Human Milk Composition Is Associated with Maternal Body Mass Index in a Cross-Sectional, Untargeted Metabolomics Analysis of Human Milk from Guatemalan Mothers.

Background: Maternal overweight and obesity has been associated with poor lactation performance including delayed lactogenesis and reduced duration. However, the effect on human milk composition is less well understood. Objectives: We evaluated the relationship of maternal BMI on the human milk metabolome among Guatemalan mothers. Methods: We used data from 75 Guatemalan mothers who participated in the Household Air Pollution Intervention Network trial. Maternal BMI was measured between 9 and <20 weeks of gestation. Milk samples were collected at a single time point using aseptic collection from one breast at 6 mo postpartum and analyzed using high-resolution mass spectrometry. A cross-sectional untargeted high-resolution metabolomics analysis was performed by coupling hydrophilic interaction liquid chromatography (HILIC) and reverse phase C18 chromatography with mass spectrometry. Metabolic features associated with maternal BMI were determined by a metabolome-wide association study (MWAS), adjusting for baseline maternal age, education, and dietary diversity, and perturbations in metabolic pathways were identified by pathway enrichment analysis. Results: The mean age of participants at baseline was 23.62 ± 3.81 y, and mean BMI was 24.27 ± 4.22 kg/m2. Of the total metabolic features detected by HILIC column (19,199 features) and by C18 column (11,594 features), BMI was associated with 1026 HILIC and 500 C18 features. Enriched pathways represented amino acid metabolism, galactose metabolism, and xenobiotic metabolic metabolism. However, no significant features were identified after adjusting for multiple comparisons using the Benjamini-Hochberg false discovery rate procedure (FDRBH < 0.2). Conclusions: Findings from this untargeted MWAS indicate that maternal BMI is associated with metabolic perturbations of galactose metabolism, xenobiotic metabolism, and xenobiotic metabolism by cytochrome p450 and biosynthesis of amino acid pathways. Significant metabolic pathway alterations detected in human milk were associated with energy metabolism-related pathways including carbohydrate and amino acid metabolism.This trial was registered at clinicaltrials.gov as NCT02944682.

Metabolome × Microbiome Changes Associated with a Diet-Induced Reduction in Hepatic Fat among Adolescent Boys.

Dietary sugar reduction is one therapeutic strategy for improving nonalcoholic fatty liver disease (NAFLD), and the underlying mechanisms for this effect warrant further investigation. Here, we employed metabolomics and metagenomics to examine systemic biological adaptations associated with dietary sugar restriction and (subsequent) hepatic fat reductions in youth with NAFLD. Data/samples were from a randomized controlled trial in adolescent boys (11-16 years, mean ± SD: 13.0 ± 1.9 years) with biopsy-proven NAFLD who were either provided a low free-sugar diet (LFSD) (n = 20) or consumed their usual diet (n = 20) for 8 weeks. Plasma metabolomics was performed on samples from all 40 participants by coupling hydrophilic interaction liquid chromatography (HILIC) and C18 chromatography with mass spectrometry. In a sub-sample (n = 8 LFSD group and n = 10 usual diet group), 16S ribosomal RNA (rRNA) sequencing was performed on stool to examine changes in microbial composition/diversity. The diet treatment was associated with differential expression of 419 HILIC and 205 C18 metabolite features (p < 0.05), which were enriched in amino acid pathways, including methionine/cysteine and serine/glycine/alanine metabolism (p < 0.05), and lipid pathways, including omega-3 and linoleate metabolism (p < 0.05). Quantified metabolites that were differentially changed in the LFSD group, compared to usual diet group, and representative of these enriched metabolic pathways included increased serine (p = 0.001), glycine (p = 0.004), 2-aminobutyric acid (p = 0.012), and 3-hydroxybutyric acid (p = 0.005), and decreased linolenic acid (p = 0.006). Microbiome changes included an increase in richness at the phylum level and changes in a few genera within Firmicutes. In conclusion, the LFSD treatment, compared to usual diet, was associated with metabolome and microbiome changes that may reflect biological mechanisms linking dietary sugar restriction to a therapeutic decrease in hepatic fat. Studies are needed to validate our findings and test the utility of these "omics" changes as response biomarkers.

PISTACHIo (PreemptIon of diSrupTive behAvior in CHIldren): real-time monitoring of sleep and behavior of children 3-7 years old receiving parent-child interaction therapy augment with artificial intelligence - the study protocol, pilot study.

