Branched-Chain Amino Acid Oxidation Is Elevated in Adults with Morbid Obesity and Decreases Significantly after Sleeve Gastrectomy.

BACKGROUND: Plasma concentrations of branched-chain amino acids (BCAAs) are elevated in obese individuals with insulin resistance (IR) and decrease after bariatric surgery. However, the metabolic mechanisms are unclear. OBJECTIVES: Our objectives are to compare leucine kinetics between morbidly obese and healthy-weight individuals cross-sectionally, and to prospectively evaluate changes in the morbidly obese after sleeve gastrectomy. We hypothesized that leucine oxidation is slower in obese individuals and increases after surgery. METHODS: Ten morbidly obese [BMI (in kg/m2) ≥32.5, age 21-50 y] and 10 healthy-weight participants (BMI <25), matched for age (median ∼30 y) but not gender, were infused with [U-13C6] leucine and [2H5] glycerol to quantify leucine and glycerol kinetics. Morbidly obese participants were studied again 6 mo postsurgery. Primary outcomes were kinetic parameters related to BCAA metabolism. Data were analyzed by nonparametric methods and presented as median (IQR). RESULTS: Participants with obesity had IR with an HOMA-IR (4.89; 4.36-8.76) greater than that of healthy-weight participants (1.32; 0.99-1.49; P < 0.001) and had significantly faster leucine flux [218; 196-259 compared with 145; 138-149 µmol · kg fat-free mass (FFM)-1 · h-1], oxidation (24.0; 17.9-29.8 compared with 16.1; 14.3-18.5 µmol · kg FFM-1 · h-1), and nonoxidative disposal (204; 190-247 compared with 138; 129-140 µmol · kg FFM-1 · h-1) (P < 0.017 for all). After surgery, the morbidly obese had a marked improvement in IR (3.54; 3.06-6.08; P = 0.008) and significant reductions in BCAA concentrations (113; 95-157 µmol/L) and leucine oxidation (9.37; 6.85-15.2 µmol · kg FFM-1 · h-1) (P = 0.017 for both). Further, leucine flux in this group correlated significantly with IR (r = 0.78, P < 0.001). CONCLUSIONS: BCAA oxidation is not impaired but elevated in individuals with morbid obesity. Plasma BCAA concentrations are lowered after surgery owing to slower breakdown of body proteins as insulin's ability to suppress proteolysis is restored. These findings suggest that IR is the underlying cause and not the consequence of elevated BCAAs in obesity.

Preovulatory exposure to a protein-restricted diet disrupts amino acid kinetics and alters mitochondrial structure and function in the rat oocyte and is partially rescued by folic acid.

BACKGROUND: Detrimental exposures during pregnancy have been implicated in programming offspring to develop permanent changes in physiology and metabolism, increasing the risk for developing diseases in adulthood such as hypertension, diabetes, heart disease and obesity. This study investigated the effects of protein restriction on the metabolism of amino acids within the oocyte, liver, and whole organism in a rat model as well as effects on mitochondrial ultrastructure and function in the cumulus oocyte complex. METHODS: Wistar outbred female rats 8-11 weeks of age (n = 24) were assigned to three isocaloric dietary groups, including control (C), low protein (LP) and low protein supplemented with folate (LPF). Animals were superovulated and 48 h later underwent central catheterization. Isotopic tracers of 1-13C-5C2H3-methionine, 2H2-cysteine, U-13C3-cysteine and U-13C3-serine were administered by a 4 h prime-constant rate infusion. After sacrifice, oocytes were denuded of cumulus cells and liver specimens were obtained. RESULTS: Oocytes demonstrated reduced serine flux in LP vs. LPF (p < 0.05), reduced cysteine flux in LP and LPF vs. C (p < 0.05), and a trend toward reduced transsulfuration in LP vs. C and LPF. Folic acid supplementation reversed observed effects on serine flux and transsulfuration. Preovulatory protein restriction increased whole-body methionine transmethylation, methionine transsulfuration and the flux of serine in LP and LPF vs. C (p = 0.003, p = 0.002, p = 0.005). The concentration of glutathione was increased in erythrocytes and liver in LP and LPF vs. C (p = 0.003 and p = 0.0003). Oocyte mitochondrial ultrastructure in LP and LPF had increased proportions of abnormal mitochondria vs. C (p < 0.01 and p < 0.05). Cumulus cell mitochondrial ultrastructure in LP and LPF groups had increased proportions of abnormal mitochondria vs. C (p < 0.001 and p < 0.05). Preovulatory protein restriction altered oocyte expression of Drp1, Opa-1, Mfn1/2, Parl and Ndufb6 (p < 0.05) and Hk2 (p < 0.01), which are genes involved in mitochondrial fission (division) and fusion, mitochondrial apoptotic mechanisms, respiratory electron transport and glucose metabolism. CONCLUSIONS: Preovulatory protein restriction resulted in altered amino acid metabolism, abnormal cumulus oocyte complex mitochondrial ultrastructure and differential oocyte expression of genes related to mitochondrial biogenesis.

