Bariatric surgery improves postprandial VLDL kinetics and restores insulin-mediated regulation of hepatic VLDL production.

Dyslipidemia in obesity results from excessive production and impaired clearance of triglyceride-rich (TG-rich) lipoproteins, which are particularly pronounced in the postprandial state. Here, we investigated the impact of Roux-en-Y gastric bypass (RYGB) surgery on postprandial VLDL1 and VLDL2 apoB and TG kinetics and their relationship with insulin-responsiveness indices. Morbidly obese patients without diabetes who were scheduled for RYGB surgery (n = 24) underwent a lipoprotein kinetics study during a mixed-meal test and a hyperinsulinemic-euglycemic clamp study before the surgery and 1 year later. A physiologically based computational model was developed to investigate the impact of RYGB surgery and plasma insulin on postprandial VLDL kinetics. After the surgery, VLDL1 apoB and TG production rates were significantly decreased, whereas VLDL2 apoB and TG production rates remained unchanged. The TG catabolic rate was increased in both VLDL1 and VLDL2 fractions, but only the VLDL2 apoB catabolic rate tended to increase. Furthermore, postsurgery VLDL1 apoB and TG production rates, but not those of VLDL2, were positively correlated with insulin resistance. Insulin-mediated stimulation of peripheral lipoprotein lipolysis was also improved after the surgery. In summary, RYGB resulted in reduced hepatic VLDL1 production that correlated with reduced insulin resistance, elevated VLDL2 clearance, and improved insulin sensitivity in lipoprotein lipolysis pathways.

Endogenous Oxalate Production in Primary Hyperoxaluria Type 1 Patients.

BACKGROUND: Primary hyperoxaluria type 1 (PH1) is an inborn error of glyoxylate metabolism, characterized by increased endogenous oxalate production. The metabolic pathways underlying oxalate synthesis have not been fully elucidated, and upcoming therapies require more reliable outcome parameters than the currently used plasma oxalate levels and urinary oxalate excretion rates. We therefore developed a stable isotope infusion protocol to assess endogenous oxalate synthesis rate and the contribution of glycolate to both oxalate and glycine synthesis in vivo . METHODS: Eight healthy volunteers and eight patients with PH1 (stratified by pyridoxine responsiveness) underwent a combined primed continuous infusion of intravenous [1- 13 C]glycolate, [U- 13 C 2 ]oxalate, and, in a subgroup, [D 5 ]glycine. Isotopic enrichment of 13 C-labeled oxalate and glycolate were measured using a new gas chromatography-tandem mass spectrometry (GC-MS/MS) method. Stable isotope dilution and incorporation calculations quantified rates of appearance and synthetic rates, respectively. RESULTS: Total daily oxalate rates of appearance (mean [SD]) were 2.71 (0.54), 1.46 (0.23), and 0.79 (0.15) mmol/d in patients who were pyridoxine unresponsive, patients who were pyridoxine responsive, and controls, respectively ( P =0.002). Mean (SD) contribution of glycolate to oxalate production was 47.3% (12.8) in patients and 1.3% (0.7) in controls. Using the incorporation of [1- 13 C]glycolate tracer in glycine revealed significant conversion of glycolate into glycine in pyridoxine responsive, but not in patients with PH1 who were pyridoxine unresponsive. CONCLUSIONS: This stable isotope infusion protocol could evaluate efficacy of new therapies, investigate pyridoxine responsiveness, and serve as a tool to further explore glyoxylate metabolism in humans.

ANGPTL3 Inhibition With Evinacumab Results in Faster Clearance of IDL and LDL apoB in Patients With Homozygous Familial Hypercholesterolemia-Brief Report.

[Figure: see text].

Exploring the metabolic fate of medium-chain triglycerides in healthy individuals using a stable isotope tracer.