BACKGROUND: Emotional behavior problems (EBP) are the most common and persistent mental health issues in early childhood. Early intervention programs are crucial in helping children with EBP. Parent-child interaction therapy (PCIT) is an evidence-based therapy designed to address personal difficulties of parent-child dyads as well as reduce externalizing behaviors. In clinical practice, parents consistently struggle to provide accurate characterizations of EBP symptoms (number, timing of tantrums, precipitating events) even from the week before in their young children. The main aim of the study is to evaluate feasibility of the use of smartwatches in children aged 3-7 years with EBP. METHODS: This randomized double-blind controlled study aims to recruit a total of 100 participants, consisting of 50 children aged 3-7 years with an EBP measure rated above the clinically significant range (T-score ≥ 60) (Eyberg Child Behavior Inventory-ECBI; Eyberg & Pincus, 1999) and their parents who are at least 18 years old. Participants are randomly assigned to the artificial intelligence-PCIT group (AI-PCIT) or the PCIT-sham biometric group. Outcome parameters include weekly ECBI and Pediatric Sleep Questionnaire (PSQ) as well as Child Behavior Checklist (CBCL) obtained weeks 1, 6, and 12 of the study. Two smartphone applications (Garmin connect and mEMA) and a wearable Garmin smartwatch are used collect the data to monitor step count, sleep, heart rate, and activity intensity. In the AI-PCIT group, the mEMA application will allow for the ecological momentary assessment (EMA) and will send behavioral alerts to the parent. DISCUSSION: Real-time predictive technologies to engage patients rely on daily commitment on behalf of the participant and recurrent frequent smartphone notifications. Ecological momentary assessment (EMA) provides a way to digitally phenotype in-the-moment behavior and functioning of the parent-child dyad. One of the study's goals is to determine if AI-PCIT outcomes are superior in comparison with standard PCIT. Overall, we believe that the PISTACHIo study will also be able to determine tolerability of smartwatches in children aged 3-7 with EBP and could participate in a fundamental shift from the traditional way of assessing and treating EBP to a more individualized treatment plan based on real-time information about the child's behavior. TRIAL REGISTRATION: The ongoing clinical trial study protocol conforms to the international Consolidated Standards of Reporting Trials (CONSORT) guidelines and is registered in clinicaltrials.gov (ID: NCT05077722), an international clinical trial registry.

Metabolomics of Aerobic Exercise in Chronic Stroke Survivors: A Pilot Study.

BACKGROUND: Understanding the metabolic response to exercise may aid in optimizing stroke management. Therefore, the purpose of this pilot study was to evaluate plasma metabolomic profiles in chronic stroke survivors following aerobic exercise training. METHODS: Participants (age: 62 ± 1 years, body mass index: 31 ± 1 kg/m2, mean ± standard error of the mean) were randomized to 6 months of treadmill exercise (N = 17) or whole-body stretching (N = 8) with preintervention and postintervention measurement of aerobic capacity (VO2peak). Linear models for microarray data expression analysis was performed to determine metabolic changes over time, and Mummichog was used for pathway enrichment analysis following analysis of plasma samples by high-performance liquid chromatography coupled to ultrahigh resolution mass spectrometry. RESULTS: VO2peak change was greater following exercise than stretching (18.9% versus -.2%; P < .01). Pathway enrichment analysis of differentially expressed metabolites results showed significant enrichment in 4 pathways following treadmill exercise, 3 of which (heparan-, chondroitin-, keratan-sulfate degradation) involved connective tissue metabolism and the fourth involve lipid signaling (linoleate metabolism). More pathways were altered in pre and post comparisons of stretching, including branched-chain amino acid, tryptophan, tyrosine, and urea cycle, which could indicate loss of lean body mass. CONCLUSIONS: These preliminary data show different metabolic changes due to treadmill training and stretching in chronic stroke survivors and suggest that in addition to improved aerobic capacity, weight-bearing activity, like walking, could protect against loss of lean body mass. Future studies are needed to examine the relationship between changes in metabolomic profiles to reductions in cardiometabolic risk after treadmill rehabilitation.

High-Resolution Metabolomics for Nutrition and Health Assessment of Armed Forces Personnel.