Arginine Metabolism Is Altered in Adults with A-ß + Ketosis-Prone Diabetes.

Background: A-ß + ketosis-prone diabetes (KPD) is a subset of type 2 diabetes in which patients have severe but reversible ß cell dysfunction of unknown etiology. Plasma metabolomic analysis indicates that abnormal arginine metabolism may be involved. Objective: The objective of this study was to determine the relation between gut microbiome and arginine metabolism and the relation between arginine availability and ß cell function in KPD patients compared with control participants. Methods: Kinetics of arginine and related metabolites were measured with stable isotope tracers, and insulin secretory responses to arginine and glucose were determined under euglycemic and hyperglycemic conditions in 6 KPD patients and 6 age-, gender-, and body mass index-matched control participants. Glucose potentiation of arginine-induced insulin secretion was performed in a different set of 6 KPD and 3 control participants. Results: Arginine availability was higher in KPD patients during euglycemia [53.5 ± 4.3 (mean ± SEM) compared with 40.3 ± 2.4 µmol · kg lean body mass (LBM)-1 · h-1, P = 0.03] but declined more in response to hyperglycemia (Δ 10.15 ± 2.6 compared with Δ 3.20 ± 1.3 µmol · kg LBM-1 · h-1, P = 0.041). During hyperglycemia, ornithine flux was not different between groups but after an arginine bolus, plasma ornithine AUC trended higher in KPD patients (3360 ± 294 compared with 2584 ± 259 min · µmol · L-1, P = 0.08). In both euglycemia and hyperglycemia, the first-phase insulin responses to glucose stimulation were lower in KPD patients (euglycemic insulin AUC 282 ± 108 compared with 926 ± 257 min · µU · mL-1, P = 0.02; hyperglycemic insulin AUC 358 ± 79 compared with 866 ± 292 min · µU · mL-1, P = 0.05), but exogenous arginine restored first-phase insulin secretion in KPD patients to the level of control participants. Conclusion: Compared with control participants, KPD patients have increased arginine availability in the euglycemic state, indicating a higher requirement. This is compromised during hyperglycemia, with an inadequate supply of arginine to sustain metabolic functions such as insulin secretion. Exogenous arginine administration restores a normal insulin secretory response.

Assessment of Nitric Oxide Production in Mitochondrial Encephalomyopathy, Lactic Acidosis, and Stroke-Like Episodes Syndrome with the Use of a Stable Isotope Tracer Infusion Technique.

Mitochondrial disorders result from dysfunctional mitochondria that are unable to generate sufficient energy to meet the needs of various organs. Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) syndrome is one of the most frequent maternally inherited mitochondrial disorders. There is growing evidence that nitric oxide (NO) deficiency occurs in MELAS syndrome and results in impaired blood perfusion that contributes significantly to several complications in this disease. NO is synthesized from arginine by NO synthase, which catalyzes the conversion of arginine to NO and citrulline. Citrulline can be recycled into arginine, and therefore, both arginine and citrulline support NO synthesis. The use of 15N2-arginine and 13C-,2H4-citrulline stable isotope infusion allows measuring arginine flux; citrulline flux; citrulline-to-arginine flux, which represents the de novo arginine synthesis rate; and arginine-to-citrulline flux, which represents the NO production rate. The objective of this review is to highlight the utility of this method in providing additional evidence for NO deficiency in MELAS syndrome, adding more insight into the potential mechanisms of NO deficiency in this syndrome, and allowing for the assessment of the effects of supplementation with the NO donors, arginine and citrulline, on improving NO production in MELAS syndrome.

Combined Vitamin B-12 and Balanced Protein-Energy Supplementation Affect Homocysteine Remethylation in the Methionine Cycle in Pregnant South Indian Women of Low Vitamin B-12 Status.

