Effect of sitagliptin on energy metabolism and brown adipose tissue in overweight individuals with prediabetes: a randomised placebo-controlled trial.

AIMS/HYPOTHESIS: The aim of this study was to evaluate the effect of sitagliptin on glucose tolerance, plasma lipids, energy expenditure and metabolism of brown adipose tissue (BAT), white adipose tissue (WAT) and skeletal muscle in overweight individuals with prediabetes (impaired glucose tolerance and/or impaired fasting glucose). METHODS: We performed a randomised, double-blinded, placebo-controlled trial in 30 overweight, Europid men (age 45.9 ± 6.2 years; BMI 28.8 ± 2.3 kg/m2) with prediabetes in the Leiden University Medical Center and the Alrijne Hospital between March 2015 and September 2016. Participants were initially randomly allocated to receive sitagliptin (100 mg/day) (n = 15) or placebo (n = 15) for 12 weeks, using a randomisation list that was set up by an unblinded pharmacist. All people involved in the study as well as participants were blinded to group assignment. Two participants withdrew from the study prior to completion (both in the sitagliptin group) and were subsequently replaced with two new participants that were allocated to the same treatment. Before and after treatment, fasting venous blood samples and skeletal muscle biopsies were obtained, OGTT was performed and body composition, resting energy expenditure and [18F] fluorodeoxyglucose ([18F]FDG) uptake by metabolic tissues were assessed. The primary study endpoint was the effect of sitagliptin on BAT volume and activity. RESULTS: One participant from the sitagliptin group was excluded from analysis, due to a distribution error, leaving 29 participants for further analysis. Sitagliptin, but not placebo, lowered glucose excursion (-40%; p < 0.003) during OGTT, accompanied by an improved insulinogenic index (+38%; p < 0.003) and oral disposition index (+44%; p < 0.003). In addition, sitagliptin lowered serum concentrations of triacylglycerol (-29%) and very large (-46%), large (-35%) and medium-sized (-24%) VLDL particles (all p < 0.05). Body weight, body composition and energy expenditure did not change. In skeletal muscle, sitagliptin increased mRNA expression of PGC1ß (also known as PPARGC1B) (+117%; p < 0.05), a main controller of mitochondrial oxidative energy metabolism. Although the primary endpoint of change in BAT volume and activity was not met, sitagliptin increased [18F] FDG uptake in subcutaneous WAT (sWAT; +53%; p < 0.05). Reported side effects were mild and transient and not necessarily related to the treatment. CONCLUSIONS/INTERPRETATION: Twelve weeks of sitagliptin in overweight, Europid men with prediabetes improves glucose tolerance and lipid metabolism, as related to increased [18F] FDG uptake by sWAT, rather than BAT, and upregulation of the mitochondrial gene PGC1ß in skeletal muscle. Studies on the effect of sitagliptin on preventing or delaying the progression of prediabetes into type 2 diabetes are warranted. TRIAL REGISTRATION: ClinicalTrials.gov NCT02294084. FUNDING: This study was funded by Merck Sharp & Dohme Corp, Dutch Heart Foundation, Dutch Diabetes Research Foundation, Ministry of Economic Affairs and the University of Granada.

Multiple effects of cold exposure on livers of male mice.

Cold exposure of mice is a common method to stimulate brown adipose tissue (BAT) activity and induce browning of white adipose tissue (WAT) that has beneficial effects on whole-body lipid metabolism, including reduced plasma triglyceride (TG) concentrations. The liver is a key regulatory organ in lipid metabolism as it can take up as well as oxidize fatty acids. The liver can also synthesize, store and secrete TGs in VLDL particles. The effects of cold exposure on murine hepatic lipid metabolism have not been addressed. Here, we report the effects of 24-h exposure to 4°C on parameters of hepatic lipid metabolism of male C57BL/6J mice. Cold exposure increased hepatic TG concentrations by 2-fold (P < 0.05) but reduced hepatic lipogenic gene expression. Hepatic expression of genes encoding proteins involved in cholesterol synthesis and uptake such as the LDL receptor (LDLR) was significantly increased upon cold exposure. Hepatic expression of Cyp7a1 encoding the rate-limiting enzyme in the classical bile acid (BA) synthesis pathway was increased by 4.3-fold (P < 0.05). Hepatic BA concentrations and fecal BA excretion were increased by 2.8- and 1.3-fold, respectively (P < 0.05 for both). VLDL-TG secretion was reduced by approximately 50% after 24 h of cold exposure (P < 0.05). In conclusion, cold exposure has various, likely intertwined effects on the liver that should be taken into account when studying the effects of cold exposure on whole-body metabolism.