OBJECTIVE: The aim of this study was to test the utility of high-resolution metabolomics (HRM) for analysis of nutritional status and health indicators in military personnel. METHODS: Serum samples from 400 military personnel were obtained from the Department of Defense Serum Repository (DoDSR) and analyzed for metabolites related to nutrition and health status. Metabolic profile organization was studied using modulated modularity clustering (MMC). RESULTS: HRM provided quantitative measures of 61 metabolites across chemical classes for use as nutritional and clinical biomarkers. Levels were comparable to reported values except for arginine and glutamine, which were above and below reference ranges, respectively. MMC generated five clusters, three of which were associated and contained amino acids. The others contained lipids and mitochondria-related metabolites. CONCLUSIONS: HRM analysis of serum is suitable for real-time and/or retrospective evaluation of nutrition and health status of specific military cohorts.

Sulfur amino acid-free diet results in increased glutamate in human midbrain: a pilot magnetic resonance spectroscopic study.

OBJECTIVE: This pilot study was designed to determine if metabolic effects in different brain regions (left and right parietal lobes, midbrain) caused by 3 d of food consumption without methionine or cysteine could be detected by proton magnetic resonance spectroscopy. METHODS: Healthy individuals 18 to 36 y old (n = 8) were studied by magnetic resonance spectroscopy after receiving a diet with adequate sulfur amino acids (SAAs) or with zero SAA for 3 d. Pulse sequences were used to selectively measure glutathione (GSH), and linear combination modeling of spectra was used to measure other high-abundance brain metabolites and expressed relative to creatine (Cr). RESULTS: Although dietary SAAs are required to maintain GSH, the 3-d SAA insufficiency resulted in no significant change in GSH/Cr in the three brain regions. Principal component analysis of 16 metabolites measured by linear combination modeling showed that the metabolic pattern in the midbrain, but not in the parietal lobes, was distinguished according to the dietary SAAs. Multivariate statistical analysis showed that the major discriminating factors were signals of glutamate/Cr, (glutamate + glutamine)/Cr, and myoinositol/Cr. Correlation analyses between midbrain metabolites and GSH-related metabolites in plasma showed that midbrain glutamate/Cr had an inverse correlation with plasma cystine. CONCLUSION: The data show that magnetic resonance spectroscopy is a non-invasive tool suitable for nutritional assessment and suggest that nutritional imbalance caused by 3 d of SAA-free food more selectively affects the midbrain than the parietal lobes.

A sulfur amino acid-free meal increases plasma lipids in humans.

The content of sulfur amino acid (SAA) in a meal affects postprandial plasma cysteine concentrations and the redox potential of cysteine/cystine. Because such changes can affect enzyme, transporter, and receptor activities, meal content of SAA could have unrecognized effects on metabolism during the postprandial period. This pilot study used proton NMR ((1)H-NMR) spectroscopy of human plasma to test the hypothesis that dietary SAA content changes macronutrient metabolism. Healthy participants (18-36 y, 5 males and 3 females) were equilibrated for 3 d to adequate SAA, fed chemically defined meals without SAA for 5 d (depletion), and then fed isoenergetic, isonitrogenous meals containing 56 mg·kg(-1)·d(-1) SAA for 4.5 d (repletion). On the first and last day of consuming the chemically defined meals, a morning meal containing 60% of the daily food intake was given and plasma samples were collected over an 8-h postprandial time course for characterization of metabolic changes by (1)H-NMR spectroscopy. SAA-free food increased peak intensity in the plasma (1)H-NMR spectra in the postprandial period. Orthogonal signal correction/partial least squares-discriminant analysis showed changes in signals associated with lipids, some amino acids, and lactate, with notable increases in plasma lipid signals (TG, unsaturated lipid, cholesterol). Conventional lipid analyses confirmed higher plasma TG and showed an increase in plasma concentration of the lipoprotein lipase inhibitor, apoC-III. The results show that plasma (1)H-NMR spectra can provide useful macronutrient profiling following a meal challenge protocol and that a single meal with imbalanced SAA content alters postprandial lipid metabolism.

Dietary sulfur amino acid effects on fasting plasma cysteine/cystine redox potential in humans.