Background: Low-quality dietary protein intake and vitamin B-12 deficiency could interact to decrease methionine transmethylation and remethylation rates during pregnancy and may affect epigenetic modifications of the fetal genome.Objective: The objective of this randomized, partially open-labeled intervention trial was to examine the effect of supplemental high-quality protein and vitamin B-12 on third-trimester methionine kinetics in pregnant Indian women with a low vitamin B-12 status.Methods: Pregnant women with low serum vitamin B-12 concentrations (<200 pmol/L) were randomly assigned to 1 of 3 groups: the first group received balanced protein-energy supplementation of 500 mL milk/d plus a 10-µg vitamin B-12 tablet/d (M+B-12 group; n = 30), the second group received milk (500 mL/d) plus a placebo tablet (M+P group; n = 30), and the third group received a placebo tablet alone (P group; n = 33). Third-trimester fasting plasma amino acid kinetics were measured by infusing 1-13C,methyl-2H3-methionine, ring-2H5-phenylalanine, ring-2H4-tyrosine,1-13C-glycine, and 2,3,3-2H3,15N-serine in a subset of participants. Placental mRNA expression of genes involved in methionine pathways, placental long interspersed nuclear elements 1 (LINE-1) methylation, and promoter methylation levels of vascular endothelial growth factor (VEGF) were analyzed.Results: Remethylation rates in the M+B-12, M+P, and P groups were 5.1 ± 1.7, 4.1 ± 1.0, and, 5.0 ± 1.4 µmol ⋅ kg-1 ⋅ h-1, respectively (P = 0.057), such that the percentage of transmethylation remethylated to methionine tended to be higher in the M+B-12 group (49.5% ± 10.5%) than in the M+P group (42.3% ± 8.4%; P = 0.053) but neither differed from the P group (44.2% ± 8.1%; P > 0.1). Placental mRNA expression, LINE-1, and VEGF promoter methylation did not differ between groups.Conclusions: Combined vitamin B-12 and balanced protein-energy supplementation increased the homocysteine remethylation rate in late pregnancy. Thus, vitamin B-12 along with balanced protein-energy supplementation is critical for optimal functioning of the methionine cycle in the third trimester of pregnancy in Indian women with low serum vitamin B-12 in early pregnancy. This trial was registered at clinicaltrials.gov as CTRI/2016/01/006578.

Impaired nitric oxide production in children with MELAS syndrome and the effect of arginine and citrulline supplementation.

Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) syndrome is one of the most frequent maternally inherited mitochondrial disorders. The pathogenesis of this syndrome is not fully understood and believed to result from several interacting mechanisms including impaired mitochondrial energy production, microvasculature angiopathy, and nitric oxide (NO) deficiency. NO deficiency in MELAS syndrome is likely to be multifactorial in origin with the decreased availability of the NO precursors, arginine and citrulline, playing a major role. In this study we used stable isotope infusion techniques to assess NO production in children with MELAS syndrome and healthy pediatric controls. We also assessed the effect of oral arginine and citrulline supplementations on NO production in children with MELAS syndrome. When compared to control subjects, children with MELAS syndrome were found to have lower NO production, arginine flux, plasma arginine, and citrulline flux. In children with MELAS syndrome, arginine supplementation resulted in increased NO production, arginine flux, and arginine concentration. Citrulline supplementation resulted in a greater increase of these parameters. Additionally, citrulline supplementation was associated with a robust increase in citrulline concentration and flux and de novo arginine synthesis rate. The greater effect of citrulline in increasing NO production is due to its greater ability to increase arginine availability particularly in the intracellular compartment in which NO synthesis takes place. This study, which is the first one to assess NO metabolism in children with mitochondrial diseases, adds more evidence to the notion that NO deficiency occurs in MELAS syndrome, suggests a better effect for citrulline because of its greater role as NO precursor, and indicates that impaired NO production occurs in children as well as adults with MELAS syndrome. Thus, the initiation of treatment with NO precursors may be beneficial earlier in life. Controlled clinical trials to assess the therapeutic effects of arginine and citrulline on clinical complications of MELAS syndrome are needed.

Energy and Protein Supplementation Does Not Affect Protein and Amino Acid Kinetics or Pregnancy Outcomes in Underweight Indian Women.

BACKGROUND: In India, the prevalence of low birth weight is high in women with a low body mass index (BMI), suggesting that underweight women are not capable of providing adequate energy and protein for fetal growth. Furthermore, as pregnancy progresses, there is increased need to provide methyl groups for methylation reactions associated with the synthesis of new proteins and, unlike normal-BMI American women, low-BMI Indian women are unable to increase methionine transmethylation and remethylation rates as pregnancy progresses from trimester 1 to 3. This also negatively influences birth weight. OBJECTIVE: The aim was to determine the effect of dietary supplementation with energy and protein from 12 ± 1 wk of gestation to time of delivery compared with no supplement on pregnancy outcomes, protein kinetics, and the fluxes of the methyl group donors serine and glycine. METHODS: Protein kinetics and serine and glycine fluxes were measured by using standard stable isotope tracer methods in the fasting and postprandial states in 24 pregnant women aged 22.9 ± 0.7 y with low BMIs [BMI (in kg/m(2)) ≤18.5] at 12 ± 1 wk (trimester 1) and 30 ± 1 wk (trimester 3) of gestation. After the first measurement, subjects were randomly assigned to either receive the supplement (300 kcal/d, 15 g protein/d) or no supplement. RESULTS: Supplementation had no significant effect on any variable of pregnancy outcome, and except for fasting state decreases in leucine flux (125 ± 7.14 compared with 113 ± 5.06 µmol ⋅ kg(-1) ⋅ h(-1); P = 0.04) and nonoxidative disposal (110 ± 6.97 compared with 101 ± 3.69 µmol ⋅ kg(-1) ⋅ h(-1); P = 0.02) from trimesters 1 to 3, it had no effect on any other leucine kinetic variable or urea, glycine, and serine fluxes. CONCLUSION: We conclude that in Indian women with a low BMI, supplementation with energy and protein from week 12 of pregnancy to time of delivery does not improve pregnancy outcome, whole-body protein kinetics, or serine and glycine fluxes.