BACKGROUND & AIMS: Medium chain triglyceride (MCT) supplementation is often recommended as treatment for patients with long-chain fatty acid ß-oxidation (lcFAO) disorders, since they can be utilized as an energy source without the use of the defective enzyme. However, studies in mice and preterm infants suggest that not all medium-chain fatty acids (MCFA) are oxidized and may undergo elongation to long-chain fatty acids (LCFA). In this single blinded study, we explored the metabolic fates of MCT in healthy individuals using a 13C-labeled MCT tracer. METHOD: Three healthy males in rest received on two test days a primed continuous infusion of glyceryl tri[1,2,3,4-13C4]-octanoate with either an isocaloric supplementation of 1) exclusively MCT (MCT-only) or 2) a mixture of MCT, proteins and carbohydrates (MCT-mix). Gas chromatography - combustion - isotope ratio mass spectrometry (GC-C-IRMS) was used to determine 13C-enrichment of long-chain fatty acids in plasma and of 13CO2 in exhaled air. RESULTS: When provided as single energy source, an estimated 42% of administered MCT was converted to CO2. In combination with carbohydrates and proteins in the diet, oxidation of MCT was higher (62%). In both diets <1% of 13C-label was incorporated in LCFA in plasma, indicating that administered MCT underwent chain-elongation to LCT. CONCLUSIONS: Although the relative MCT oxidation rate was higher when combined with carbohydrates and protein, quantitatively more MCT was oxidized when given an isocaloric meal with solely MCT. As these results were obtained in the resting state opposed to during exercise, it is too early to give a recommendation concerning the use of MCT in lcFAO disorders. The data show that in resting healthy individuals only a very small part of the MCT is traced back as LCFA in plasma, suggesting that MCT treatment does not result in a large LCFA burden, however further research on storage of MCT in tissues is warranted. REGISTRATION: The study was registered in the Nederlands Trialregister. Protocol ID: Trial NL7417 (NTR7650).

Simultaneous assessment of intestinal permeability and lactase activity in human-milk-fed preterm infants by sugar absorption test: Clinical implementation and analytical method.

BACKGROUND & AIMS: Experimental (nutritional) interventions in preterm infants frequently focus on intestinal maturation, as improving tolerance to enteral nutrition is a major goal. Intestinal permeability and lactase activity serve as markers for intestinal maturation. We aimed to develop a protocol for the simultaneous assessment of both markers in human-milk-fed preterm infants by a sugar absorption test. In addition, we developed a new gas chromatography-mass spectrometry (GC-MS) method for the analysis of lactulose, lactose, and mannitol in urine and milk collected during the sugar absorption test. METHODS: The sugar absorption test was performed on days 4, 7, and 14 postpartum in 12 preterm infants (gestational age of 26-32 weeks). Human milk was collected, pooled, and divided into equal portions to provide a stable lactose intake for 24 h. Urine was collected in the last 6 h of this 24 h period, after administration of a bolus test sugar solution. Samples were analyzed by GC-MS after derivatization by oxime formation combined with acetylation. RESULTS: The GC-MS method was validated and used for the accurate measurement of lactulose, lactose, and mannitol concentrations. The urinary lactulose/mannitol ratio declined with time, suggesting a decreased intestinal permeability. The urine-to-milk-lactulose/lactose ratio increased as a result of increased lactase activity with time. CONCLUSIONS: The developed protocol for simultaneous assessment of intestinal permeability and lactase activity can be used to monitor the effect of experimental (nutritional) interventions in human-milk-fed preterm infants. Urine and milk samples obtained during the sugar absorption test can be accurately analyzed by GC-MS.

Development and validation of a gas chromatography-mass spectrometry method to analyze octanoate enrichments at low concentrations in human plasma.

A new method for accurately analyzing octanoate enrichment in plasma was developed and validated. Samples were derivatized directly in plasma by transesterification with isobutanol and were analyzed by gas chromatography-mass spectrometry (GC-MS). This method was developed to analyze the precursor enrichment in a stable isotope tracer protocol. Glyceryl tri[1,2,3,4-13C4] octanoate, a stable isotope-labeled medium-chain triglyceride (MCT), was orally administered in combination with (1) exclusively MCT or (2) a combination of protein, carbohydrates, and MCT to investigate the metabolic route of oral MCT under various conditions. Accurate analysis of octanoate enrichment in plasma at concentrations as low as 0.43 µM (lower limit of quantification, LLOQ) was performed. This is an improvement of about twenty times for the LLOQ for analysis of the enrichment of octanoate when compared with the gold-standard method for fatty acid analysis (methyl esterification). Moreover, we found that' with this gold-standard method, study samples were easily contaminated with (unlabeled) octanoate from other sources, leading to biased, incorrect results. The precision and linearity obtained using the new method were good (coefficient of variation intraday < 9.1%, interday < 9.3%, R2 of the calibration curve > 0.99). The sensitivity was sufficient for analyzing samples obtained using the stable isotope protocol. This new method is more sensitive than methyl esterification and it minimizes the risk of contamination. Graphical abstract.

The effect of dapagliflozin on apolipoprotein B and glucose fluxes in patients with type 2 diabetes and well-controlled plasma LDL cholesterol.