OBJECTIVE: Oxidation of plasma cysteine/cystine (Cys/CySS) redox potential (E(h)CySS) has been associated with risk factors for cardiovascular disease in humans. Cys and CySS are derived from dietary sulfur amino acids (SAA), but the specific effects of SAA depletion and repletion on Cys/CySS redox indices are unknown. The present study examined the effect of dietary SAA intake level on free Cys, free CySS, and E(h)CySS in human plasma under fasting conditions. METHODS: Healthy individuals aged 18-36 y (n = 13) were equilibrated to foods providing the RDA for SAA and then fed chemically defined diets without SAA (0 mg · kg(-1) · d(-1); n = 13) followed by SAA at levels approximating the mean (56 mg · kg(-1) · d(-1); n = 8) or 99th percentile (117 mg · kg(-1) · d(-1); n = 5) intake levels of Americans. Fasting plasma samples were collected daily during 4-d study periods and analyzed for free Cys, free CySS, and the E(h)CySS. RESULTS: The SAA-free diet significantly (P < 0.05) decreased plasma-free Cys concentrations and oxidized E(h)CySS values after 4 d of SAA depletion. With SAA repletion at 56 mg · kg(-1) · d(-1), plasma-free Cys increased significantly and values for E(h)CySS became more reduced. Administration of a diet providing a higher dose of SAA (117 mg · kg(-1) · d(-1)) resulted in a significantly higher level of free Cys and a more reduced E(h)CySS. CONCLUSIONS: These results show that free Cys and Cys/CySS redox potential (E(h)CySS) in fasting plasma are affected by dietary SAA intake level in humans. Significant changes occur slowly over 4 d with insufficient SAA intake, but rapidly (after 1 d) with repletion.

Postprandial cysteine/cystine redox potential in human plasma varies with meal content of sulfur amino acids.

Few data are available on plasma redox responses to sulfur amino acid (SAA) loads. In this study, we had 2 aims: to determine whether the SAA content of a meal affected postprandial plasma cysteine (Cys), cystine (CySS), or redox potential (E(h)CySS) in humans and whether SAA intake level (adequate or inadequate) in the days preceding the meal challenge affected these postprandial levels. Eight healthy individuals aged 18-36 y were equilibrated for 3 d to adequate SAA, fed chemically defined meals without SAA for 5 d (inadequate SAA) and then fed isoenergetic, isonitrogenous meals with adequate SAA for 5 d. On the first and last days with the chemically defined meals, a morning meal containing 60% of the daily food intake was given, and plasma Cys, CySS, and E(h)CySS were determined over an 8-h postprandial time course. Following equilibration to adequate intake, provision of the meal with SAA resulted in increased plasma Cys and CySS concentrations and more reduced plasma E(h)CySS compared with the postprandial values following the same meal without SAA. Equilibration to inadequate SAA intake for the days preceding the meal challenge did not affect this response. The magnitude of the difference in postprandial plasma E(h)CySS (10 mV) due to meal content of SAA was comparable to those which alter physiologic signaling and/or are associated with disease risk. Consequently, the SAA content of meals could affect physiologic signaling and associated disease mechanisms in the postprandial period by changes in Cys, CySS, or E(h)CySS.

Individual variation in macronutrient regulation measured by proton magnetic resonance spectroscopy of human plasma.

Proton nuclear magnetic resonance ((1)H-NMR) spectroscopy of plasma provides a global metabolic profiling method that shows promise for clinical diagnostics. However, cross-sectional studies are complicated by a lack of understanding of intraindividual variation, and this limits experimental design and interpretation of data. The present study determined the diurnal variation detected by (1)H NMR spectroscopy of human plasma. Data reduction methods revealed three time-of-day metabolic patterns, which were associated with morning, afternoon, and night. Major discriminatory regions for these time-of-day patterns included the various kinds of lipid signals (-CH(2)- and -CH(2)OCOR), and the region between 3 and 4 ppm heavily overlapped with amino acids that had alpha-CH and alpha-CH(2). The phasing and duration of time-of-day patterns were variable among individuals, apparently because of individual difference in food processing/digestion and absorption and clearance of macronutrient energy sources (fat, protein, carbohydrate). The times of day that were most consistent among individuals, and therefore most useful for cross-sectional studies, were fasting morning (0830-0930), postprandial afternoon (1430-1630), and nighttime samples (0430-0530). Importantly, the integrated picture of metabolism provided by (1)H-NMR spectroscopy of plasma suggests that this approach is suitable to study complex regulatory processes, including eating patterns/eating disorders, upper gastrointestinal functions (gastric emptying, pancreatic, biliary functions), and absorption/clearance of macronutrients. Hence, (1)H-NMR spectroscopy of plasma could provide a global metabolic tolerance test to assess complex processes involved in disease, including eating disorders and the range of physiological processes causing dysregulation of energy homeostasis.

Cysteine redox potential determines pro-inflammatory IL-1beta levels.