Unlike pregnant adult women, pregnant adolescent girls cannot maintain glycine flux during late pregnancy because of decreased synthesis from serine.

During pregnancy, glycine and serine become more important because they are the primary suppliers of methyl groups for the synthesis of fetal DNA, and more glycine is required for fetal collagen synthesis as pregnancy progresses. In an earlier study, we reported that glycine flux decreased by 39% from the first to the third trimester in pregnant adolescent girls. As serine is a primary precursor for glycine synthesis, the objective of this study was to measure and compare glycine and serine fluxes and inter-conversions in pregnant adolescent girls and adult women in the first and third trimesters. Measurements were made after an overnight fast by continuous intravenous infusions of 2H2-glycine and 15N-serine in eleven adolescent girls (17·4 (se 0·1) years of age) and in ten adult women (25·8 (se 0·5) years of age) for 4 h. Adolescent girls had significantly slower glycine flux and they made less glycine from serine in the third (P<0·05) than in the first trimester. Baby birth length was significantly shorter of adolescent girls (P=0·04) and was significantly associated with third trimester glycine flux. These findings suggest that the pregnant adolescent cannot maintain glycine flux in late pregnancy compared with early pregnancy because of decreased synthesis from serine. It is possible that the inability to maintain glycine synthesis makes her fetus vulnerable to impaired cartilage synthesis, and thus linear growth.

The Microbiome, Intestinal Function, and Arginine Metabolism of Healthy Indian Women Are Different from Those of American and Jamaican Women.

BACKGROUND: Indian women have slower arginine flux during pregnancy compared with American and Jamaican women. Arginine is a semi-essential amino acid that becomes essential during periods of rapid lean tissue deposition. It is synthesized only from citrulline, a nondietary amino acid produced mainly in the gut. The gut is therefore a key site of arginine and citrulline metabolism, and gut microbiota may affect their metabolism. OBJECTIVE: The objective of this study was to identify differences in the gut microbiota of nonpregnant American, Indian, and Jamaican women and characterize the relations between the gut microbiota, gut function, and citrulline and arginine metabolism. METHODS: Thirty healthy American, Indian, and Jamaican women (n = 10/group), aged 28.3 ± 0.8 y, were infused intravenously with [guanidino-15N2]arginine, [5,5-2H2]citrulline, and [15N2]ornithine and given oral [U-13C6]arginine in the fasting and postprandial states. Fecal bacterial communities were characterized by 16S rRNA gene sequencing. RESULTS: In the fasting state, Indian women had lower citrulline flux than did American and Jamaican women [7.0 ± 0.4 compared with 9.1 ± 0.4 and 8.9 ± 0.2 µmol ⋅ kg fat-free mass (FFM)-1 ⋅ h-1, P = 0.01] and greater enteral arginine conversion to ornithine than did American women (1.4 ± 0.11 compared with 1.0 ± 0.08 µmol ⋅ kg FFM-1 ⋅ h-1, P = 0.04). They also had lower mannitol excretion than American and Jamaican women (154 ± 37.1 compared with 372 ± 51.8 and 410 ± 39.6 mg/6 h, P < 0.01). Three dominant stool community types characterized by increased abundances of the genera Prevotella, Bacteroides, and Bacteroides with Clostridium were identified. Indian women had increased mean relative abundances of Prevotella (42%) compared to American and Jamaican women (7% and < 1%, P = 0.03) which were associated with diet, impaired intestinal absorptive capacity, and arginine flux. CONCLUSIONS: These findings suggest that dysregulated intestinal function and a unique gut microbiome may contribute to altered arginine metabolism in Indian women.

Indian women of childbearing age do not metabolically conserve arginine as do American and Jamaican women.