AIM: To dissect the effects of the sodium-glucose linked transporter 2 inhibitor dapagliflozin on lipid metabolism and assess whether these effects could potentially offset cardiovascular benefit with this drug-class. MATERIALS AND METHODS: We assessed the effect of dapagliflozin on lipid metabolism in 11 adults with uncomplicated type 2 diabetes. After 4 weeks of statin wash-out and 4 weeks of rosuvastatin 10 mg treatment, participants were treated with dapagliflozin 10 mg once-daily for 5 weeks. Before and after dapagliflozin, plasma lipids were measured and very low-density lipoprotein (VLDL)-1 and VLDL-2 apolipoprotein (Apo)B fluxes were assessed using (5.5.5-2 H3 )-leucine tracer infusion. In addition, hepatic and peripheral insulin sensitivity as well as insulin-mediated inhibition of peripheral lipolysis were measured during a two-step hyperinsulinemic-euglycaemic clamp using (6,6-2 H2 )-glucose and (1,1,2,3,3-2 H5 )-glycerol tracers. RESULTS: Rosuvastatin decreased all plasma lipids significantly: total cholesterol from 4.5 (3.2-6.2) to 3.1 (2.5-3.8) mmol/L, LDL cholesterol from 2.6 (1.7-3.4) to 1.5 (1.1-2.2) mmol/L, HDL cholesterol from 1.34 (0.80-2.02) to 1.19 (0.74-1.89) mmol/L and triglycerides from 0.92 (0.31-3.91) to 0.79 (0.32-2.10) mmol/L. The addition of dapaglifozin to rosuvastatin did not raise either LDL cholesterol or total cholesterol, and only increased HDL cholesterol by 0.08 (-0.03-0.13) mmol/L (P = 0.03). In line with this, dapagliflozin did not affect VLDL-1 or VLDL-2 ApoB fluxes. Fasting endogenous glucose production tended to increase by 0.9 (-3.4-3.1) µmol kg-1 min-1 (P = 0.06), but no effect on hepatic and peripheral insulin sensitivity or on peripheral lipolysis was observed. CONCLUSIONS: Dapagliflozin has no effect on plasma LDL-cholesterol levels or VLDL-apoB fluxes in the context of optimal lipid-lowering treatment, which will thus not limit cardiovascular benefit when lipids are adequately controlled.

Development and Validation of a New Gas Chromatography-Tandem Mass Spectrometry Method for the Measurement of Enrichment of Glyoxylate Metabolism Analytes in Hyperoxaluria Patients Using a Stable Isotope Procedure.

Primary hyperoxalurias (PH) are inborn errors of glyoxylate metabolism characterized by an increase in endogenous oxalate production. Oxalate overproduction may cause calcium-oxalate crystal formation leading to kidney stones, nephrocalcinosis, and ultimately kidney failure. Twenty-four hour urine oxalate excretion is an inaccurate measure for endogenous oxalate production in PH patients and not applicable in those with kidney failure. Treatment efficacy cannot be assessed with this measure during clinical trials. We describe the development and validation of a gas chromatography-tandem mass spectrometry method to analyze the samples obtained following a stable isotope infusion protocol of 13C2-oxalate and 1-13C-glycolate in both healthy individuals and PH patients. Isotopic enrichments of plasma oxalate, glycolate, and glyoxylate were measured on a gas chromatography-triple quadrupole mass spectrometry system using ethylhydroxylamine and N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide (MTBSTFA) for analyte derivatization. Method precision was good for oxalate and glycolate (coefficients of variation [CV] were <6.3% and <4.2% for inter- and intraday precision, respectively) and acceptable for glyoxylate (CV <18.3% and <6.7% for inter- and intraday precision, respectively). The enrichment curves were linear over the specified range. Sensitivity was sufficient to accurately analyze enrichments. This new method allowed calculation of kinetic features of these metabolites, thus enabling a detailed analysis of the various pathways involved in glyoxylate metabolism. The method will further enhance the investigation of the metabolic PH derangements, provides a tool to accurately assess the therapeutic efficacy of new promising therapeutic interventions for PH, and could serve as a clinical tool to improve personalized therapeutic strategies.

A modified low-protein infant formula supports adequate growth in healthy, term infants: a randomized, double-blind, equivalence trial.