Phenylalanine requirements of enterally fed term and preterm neonates.

BACKGROUND: Phenylalanine, which is an essential aromatic amino acid, is either used for protein synthesis or irreversibly hydroxylated to tyrosine. The provision of optimal amounts of dietary phenylalanine is not only important for growth and development but might also influence catecholamine synthesis and release rates. The current recommended aromatic amino acid requirement for infants aged 0-6 mo is based on the amino acid content of human milk. OBJECTIVE: We quantified the requirements for phenylalanine in the presence of excess tyrosine (166 or 177 mg/kg per day for term and preterm infants, respectively) for term and preterm neonates by using the indicator amino acid oxidation method with l-[1-(13)C]lysine 2HCl as an indicator. Hence, we determined the minimum obligatory phenylalanine requirement. DESIGN: Fully enterally fed term and preterm infants received randomly graded amounts of phenylalanine (5-177 mg/kg per day) as part of an elemental formula. Data are expressed as means ± SDs. RESULTS: Twenty term (birth weight: 3.19 ± 0.34 kg; gestational age: 38.9 ± 1 wk) and 16 preterm (birth weight: 1.75 ± 0.17 kg; gestational age: 32.5 ± 0.6 wk) Asian infants participated at a postnatal age of 17 ± 8 d. In total, 44 studies were performed. The minimum obligatory phenylalanine requirement was 58 mg/kg per day (95% CI: 38-78 mg/kg per day) and 80 mg/kg per day (95% CI: 40-119 mg/kg per day) for term and preterm infants, respectively. CONCLUSION: The determined mean phenylalanine-requirement estimates are lower than the contents of term and preterm formulas currently on the market. This trial was registered at www.trialregister.nl as NTR1610.

Threonine Requirement of the Enterally Fed Term Infant in the First Month of Life.

OBJECTIVE: Threonine is one of the essential amino acids. Its major fate is incorporation into intestinal mucosal proteins and synthesis of secretory glycoproteins. Therefore, it has an important function in the neonatal gut barrier integrity. The objective was to quantify the threonine requirement in fully enterally fed term neonates by means of the indicator amino acid oxidation (IAAO) method, using L-[1-C]phenylalanine as indicator. METHODS: After a 24-hour test diet adaptation, containing randomly assigned amounts of threonine (range 5-182 mg · kg · day), the participating neonates received a primed continuous infusion of [C]bicarbonate and L-[1-C]phenylalanine. At baseline and during the plateau phase of both infusions, breath samples were obtained for CO2. The fractional L-[1-C]phenylalanine oxidation (FCO2) was estimated and plotted against the threonine intakes. Biphasic linear regression crossover analysis was used to calculate the breakpoint of the FCO2, representing the mean threonine requirement. Data are presented as mean ±â€ŠSD. RESULTS: Thirty-two term neonates (gestational age 39 ±â€Š1 weeks, birth weight 3.3 ±â€Š0.3 kg, mean postnatal age 10 ±â€Š4 days) were studied. The mean threonine requirement was estimated to be 68 mg · kg · day with an upper and lower 95% confidence interval of 104 and 32 mg · kg · day, respectively (r = 0.37). CONCLUSIONS: The determined threonine requirement is extremely close to the existing requirement recommendations (∼90% of the present World Health Organization requirement guidelines). Infant formula preparations presently on the market, however, contain up to twice as much threonine as recommended. The threonine intake in formula-fed infants may therefore be reduced considerably.

Tryptophan requirement of the enterally fed term infant in the first month of life.