BACKGROUND: In a previous study in pregnant American women, we reported that arginine flux and nitric oxide synthesis increased in trimester 2. More recently, we reported that Indian women do not increase arginine flux during pregnancy as their American or Jamaican counterparts do. OBJECTIVE: The purpose of this study was to determine whether Indian women of childbearing age are producing less arginine and/or catabolizing more arginine and therefore have less available for anabolic pathways than do Jamaican and American women. METHODS: Thirty healthy women aged 28.3 ± 0.8 y from the United States, India, and Jamaica (n = 10/group) were given 6 h primed, constant intravenous infusions of guanidino-¹5N2-arginine, 5,5-²H2-citrulline, ¹5N2-ornithine, and ring-²H5-phenylalanine, in addition to primed, oral doses of U-¹³C6-arginine in both the fasting and postprandial states. An oral dose of deuterium oxide was also given to determine fat-free mass (FFM). RESULTS: Compared with American women, Indian and Jamaican women had greater ornithine fluxes (µmol · kg fat FFM⁻¹ · h⁻¹) in the fasting and postprandial states (27.3 ± 2.5 vs. 39.6 ± 3.7 and 37.2 ± 2.0, respectively, P = 0.01), indicating greater arginine catabolism. However, Jamaican women had a higher endogenous arginine flux than did Indian and American women in the fasting (66.1 ± 3.1 vs. 54.2 ± 3.1 and 56.1 ± 2.1, respectively, P = 0.01) and postprandial (53.8 ± 2.2 vs. 43.7 ± 4.9 and 42.8 ± 3.1, respectively, P = 0.06) states. As a consequence, Indian women had lower arginine bioavailability (µmol · kg FFM⁻¹ · h⁻¹) in the fasting state (42.0 ± 2.6) than did American (49.9 ± 1.3, P = 0.045) and Jamaican (55.5 ± 3.5, P = 0.004) women, as well as in the postprandial state (40.7 ± 3.5 vs. 51.8 ± 1.2 and 57.5 ± 3.2, respectively, P = 0.001). CONCLUSION: Compared with American and Jamaican women, Indian women of childbearing age have a decreased arginine supply because of increased arginine catabolism without an increase in arginine flux.

Dietary supplementation with aromatic amino acids increases protein synthesis in children with severe acute malnutrition.

Although 2 earlier studies reported that aromatic amino acid (AAA) supplementation of children with severe acute malnutrition (SAM) improved whole-body protein anabolism during the early postadmission (maintenance) phase of rehabilitation, it is not known whether this positive effect was maintained during the catch-up growth and recovery phases of treatment. This study aimed to determine whether supplementation with an AAA cocktail (330 mg · kg(-1) · d(-1)) vs. isonitrogenous Ala would improve measures of protein kinetics in 22 children, aged 4-31 mo, during the catch-up growth and recovery phases of treatment for SAM. Protein kinetics were assessed by measuring leucine, phenylalanine, and urea kinetics with the use of standard stable isotope tracer methods in the fed state. Supplementation started at the end of the maintenance period when the children were clinically/metabolically stable and continued up to full nutritional recovery. Three experiments were performed: at the end of maintenance (at ∼13 d postadmission), at mid-catch-up growth (at ∼23 d post- admission when the children had replenished 50% of their weight deficit), and at recovery (at ∼48 d postadmission when they had achieved at least 90% weight for length). Children in the AAA group had significantly faster protein synthesis compared with those in the Ala group at mid-catch-up growth (101 ± 10 vs. 72 ± 7 µmol phenylalanine · kg(-1) · h(-1); P < 0.05) and better protein balance at mid-catch-up growth (49 ± 5 vs. 30 ± 2 µmol phenylalanine · kg(-1) · h(-1); P < 0.05) and at recovery (37 ± 8 vs. 11 ± 3 µmol phenylalanine · kg(-1) · h(-1); P < 0.05). We conclude that dietary supplementation with AAA accelerates net protein synthesis in children during nutritional rehabilitation for SAM.

Adaptation of in vivo amino acid kinetics facilitates increased amino acid availability for fetal growth in adolescent and adult pregnancies alike.

During pregnancy, adult women with a normal BMI synthesise extra amino acids after an overnight fast by increasing body protein breakdown and decreasing amino acid oxidation. It is not known whether adolescent girls can make these adaptations during pregnancy. The present study aimed to measure and compare the protein, glutamine and alanine kinetics of adult women and adolescent girls at early-, mid- and late-pregnancy. Kinetics were measured in the overnight fasted state using intravenous infusions of 13C-leucine, 15N-glutamine and 15N-alanine in ten adults and twenty adolescents aged 14-17 years in the first and second trimesters (phase 1 study) and infusions of 13C-leucine and 15N2-urea in ten adults and eleven adolescents aged 16-17 years in the first and third trimesters (phase 2 study). In phase 1 study, there were no significant differences between the groups with regard to any of the kinetic parameters measured. In both groups, leucine flux increased (P< 0.05), the percentage of leucine flux oxidised decreased (P< 0.05) and non-oxidative leucine disposal to protein synthesis increased (P< 0.05) from the first to the second trimester. In phase2 study, leucine flux was significantly slower (P< 0.05) in the adult group than in the adolescent group during both trimesters, and whole-body leucine flux and non-oxidative leucine disposal increased significantly in the adolescent group (P< 0.05, respectively) and were higher in the adult group from the first to the third trimester. These results suggest that similar to their adult counterparts after an overnight fast, adolescent girls with a normal BMI provide extra amino acids required for net protein deposition during pregnancy by increasing protein breakdown and decreasing amino acid oxidation.

The impact of obesity-associated glycine deficiency on the elimination of endogenous and exogenous metabolites via the glycine conjugation pathway.