BACKGROUND: A high protein intake in early life is associated with a risk of obesity later in life. The essential amino acid requirements of formula-fed infants have been reassessed recently, enabling a reduction in total protein content and thus in protein intake. OBJECTIVES: We aimed to assess the safety of an infant formula with a modified amino acid profile and a modified low-protein (mLP) content in healthy term-born infants. Outcomes were compared with a specifically designed control (CTRL) infant formula. METHODS: In this double-blind, randomized controlled equivalence trial, infants received either mLP (1.7 g protein/100 kcal; n = 90) or CTRL formula (2.1 g protein/100 kcal; n = 88) from enrollment (age ≤ 45 d) to 6 mo of age. A breastfed group served as a reference (n = 67). Anthropometry and body composition were determined at baseline, 17 wk (including safety blood parameters), and 6 mo of age. The primary outcome was daily weight gain from enrollment up until the age of 17 wk (at an equivalence margin of ±3.0 g/d). RESULTS: Weight gain from baseline (mean ± SD age: 31 ± 9 d) up to the age of 17 wk was equivalent between the mLP and CTRL formula groups (27.9 and 28.8 g/d, respectively; difference: -0.86 g/d; 90% CI: -2.36, 0.63 g/d). No differences in other growth parameters, body composition, or in adverse events were observed. Urea was significantly lower in the mLP formula group than in the CTRL formula group (-0.74 mmol/L; 95% CI: -0.97, -0.51 mmol/L; P < 0.001). Growth rates, fat mass, fat-free mass, and several essential amino acids were significantly higher in both formula groups than in the breastfed reference group. CONCLUSIONS: Feeding an infant formula with a modified amino acid profile and a lower protein content from an average age of 1 mo until the age of 6 mo is safe and supports an adequate growth, similar to that of infants consuming CTRL formula. This trial was registered at www.trialregister.nl as Trial NL4677.

The 1-13 C galactose breath test in GALT deficient patients distinguishes NBS detected variant patients but does not predict outcome in classical phenotypes.

Classical galactosemia (CG) patients frequently develop long-term complications despite early dietary treatment. The highly variable clinical outcome is poorly understood and a lack of prognostic biomarkers hampers individual prognostication and treatment. The aim of this study was to investigate the association between residual galactose oxidation capacity and clinical and biochemical outcomes in CG patients with varying geno- and phenotypes. The noninvasive 1-13 C galactose breath test was used to assess whole body galactose oxidation capacity. Participants received a 7 mg/kg oral dose of 1-13 C labelled galactose. The galactose oxidation capacity was determined by calculating the cumulative percentage dose of the administered galactose (CUMPCD) recovered as 13 CO2 in exhaled air. Forty-one CG patients (5-47 years) and four adult controls were included. The median galactose oxidation capacity after 120 minutes (CUMPCDT120) of 34 classical patients (0.29; 0.08-7.51) was significantly lower when compared to two homozygous p.Ser135Leu patients (9.44; 8.66-10.22), one heterozygous p.Ser135Leu patient 18.59, four NBS detected variant patients (13.79; 12.73-14.87) and four controls (9.29; 8.94-10.02). There was a clear correlation between Gal-1-P levels and CUMPCDT120 (P < .0005). In the classical patients, the differences in CUMPCDT120 were small and did not distinguish between patients with poor and normal clinical outcomes. The galactose breath test distinguished classical patients from homo- and heterozygous p.Ser135Leu and NBS detected variant patients, but was not able to predict clinical outcomes in classical patients. Future studies are warranted to enable individualised prognostication and treatment, especially in NBS variants with galactose oxidation capacities in the control range.

In vivo kinetic study of materno-fetal fatty acid transfer in obese and normal weight pregnant women.

KEY POINTS: Placental structure and function can be modified as a result of maternal obesity affecting materno-fetal fatty acids (FA) transport. We report for the first time, in humans and in vivo, the kinetics of placental FA transfer in normo-weight and in normolipemic obese pregnant women using stable isotopes. The administration of different tracer FA with similar behaviour to the mother at different time points allows the collection of kinetic information on materno-fetal transfer of FA despite only one sample of placenta and cord can be collected per subject. Computational modelling showed a good fit to the data when considering all maternal plasma lipid classes but not when based only on non-esterified FA. The novel approach using multiple tracer FA administration combined with computational modelling shows a consistent time course of placental tracer FA and predicted total FA accumulation. ABSTRACT: We analyse for the first time the in vivo materno-fetal kinetic transfer of fatty acids (FA) labelled with stable isotopes in control and obese (OB) pregnant women. Labelled FA with a similar metabolism (stearic acid: 13 C-SA; palmitic acid: 13 C-PA; oleic acid: 13 C-OA) were orally administered at -4 h, -8 h and -12 h, respectively prior to elective caesarean section to 10 pregnant women with a body mass index >30 (OB) and 10 with a body mass index in the range 20-25 (NW). Placenta, venous and arterial cord blood were collected obtaining a wide range of FA enrichments. A combined experimental and computational modelling analysis was applied. FA fractional synthesis rate (FSR) in placenta was 11-12% h-1 . No differences were observed between NW and normo-lipidemic OB. It was not possible to estimate FA FSR in cord blood with this oral bolus dose approach. Computational modelling demonstrated a good fit to the data when all maternal plasma lipid classes were included but not with modelling based only on the non-esterified FA fraction. The estimated materno-fetal 13 C-FA transfer was ∼1%. In conclusion, our approach using multiple 13 C-FA tracers allowed us to estimated FSR in placental/maternal plasma but not in fetal/maternal compartments. Computational modelling showed a consistent time course of placental 13 C-FA transfer and predicted total fetal FA accumulation during the experiment. We conclude that, in addition to non-esterified FA fraction in the maternal circulation, maternal plasma very low-density lipoprotein and other lipoproteins are important contributors to placental FA transfer to the fetus.