OBJECTIVES: Tryptophan not only is an amino acid essential to protein synthesis but also serves as a precursor in 2 important metabolic pathways: the serotonin and the kynurenine pathways. Tryptophan is related to sleeping patterns. The objective of the present study was to determine the tryptophan requirement of term infants using the indicator amino acid oxidation (IAAO) method with L-[1-C]phenylalanine as the indicator. METHODS: Enterally fed infants were randomly assigned to tryptophan intakes ranging from 0.5 to 73 mg ·â€Škg ·â€Šday as part of an elemental diet. After 1-day adaptation to the test diet, [C]bicarbonate and L-[1-C]phenylalanine tracers were given enterally. Breath samples were collected at baseline and during isotopic plateaus. The mean tryptophan requirement was determined by using the biphasic linear regression crossover analysis on the fraction of CO2 recovery from L-[1-C]phenylalanine oxidation (FCO2). Data are presented as mean ±â€Šstandard deviation. RESULTS: A total of 30 term neonates (gestational age 39 ±â€Š1 weeks) were studied at 9 ±â€Š4 days. FCO2 decreased until a tryptophan intake of 15 mg ·â€Škg ·â€Šday; additional increases in tryptophan intake did not affect FCO2. Mean requirement was determined to be 15 mg ·â€Škg ·â€Šday. CONCLUSIONS: The mean tryptophan requirement for elemental formula-fed term infants is 15 mg ·â€Škg ·â€Šday. This requirement is lower than the present recommended intake of 29 mg ·â€Škg ·â€Šday, which is based on the average intake of a breastfed infant.

Branched-chain amino acid requirements for enterally fed term neonates in the first month of life.

BACKGROUND: Knowledge of essential amino acid requirements in infants is important because excessive intake of protein can lead to increased long-term morbidity such as obesity. A deficient intake may lead to suboptimal growth and impaired neurodevelopment. The current recommended branched-chain amino acid requirements in infants aged 0-1 mo are based on the amino acid content of human milk. OBJECTIVE: We quantified the requirements for isoleucine, leucine, and valine for term neonates by using the indicator amino acid oxidation method with [1-(13)C]phenylalanine as the indicator. DESIGN: Fully enterally fed term infants received randomly graded amounts of isoleucine (5-216 mg · kg(-1) · d(-1)), leucine (5-370 mg · kg(-1) · d(-1)), or valine (5-236 mg · kg(-1) · d(-1)) as part of an elemental formula. Data are expressed as means ± SDs. RESULTS: Eighty-three Asian, term neonates (mean ± SD birth weight: 3.3 ± 0.4 kg; gestational age: 39.4 ± 1.3 wk) were studied at a postnatal age of 13 ± 5 d. Mean requirements for isoleucine, leucine, and valine (measured in boys only) were 105 mg · kg(-1) · d(-1) (r(2) = 0.61, P < 0.001), 140 mg · kg(-1) · d(-1) (r(2) = 0.26, P < 0.01), and 110 mg · kg(-1) · d(-1) (r(2) = 0.35, P = 0.001), respectively. CONCLUSIONS: Current human milk-based recommendations for isoleucine and valine in term infants aged 0-1 mo are correct. However, the current recommendation for leucine (166 mg · kg(-1) · d(-1)) is higher than the mean requirement of 140 mg · kg(-1) · d(-1) that we determined in this study. This trial was registered at www.trialregister.nl as NTR1610.

New insights into the methodological issues of the indicator amino acid oxidation method in preterm neonates.

BACKGROUND: We determined the effect of adaptation to the study diet on oxidation of the indicator amino acid and the required tracer washout time in preterms. METHODS: Subjects received a study diet for 6 d that entailed a 50% reduction in leucine. Tracer studies using enterally infused [(13)C]bicarbonate and [1-(13)C]phenylalanine were performed on days 1, 2, 4, and 6. Breath samples containing (13)CO2 were collected during steady state and measured by infrared spectrometric analysis, and the fraction of (13)CO2 recovery from (13)C oxidation (F(13)CO2) was calculated. RESULTS: Preterm infants (n = 11, birth weight 1.9 ± 0.1 kg, gestational age 32.6 ± 1.5 wk) received 166 mg/kg/d of leucine. Baseline enrichment changed significantly at day 1 of the study diet. F(13)CO2 did not change significantly between days 2 and 4 but was significantly lower at day 6. The tracer washout time was determined to be 7.5 h using a biphasic regression analysis. CONCLUSION: One day of adaptation to a new diet is necessary to adapt to the (13)C enrichment of the study formula before starting infant requirement studies. Adaptation for a period of 5 d results in a protein-sparing response. The minimal time between two studies within the same subject is 7.5 h.