Background: Glycine is an integral component of the human detoxification system as it reacts with potentially toxic exogenous and endogenously produced compounds and metabolites via the glycine conjugation pathway for urinary excretion. Because individuals with obesity have reduced glycine availability, this detoxification pathway may be compromised. However, it should be restored after bariatric surgery because of increased glycine production. Objective: To examine the impact of obesity-associated glycine deficiency on the glycine conjugation pathway. We hypothesize that the synthesis rates of acylglycines from endogenous and exogenous sources are significantly reduced in individuals with obesity but increase after bariatric surgery. Methods: We recruited 21 participants with class III obesity and 21 with healthy weight as controls. At baseline, [1,2-13C2] glycine was infused to study the glycine conjugation pathway by quantifying the synthesis rates of several acylglycines. The same measurements were repeated in participants with obesity six months after bariatric surgery. Data are presented as mean ± standard deviation, and p-value< 0.05 is considered statistically significant. Results: Baseline data of 20 participants with obesity were first compared to controls. Participants with obesity were significantly heavier than controls (mean BMI 40.5 ± 7.1 vs. 20.8 ± 2.1 kg/m2). They had significantly lower plasma glycine concentration (168 ± 30 vs. 209 ± 50 µmol/L) and slower absolute synthesis rates of acetylglycine, isobutyrylglycine, tigylglycine, isovalerylglycine, and hexanoylglycine. Pre- and post-surgery data were available for 16 participants with obesity. Post-surgery BMI decreased from 40.9 ± 7.3 to 31.6 ± 6.0 kg/m2. Plasma glycine concentration increased from 164 ± 26 to 212 ± 38 µmol/L) and was associated with significantly higher rates of excretion of acetylglycine, isobutyrylglycine, tigylglycine, isovalerylglycine, and hexanoylglycine. Benzoic acid (a xenobiotic dicarboxylic acid) is excreted as benzoylglycine; its synthesis rate was significantly slower in participants with obesity but increased after bariatric surgery. Conclusion: Obesity-associated glycine deficiency impairs the human body's ability to eliminate endogenous and exogenous metabolites/compounds via the glycine conjugation pathway. This impairment is ameliorated when glycine supply is restored after bariatric surgery. These findings imply that dietary glycine supplementation could treat obesity-associated metabolic complications due to the accumulation of intramitochondrial toxic metabolites. Clinical trial registration: https://clinicaltrials.gov/study/NCT04660513, identifier NCT04660513.

Alterations in glutamine metabolism and its conversion to citrulline in sepsis.

In enterocytes, glutamine serves as the major source of energy; another metabolic fate of glutamine is conversion to citrulline. Because sepsis can affect gut function and integrity, alterations in glutamine metabolism may exist and lead to decreased citrulline production. This study aimed to investigate how sepsis affects glutamine metabolism, including its conversion to citrulline, by measuring glutamine and citrulline flux, fractional splanchnic extraction of glutamine and leucine, and the contribution of glutamine nitrogen to citrulline in septic patients and healthy controls. Eight patients with severe sepsis and 10 healthy controls were given primed, constant intravenous infusion of [(2)H2]citrulline and sequential administration of intravenous and enteral [α-(15)N]glutamine and [(13)C]leucine in the postabsorptive state. The results showed that, compared with healthy controls, septic patients had a significantly lower whole body citrulline flux and plasma concentration, higher endogenous leucine flux, and higher glutamine clearance. Fractional splanchnic extraction of leucine was higher in septic patients than in controls, but fractional extraction of glutamine was not different. The majority of the (15)N label transferred from glutamine to citrulline was found at the α-position. These results demonstrate that lower glutamine plasma concentrations in sepsis were a result of increased glutamine clearance. Despite adequate splanchnic uptake of glutamine, there is decreased production of citrulline, suggesting a defect in the metabolic conversion of glutamine to citrulline, decreased uptake of glutamine by the enterocyte but increased uptake by the liver, and/or shunting of glutamine to other metabolic pathways.

Free living total energy expenditure in young South Indian children at risk of environmental enteric dysfunction and its relation to faltered linear growth.