Branched-chain amino acid and branched-chain ketoacid ingestion increases muscle protein synthesis rates in vivo in older adults: a double-blind, randomized trial.

BACKGROUND: Protein ingestion increases muscle protein synthesis rates. However, limited data are currently available on the effects of branched-chain amino acid (BCAA) and branched-chain ketoacid (BCKA) ingestion on postprandial muscle protein synthesis rates. OBJECTIVE: The aim of this study was to compare the impact of ingesting 6 g BCAA, 6 g BCKA, and 30 g milk protein (MILK) on the postprandial rise in circulating amino acid concentrations and subsequent myofibrillar protein synthesis rates in older males. METHODS: In a parallel design, 45 older males (age: 71 ± 1 y; BMI: 25.4 ± 0.8 kg/m2) were randomly assigned to ingest a drink containing 6 g BCAA, 6 g BCKA, or 30 g MILK. Basal and postprandial myofibrillar protein synthesis rates were assessed by primed continuous l-[ring-13C6]phenylalanine infusions with the collection of blood samples and muscle biopsies. RESULTS: Plasma BCAA concentrations increased following test drink ingestion in all groups, with greater increases in the BCAA and MILK groups compared with the BCKA group (P < 0.05). Plasma BCKA concentrations increased following test drink ingestion in all groups, with greater increases in the BCKA group compared with the BCAA and MILK groups (P < 0.05). Ingestion of MILK, BCAA, and BCKA significantly increased early myofibrillar protein synthesis rates (0-2 h) above basal rates (from 0.020 ± 0.002%/h to 0.042 ± 0.004%/h, 0.022 ± 0.002%/h to 0.044 ± 0.004%/h, and 0.023 ± 0.003%/h to 0.044 ± 0.004%/h, respectively; P < 0.001), with no differences between groups (P > 0.05). Myofibrillar protein synthesis rates during the late postprandial phase (2-5 h) remained elevated in the MILK group (0.039 ± 0.004%/h; P < 0.001), but returned to baseline values following BCAA and BCKA ingestion (0.024 ± 0.005%/h and 0.024 ± 0.005%/h, respectively; P > 0.05). CONCLUSIONS: Ingestion of 6 g BCAA, 6 g BCKA, and 30 g MILK increases myofibrillar protein synthesis rates during the early postprandial phase (0-2 h) in vivo in healthy older males. The postprandial increase following the ingestion of 6 g BCAA and BCKA is short-lived, with higher myofibrillar protein synthesis rates only being maintained following the ingestion of an equivalent amount of intact milk protein. This trial was registered at Nederlands Trial Register (www.trialregister.nl) as NTR6047.

Myofibrillar and Mitochondrial Protein Synthesis Rates Do Not Differ in Young Men Following the Ingestion of Carbohydrate with Milk Protein, Whey, or Micellar Casein after Concurrent Resistance- and Endurance-Type Exercise.

