In the overweight/obese men with T2DM, only visceral adipose tissue (VAT) and the ratio of VAT to abdominal subcutaneous adipose tissue (SAT), a measure of relative body fat distribution between VAT and SAT depots, were positively associated with sCD163. In a multivariate analysis, including VAT, upper-body SAT, and lower-body fat, adjusted for BMI and age, VAT remained a significant predictor of sCD163 in the overweight/obese T2DM men, but not in the overweight/obese non-diabetic men. Our results indicate that VAT inflammation is exaggerated in men with T2DM, and that propensity to store excess body fat viscerally is particularly detrimental in men with T2DM.