BACKGROUND/OBJECTIVES: Chronic immune activation in Environmental Enteric Dysfunction (EED) could lead to increased fed-state metabolic rate (MR) or total energy expenditure (TEE) and limit the energy available for optimal linear growth. In a secondary analysis, MR and TEE were compared in young Indian children from urban slums, with and without stunting or EED. SUBJECTS/METHODS: Children (18-24 months, n = 69) were classified into non-stunted (LAZ ≥ -2) and stunted (LAZ < -2), and no-EED (lactulose rhamnose ratio, LRR < 0.068) and EED (LRR ≥ 0.068) groups. Associations between MR and TEE (kcal per kg bodyweight [BW] or fat free mass [FFM]) with stunting and EED were examined using logistic regression. RESULTS: Median TEE was significantly higher in the stunted compared to non-stunted group (76.8 versus 92.0 kcal/kg BW/day, p = <0.01). The adjusted (for sex, FFM, EED) odds ratio (AOR) for stunting with TEE (kcal/day) was 1.01 (95% CI: 1.00, 1.01), but importantly, there was no interaction between EED and TEE. The median TEE was also significantly higher in the EED compared to the no-EED group (89.1 vs 76.8 kcal/kg BW/day, p = 0.02), and the AOR (adjusted for sex and TEE) for stunting with EED was 3.56 (95% CI:1.09, 11.63). MR (per kg BW or FFM) was not associated with stunting or EED. CONCLUSION: Higher TEE and presence of EED were independently and positively associated with stunting. Children with EED also had higher TEE but not MR. Energetically, the higher TEE in stunted children may not specifically be linked to the presence of EED, although the latter independently had higher odds of stunting.

The Systemic Availability of Indispensable Amino Acids from Orally Ingested Algal and Legume Protein in Young Children at Risk of Environmental Enteric Dysfunction.

BACKGROUND: The digestion and absorption of ingested protein may be reduced in children with environmental enteric dysfunction (EED), reducing systemic amino acid availability for protein synthesis with resultant growth faltering. This has not been directly measured in children with EED and associated growth faltering. OBJECTIVES: To evaluate the systemic availability of algal (spirulina) and legume (mung bean) indispensable amino acids (IAAs) in children with EED. METHODS: Indian children (18-24 mo) from urban slums were assigned to EED (n = 24) or no-EED (control, n = 17) groups based on the lactulose rhamnose test, where the lactulose rhamnose ratio cutoff for diagnosing EED (≥0.068) was the mean + 2SD of its distribution in healthy, age-, and sex-matched children of high socioeconomic status. Fecal biomarkers of EED were also measured. Systemic IAA availability was calculated from the plasma: meal IAA enrichment ratio for each protein. True ileal mung bean IAA digestibility was measured by the dual isotope tracer method using spirulina protein as reference. Co-administration of free 13C6-phenylalanine allowed for estimating true ileal phenylalanine digestibility of both proteins, and a phenylalanine absorption index. RESULTS: There was no significant difference (independent t-test) in the systemic IAA availability from spirulina or mung bean protein between EED and no-EED groups. There was no between-group difference in true ileal phenylalanine digestibility and its absorption index, or in mung bean IAA digestibility. CONCLUSIONS: The systemic IAA availability of algal and legume protein, or the latter's IAA/phenylalanine digestibility, is not significantly reduced in children with EED and does not correlate with linear growth. This study was registered in Clinical Trials Registry of India (CTRI) with registration number: CTRI/2017/02/007921.

Supplementing Glycine and N-Acetylcysteine (GlyNAC) in Older Adults Improves Glutathione Deficiency, Oxidative Stress, Mitochondrial Dysfunction, Inflammation, Physical Function, and Aging Hallmarks: A Randomized Clinical Trial.

BACKGROUND: Elevated oxidative stress (OxS), mitochondrial dysfunction, and hallmarks of aging are identified as key contributors to aging, but improving/reversing these defects in older adults (OA) is challenging. In prior studies, we identified that deficiency of the intracellular antioxidant glutathione (GSH) could play a role and reported that supplementing GlyNAC (combination of glycine and N-acetylcysteine [NAC]) in aged mice improved GSH deficiency, OxS, mitochondrial fatty-acid oxidation (MFO), and insulin resistance (IR). To test whether GlyNAC supplementation in OA could improve GSH deficiency, OxS, mitochondrial dysfunction, IR, physical function, and aging hallmarks, we conducted a placebo-controlled randomized clinical trial. METHODS: Twenty-four OA and 12 young adults (YA) were studied. OA was randomized to receive either GlyNAC (N = 12) or isonitrogenous alanine placebo (N = 12) for 16-weeks; YA (N = 12) received GlyNAC for 2-weeks. Participants were studied before, after 2-weeks, and after 16-weeks of supplementation to assess GSH concentrations, OxS, MFO, molecular regulators of energy metabolism, inflammation, endothelial function, IR, aging hallmarks, gait speed, muscle strength, 6-minute walk test, body composition, and blood pressure. RESULTS: Compared to YA, OA had GSH deficiency, OxS, mitochondrial dysfunction (with defective molecular regulation), inflammation, endothelial dysfunction, IR, multiple aging hallmarks, impaired physical function, increased waist circumference, and systolic blood pressure. GlyNAC (and not placebo) supplementation in OA improved/corrected these defects. CONCLUSION: GlyNAC supplementation in OA for 16-weeks was safe and well-tolerated. By combining the benefits of glycine, NAC and GSH, GlyNAC is an effective nutritional supplement that improves and reverses multiple age-associated abnormalities to promote health in aging humans. Clinical Trials Registration Number: NCT01870193.