BACKGROUND: Whey and micellar casein are high-quality dairy proteins that can stimulate postprandial muscle protein synthesis rates. How whey and casein compare with milk protein in their capacity to stimulate postprandial myofibrillar (MyoPS) and mitochondrial (MitoPS) protein synthesis rates during postexercise recovery is currently unknown. OBJECTIVE: The objective of this study was to compare postprandial MyoPS and MitoPS rates after protein-carbohydrate co-ingestion with milk protein, whey, or micellar casein during recovery from a single bout of concurrent resistance- and endurance-type exercise in young healthy men. METHODS: In a randomized, double-blind, parallel-group design, 48 healthy, young, recreationally active men (mean ± SEM age: 23 ± 0.3 y) received a primed continuous infusion of L-[ring-13C6]-phenylalanine and L-[ring-3,5-2H2]-tyrosine and ingested 45 g carbohydrate with 0 g protein (CHO), 20 g milk protein (MILK), 20 g whey protein (WHEY), or 20 g micellar casein protein (CASEIN) after a sequential bout of resistance- and endurance-type exercise (i.e., concurrent exercise). Blood and muscle biopsies were collected over 360 min during recovery from exercise to assess MyoPS and MitoPS rates and signaling through mammalian target of rapamycin complex 1 (mTORC1). RESULTS: Despite temporal differences in postprandial plasma leucine concentrations between treatments (P < 0.001), MyoPS rates over 360 min of recovery did not differ between treatments (CHO: 0.049% ± 0.003%/h; MILK: 0.059% ± 0.003%/h; WHEY: 0.054% ± 0.002%/h; CASEIN: 0.059% ± 0.005%/h; P = 0.11). When MILK, WHEY, and CASEIN were pooled into a single group (PROTEIN), protein co-ingestion resulted in greater MyoPS rates compared with CHO (PROTEIN: 0.057% ± 0.002%/h; CHO: 0.049% ± 0.003%/h; P = 0.04). MitoPS rates and signaling through the mTORC1 pathway were similar between treatments. CONCLUSION: MyoPS and MitoPS rates do not differ after co-ingestion of either milk protein, whey protein, or micellar casein protein with carbohydrate during recovery from a single bout of concurrent resistance- and endurance-type exercise in recreationally active young men. Co-ingestion of protein with carbohydrate results in greater MyoPS, but not MitoPS rates, when compared with the ingestion of carbohydrate only during recovery from concurrent exercise. This trial was registered at Nederlands Trial Register: NTR5098.

Myofibrillar and Mitochondrial Protein Synthesis Rates Do Not Differ in Young Men Following the Ingestion of Carbohydrate with Whey, Soy, or Leucine-Enriched Soy Protein after Concurrent Resistance- and Endurance-Type Exercise.

BACKGROUND: Protein ingestion during recovery from resistance-type exercise increases postexercise muscle protein synthesis rates. Whey protein has been reported to have greater anabolic properties than soy protein, an effect which may be attributed to the higher leucine content of whey. OBJECTIVE: The objective of this study was to compare postprandial myofibrillar (MyoPS) and mitochondrial (MitoPS) protein synthesis rates after ingestion of carbohydrate with whey, soy, or soy protein enriched with free leucine (to match the leucine content of whey) during recovery from a single bout of concurrent resistance- and endurance-type exercise in young healthy men. METHODS: In a randomized, double-blind, parallel-group design, 36 healthy young recreationally active men (mean ± SEM age: 23 ± 0.4 y) received a primed continuous infusion of l-[ring-13C6]-phenylalanine and l-[ring-3,5-2H2]-tyrosine and ingested 45 g carbohydrate with 20 g protein from whey (WHEY), soy (SOY), or leucine-enriched soy (SOY + LEU) after concurrent resistance- and endurance-type exercise. Blood and muscle biopsies were collected over a 360 min postexercise recovery period to assess MyoPS and MitoPS rates, and associated signaling through the mammalian target of rapamycin complex 1 (mTORC1). RESULTS: Postprandial peak plasma leucine concentrations were significantly higher in WHEY (mean ± SEM: 322 ± 10 µmol/L) and SOY + LEU (328 ± 14 µmol/L) compared with SOY (216 ± 6 µmol/L) (P < 0.05). Despite the apparent differences in plasma leucinemia, MyoPS (WHEY: 0.054 ± 0.002; SOY: 0.053 ± 0.004; SOY + LEU: 0.056 ± 0.004%·h-1; P = 0.83), and MitoPS (WHEY: 0.061 ± 0.004; SOY: 0.061 ± 0.006; SOY + LEU: 0.063 ± 0.004%·h-1; P = 0.96) rates over the entire 360 min recovery period did not differ between treatments. Similarly, signaling through mTORC1Ser2448, p70S6kThr389, 4E-BP1Thr37/46, and rpS6Ser235/236 was similar between treatments. CONCLUSION: Postexercise MyoPS and MitoPS rates do not differ after co-ingestion of carbohydrate with 20 g protein from whey, soy, or leucine-enriched soy protein during 360 min of recovery from concurrent resistance- and endurance-type exercise in young, recreationally active men. This trial was registered at Nederlands Trial Register as NTR5098.