Methionine requirement of the enterally fed term infant in the first month of life in the presence of cysteine.

BACKGROUND: The essential amino acid methionine can be used for protein synthesis but also serves as a precursor for homocysteine and cysteine. OBJECTIVE: The objective of this study was to determine the minimal obligatory methionine requirement of infants in the presence of excess cysteine (91 mg ⋅ kg(-1) ⋅ d(-1)) by using the indicator amino acid oxidation (IAAO) method with l-[1-(13)C]phenylalanine as the indicator. DESIGN: Fully enterally fed term infants <1 mo of age were randomly assigned to methionine intakes that ranged from 3 to 59 mg ⋅ kg(-1) ⋅ d(-1) as part of an elemental formula. After 1 d of adaptation to the test diet, [(13)C]bicarbonate and l-[1-(13)C]phenylalanine tracers were given enterally. Breath samples were collected at baseline and during isotopic plateaus. The mean methionine requirement was determined by using biphasic linear regression crossover analysis on the fraction of (13)CO(2) recovery from l-[1-(13)C]phenylalanine oxidation (F(13)CO(2)). Data are presented as means ± SDs. RESULTS: Thirty-three neonates (gestational age: 39 ± 1 wk) were studied at 13 ± 6 d. With increasing methionine intakes, F(13)CO(2) decreased until a methionine intake of 38 mg ⋅ kg(-1) ⋅ d(-1); additional increases in methionine intake did not affect F(13)CO(2). The mean methionine requirement was determined at 38 mg ⋅ kg(-1) ⋅ d(-1), and the upper and lower CIs were 48 and 27 mg ⋅ kg(-1) ⋅ d(-11), respectively (P < 0.0001, r(2) = 0.59). CONCLUSIONS: Although the current recommended methionine intake of 28 mg ⋅ kg(-1) ⋅ d(-1) is within the CIs of our study, the estimated mean requirement is substantially higher. However, most of the infant formulas provide a methionine intake of 49-80 mg ⋅ kg(-1) ⋅ d(-1), which is above the upper CI of our study. This trial was registered at www.trialregister.nl as NTR1610.

Lysine requirement of the enterally fed term infant in the first month of life.

BACKGROUND: Infant nutrition has a major impact on child growth and functional development. Low and high intakes of protein or amino acids could have a detrimental effect. OBJECTIVE: The objective of the study was to determine the lysine requirement of enterally fed term neonates by using the indicator amino acid oxidation (IAAO) method. L-[1-(13)C]phenylalanine was used as an indicator amino acid. DESIGN: Twenty-one neonates were randomly assigned to lysine intakes that ranged from 15 to 240 mg · kg(-1) · d(-1). Breath, urine, and blood samples were collected at baseline and during the plateau. The mean lysine requirement was determined by using biphasic linear regression crossover analysis on the fraction of (13)CO(2) recovery from L-[1-(13)C]phenylalanine oxidation (F(13)CO(2)) and phenylalanine oxidation rates calculated from the L-[1-(13)C]phenylalanine enrichment of urine and plasma. RESULTS: The mean (±SD) phenylalanine flux calculated from urine and plasma L-[1-(13)C]phenylalanine enrichment data were 88.3 ± 6.9 and 84.5 ± 7.4 µmol · kg(-1) · h(-1), respectively. Graded intakes of lysine had no effect on phenylalanine fluxes. The mean lysine requirement determined by F(13)CO(2) was 130 mg · kg(-1) · d(-1) (upper and lower CIs: 183.7 and 76.3 mg · kg(-1) · d(-1), respectively). The mean requirement was identical to the requirement determined by using phenylalanine oxidation rates in urine and plasma. CONCLUSIONS: The mean lysine requirement of enterally fed term neonates was determined by using F(13)CO(2) and phenylalanine oxidation rates calculated from the L-[1-(13)C]phenylalanine enrichment of urine and plasma. These methods yielded a similar result of 130 mg lysine · kg(-1) · d(-1). This study demonstrates that sampling of (13)CO(2) in expired air is sufficient to estimate the lysine requirement by using the IAAO method in infants. This trial was registered at www.trialregister.nl as NTR1610.