Plasma amino acid signatures define types of pediatric diabetes.

BACKGROUND & AIMS: Metabolic biomarkers with pathophysiological relevance is lacking in pediatric diabetes. We aimed to identify novel metabolic biomarkers in pediatric type 1 (T1D) and type 2 diabetes (T2D). We hypothesized that (1) targeted plasma metabolomics, focused on plasma amino acid concentrations, could identify distinctively altered patterns in children with T1D or T2D, and (2) there are specific changes in concentrations of metabolites related to branch chain amino acids (BCAA) and arginine metabolism in children with T2D. METHODS: In a pilot study, we enrolled children with T1D (n = 15) and T2D (n = 13), and healthy controls (n = 15). Fasting plasma amino acid concentrations were measured by ultra-performance liquid chromatography, and compared between the groups after adjustment for confounding factors. RESULTS: The mean age (SD) of participants was 16.4 (0.9) years. There were no group differences in age, gender, race/ethnicity, or 24-h protein intake. Mean BMI percentile was higher in the T2D than the T1D group or controls (p < 0.001). The T2D group had lower arginine, citrulline, glutamine, glycine, phenylalanine, methionine, threonine, asparagine and symmetric dimethylarginine (SDMA) but higher aspartate than controls, after adjusting for BMI percentiles (all p < 0.05). Children with T2D also had lower glycine but higher ornithine, proline, leucine, isoleucine, valine, total BCAA, lysine and tyrosine than those with T1D after adjusting for confounding factors (all p < 0.05). Children with T1D had lower phenylalanine, methionine, threonine, glutamine, tyrosine, asymmetric dimethylarginine (ADMA) and SDMA than controls (all p < 0.05). CONCLUSIONS: Children with T2D and T1D have distinct fasting plasma amino acid signatures that suggest varying pathogenic mechanisms and could serve as biomarkers for these conditions.

Restoration of impaired nitric oxide production in MELAS syndrome with citrulline and arginine supplementation.

Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) syndrome is one of the most common mitochondrial disorders. Although the pathogenesis of stroke-like episodes remains unclear, it has been suggested that mitochondrial proliferation may result in endothelial dysfunction and decreased nitric oxide (NO) availability leading to cerebral ischemic events. This study aimed to assess NO production in subjects with MELAS syndrome and the effect of the NO precursors arginine and citrulline. Using stable isotope infusion techniques, we assessed arginine, citrulline, and NO metabolism in control subjects and subjects with MELAS syndrome before and after arginine or citrulline supplementation. The results showed that subjects with MELAS had lower NO synthesis rate associated with reduced citrulline flux, de novo arginine synthesis rate, and plasma arginine and citrulline concentrations, and higher plasma asymmetric dimethylarginine (ADMA) concentration and arginine clearance. We conclude that the observed impaired NO production is due to multiple factors including elevated ADMA, higher arginine clearance, and, most importantly, decreased de novo arginine synthesis secondary to decreased citrulline availability. Arginine and, to a greater extent, citrulline supplementation increased the de novo arginine synthesis rate, the plasma concentrations and flux of arginine and citrulline, and NO production. De novo arginine synthesis increased markedly with citrulline supplementation, explaining the superior efficacy of citrulline in increasing NO production. The improvement in NO production with arginine or citrulline supplementation supports their use in MELAS and suggests that citrulline may have a better therapeutic effect than arginine. These findings can have a broader relevance for other disorders marked by perturbations in NO metabolism.

Nutritional repletion of children with severe acute malnutrition does not affect VLDL apolipoprotein B-100 synthesis rate.

VLDL apo B-100 is essential for the secretion of liver fat. It is thought that synthesis of this lipoprotein is impaired in childhood severe acute malnutrition (SAM), especially in the edematous syndromes, and that this contributes to the common occurrence of hepatic steatosis in this condition. However, to our knowledge, it has not been confirmed that VLDL apo B-100 synthesis is slower in edematous compared with nonedematous SAM and that it is impaired in the malnourished compared with the well-nourished state. Therefore, VLDL apo B-100 kinetics were measured in 2 groups of children with SAM (15 edematous and 7 nonedematous), aged 4-20 mo, at 3 stages during treatment. Measurements were done at 4 ± 1 d postadmission, mid- catch-up growth in weight, and at recovery (normal weight-for-length). VLDL apo B-100 synthesis was determined using stable isotope methodology to measure the rate of incorporation of (2)H(3)-leucine into its apoprotein moiety. The fractional and absolute synthesis of VLDL apo B-100 did not differ between the groups or from the initial malnourished stage to the recovery stage. Concentrations of VLDL apo B-100 were greater in the edematous than in the nonedematous group (P < 0.04) and did not differ from the initial stage to recovery. The data indicate that VLDL apo B-100 synthesis is not reduced when children develop either edematous or nonedematous SAM.