Growth and Clinical Variables in Nitrogen-Restricted Piglets Fed an Adjusted Essential Amino Acid Mix: Effects of Free Amino Acid-Based Diets.

Background: Excess protein intake in early life has been linked to obesity and metabolic syndrome in later life. Yet protein, and in particular the essential amino acids (EAAs), need to be present in adequate quantity to support growth. Objective: With the use of a piglet model restricted in dietary amino acids (AAs), we compared the efficacy and safety of a standard formula with a low-AA formula containing an adjusted AA composition. Methods: Female piglets (3-7 d old; Landrace × Yorkshire × Duroc) were fed 1 of 4 isoenergetic AA-based formulas for 14 d (700 kJ · kg body weight-1 · d-1). The formulas contained a set control amount (44 g/L) and AA compositions referred to as the experimental standard (ST-100, n = 22), or 20% or 50% lower total AAs (respectively, ST-80, n = 19 and ST-50, n = 13), or 20% lower total AAs with an optimally adjusted EAA composition (O-80, n = 17). A series of clinical and paraclinical endpoints were measured. Results: Growth rates were similar for ST-100, O-80 and ST-80 piglets (all ∼15 g · kg-1 · d-1), whereas ST-50 had a markedly lower weight gain relative to all groups (all P < 0.05). Relative to ST-100, all groups with reduced AA intake showed ∼16% reduction in plasma albumin and ∼30% reduction in plasma urea (both P < 0.05). The absolute leucine oxidation rate was ∼30% lower for O-80 than for ST-100 piglets (P < 0.05). Conclusions: These data show that a 20% reduction in total AA intake for both the control (ST-80) and the adjusted AA (O-80) formula did not have any short-term adverse effects on growth in artificially reared, AA-restricted piglets. The lower absolute leucine oxidation rate observed in O-80 supports the development of an infant formula with an improved AA composition and a moderate reduction in total protein to support adequate growth in healthy infants.

Bioimpedance Spectroscopy Imprecisely Assesses Lean Body Mass in Pediatric Dialysis Patients.

OBJECTIVES: Alterations in body compositions are strongly associated with poor outcomes in end-stage renal disease patients. Hence, assessment of lean body mass is crucial for clinically monitoring these patients. The use of multifrequency bioimpedance spectroscopy measurements has also been advocated, but their usefulness in children is questioned. We investigated whether their application is appropriate for lean body mass measurement in pediatric patients receiving chronic dialysis. METHODS: Lean body mass estimates as assessed by multifrequency bioimpedance spectroscopy and by deuterium dilution were obtained for 15 patients (mean age 10.9 ±â€Š3.6 years). RESULTS: Lean body mass (mean ±â€Šstandard deviation) determined by bioimpedance was 24.2 ±â€Š10.7 and 24.4 ±â€Š10.3 kg by deuterium technique. Bland-Altman analysis showed a mean (±standard deviation) difference between the 2 methods of -0.25 ±â€Š2.30 kg with 95% limits of agreement of -4.80 to 4.25 kg. In a multiple linear regression model, the hydration status was associated with measurement bias after adjusting for age, sex, weight, and body surface area. CONCLUSIONS: Our results show a high level of agreement between measurements by bioimpedance and deuterium technique, but the limits of agreement were wide. These findings do not support the use of bioimpedance to individually assess lean body mass in pediatric dialysis patients with and without overhydration.

A new key player in VEGF-dependent angiogenesis in human hepatocellular carcinoma: dimethylarginine dimethylaminohydrolase 1.

BACKGROUND: Anti-angiogenic therapies, targeting VEGF, are a promising treatment for hepatocellular carcinoma (HCC). To enhance this potential therapy, identification of novel targets in this pathway is of major interest. Nitric oxide (NO) plays a crucial role in VEGF-dependent angiogenesis. NO production depends on arginine as substrate and asymmetric dimethylarginine (ADMA) as inhibitor. Dimethylarginine dimethylaminohydrolase 1 (DDAH-1) catabolizes ADMA and therefore regulates NO and VEGF expression. This study unravels additional mechanisms to improve VEGF targeting therapies. METHODS: The expression of DDAH-1 was examined in HCC specimen and non-tumorous background liver of 20 patients undergoing liver resection. Subsequently, arginine/ADMA balance, NO production, and VEGF expression were analyzed. The influence of hypoxia on DDAH-1 and angiogenesis promoting factors was evaluated in HepG2 cells and primary human hepatocytes. RESULTS: DDAH-1 expression was significantly induced in primary HCC tumors compared to non-tumorous background liver. This was associated with an increased arginine/ADMA ratio, higher NO formation, and higher VEGF expression in human HCC compared to non-tumorous liver. Hypoxia induced DDAH-1, iNOS, and VEGF expression in a time-dependent manner in HepG2 cells. CONCLUSIONS: Our results indicate that DDAH-1 expression is increased in human HCC, which is associated with an increase in the arginine/ADMA ratio and enhanced NO formation. Hypoxia may be an initiating factor for the increase in DDAH-1 expression. DDAH-1 expression is associated with promotion of angiogenesis stimulating factor VEGF. Together, our findings for the first time identified DDAH-1 as a key player in the regulation of angiogenesis in human HCC, and by understanding this mechanism, future therapeutic strategies targeting VEGF can be improved.

Bioimpedance and Fluid Status in Children and Adolescents Treated With Dialysis.

BACKGROUND: Assessment of hydration status in patients with chronic kidney failure treated by dialysis is crucial for clinical management decisions. Dilution techniques are considered the gold standard for measurement of body fluid volumes, but they are unfit for day-to-day care. Multifrequency bioimpedance has been shown to be of help in clinical practice in adults and its use in children and adolescents has been advocated. We investigated whether application of multifrequency bioimpedance is appropriate for total-body water (TBW) and extracellular water (ECW) measurement in children and adolescents on dialysis therapy. STUDY DESIGN: A study of diagnostic test accuracy. SETTING & PARTICIPANTS: 16 young dialysis patients (before a hemodialysis session or after peritoneal dialysis treatment) from the Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy, and the Emma Children's Hospital-Academic Medical Center, Amsterdam, the Netherlands. INDEX TEST: TBW and ECW volumes assessed by multifrequency bioimpedance. REFERENCE TESTS: TBW and ECW volumes measured by deuterium and bromide dilution, respectively. RESULTS: Mean TBW volumes determined by multifrequency bioimpedance and deuterium dilution were 19.2±8.7 (SD) and 19.3±8.3L, respectively; Bland-Altman analysis showed a mean bias between the 2 methods of -0.09 (95% limits of agreement, -2.1 to 1.9) L. Mean ECW volumes were 8.9±4.0 and 8.3±3.3L measured by multifrequency bioimpedance and bromide dilution, respectively; mean bias between the 2 ECW measurements was +0.6 (95% limits of agreement, -2.3 to 3.5). LIMITATIONS: Participants ingested the deuterated water at home without direct supervision by investigators, small number of patients, repeated measurements in individual patients were not performed. CONCLUSIONS: Multifrequency bioimpedance measurements were unbiased but imprecise in comparison to dilution techniques. We conclude that multifrequency bioimpedance measurements cannot precisely estimate TBW and ECW in children receiving dialysis.

Early micronutrient supplementation protects against early stress-induced cognitive impairments.

Early-life stress (ES) impairs cognition later in life. Because ES prevention is problematic, intervention is needed, yet the mechanisms that underlie ES remain largely unknown. So far, the role of early nutrition in brain programming has been largely ignored. Here, we demonstrate that essential 1-carbon metabolism-associated micronutrients (1-CMAMs; i.e., methionine and B vitamins) early in life are crucial in programming later cognition by ES. ES was induced in male C57Bl/6 mice from postnatal d (P)2-9. 1-CMAM levels were measured centrally and peripherally by using liquid chromatography-mass spectroscopy. Next, we supplemented the maternal diet with 1-CMAM only during the ES period and studied cognitive, neuroendocrine, neurogenic, transcriptional, and epigenetic changes in adult offspring. We demonstrate that ES specifically reduces methionine in offspring plasma and brain. Of note, dietary 1-CMAM enrichment during P2-9 restored methionine levels and rescued ES-induced adult cognitive impairments. Beneficial effects of this early dietary enrichment were associated with prevention of the ES-induced rise in corticosterone and adrenal gland hypertrophy did not involve changes in maternal care, hippocampal volume, neurogenesis, or global/Nr3c1-specific DNA methylation. In summary, nutrition is important in brain programming by ES. A short, early supplementation with essential micronutrients can already prevent lasting effects of ES. This concept opens new avenues for nutritional intervention.-Naninck, E. F. G., Oosterink, J. E., Yam, K.-Y., de Vries, L. P., Schierbeek, H., van Goudoever, J. B., Verkaik-Schakel, R.-N., Plantinga, J. A., Plosch, T., Lucassen, P. J., Korosi, A. Early micronutrient supplementation protects against early stress-induced cognitive impairments.