BACKGROUND: Cysteine (Cys) and its disulfide, cystine (CySS) represent the major extracellular thiol/disulfide redox control system. The redox potential (E(h)) of Cys/CySS is centered at approximately -80 mV in the plasma of healthy adults, and oxidation of E(h) Cys/CySS is implicated in inflammation associated with various diseases. METHODOLOGY/PRINCIPAL FINDINGS: The purpose of the present study was to determine whether oxidized E(h) Cys/CySS is a determinant of interleukin (IL)-1beta levels. Results showed a 1.7-fold increase in secreted pro-IL-1beta levels in U937 monocytes exposed to oxidized E(h) Cys/CySS (-46 mV), compared to controls exposed to a physiological E(h) of -80 mV (P<0.01). In LPS-challenged mice, preservation of plasma E(h) Cys/CySS from oxidation by dietary sulfur amino acid (SAA) supplementation, was associated with a 1.6-fold decrease in plasma IL-1beta compared to control mice fed an isonitrogenous SAA-adequate diet (P<0.01). Analysis of E(h) Cys/CySS and IL-1beta in human plasma revealed a significant positive association between oxidized E(h) Cys/CySS and IL-1beta after controlling for age, gender, and BMI (P<0.001). CONCLUSIONS/SIGNIFICANCE: These data show that oxidized extracellular E(h) Cys/CySS is a determinant of IL-1beta levels, and suggest that strategies to preserve E(h) Cys/CySS may represent a means to control IL-1beta in inflammatory disease states.

Efficacy of parenteral nutrition supplemented with glutamine dipeptide to decrease hospital infections in critically ill surgical patients.

BACKGROUND: Nosocomial infections are an important cause of morbidity and mortality in the surgical intensive care unit (SICU). Clinical benefits of glutamine-supplemented parenteral nutrition may occur in hospitalized surgical patients, but efficacy data in different surgical subgroups are lacking. The objective was to determine whether glutamine-supplemented parenteral nutrition differentially affects nosocomial infection rates in selected subgroups of SICU patients. METHODS: This was a double-blind, randomized, controlled study of alanyl-glutamine dipeptide-supplemented parenteral nutrition in SICU patients requiring parenteral nutrition and SICU care after surgery for pancreatic necrosis, cardiac, vascular, or colonic surgery. Subjects (n = 59) received isocaloric/isonitrogenous parenteral nutrition, providing 1.5 g/kg/d standard glutamine-free amino acids (STD-PN) or 1.0 g/kg/d standard amino acids + 0.5 g/kg/d glutamine dipeptide (GLN-PN). Enteral feedings were advanced as tolerated. Nosocomial infections were determined until hospital discharge. RESULTS: Baseline clinical/metabolic data were similar between groups. Plasma glutamine concentrations were low in all groups and were increased by GLN-PN. GLN-PN did not alter infection rates after pancreatic necrosis surgery (17 STD-PN and 15 GLN-PN patients). In nonpancreatic surgery patients (12 STD-PN and 15 GLN-PN), GLN-PN was associated with significantly decreased total nosocomial infections (STD-PN 36 vs GLN-PN 13, P < .030), bloodstream infections (7 vs 0, P < .01), pneumonias (16 vs 6, P < .05), and infections attributed to Staphylococcus aureus (P < .01), fungi, and enteric Gram-negative bacteria (each P < .05). CONCLUSIONS: Glutamine dipeptide-supplemented parenteral nutrition did not alter infection rates following pancreatic necrosis surgery but significantly decreased infections in SICU patients after cardiac, vascular, and colonic surgery.

Depletion of plasma antioxidants in surgical intensive care unit patients requiring parenteral feeding: effects of parenteral nutrition with or without alanyl-glutamine dipeptide supplementation.

OBJECTIVES: Antioxidant depletion is common in critically ill patients. This study was designed to determine the effects of parenteral nutrition (PN), with or without glutamine (Gln) supplementation, on systemic antioxidant status in adult patients after major surgery who required PN in the surgical intensive care unit (SICU) setting. METHODS: Fifty-nine patients in the SICU who required PN after pancreatic surgery or cardiac, vascular, or colonic (non-pancreatic) surgery were randomized in a double-blinded study to receive standard PN (Gln-free) or Gln-supplemented PN (Gln-PN) in which Gln was provided as alanyl-Gln dipeptide. Conventional PN vitamin and mineral doses were administered to all subjects. Plasma concentrations of the antioxidant glutathione (GSH) and the antioxidant nutrients alpha-tocopherol, vitamin C, and zinc were determined at baseline (initiation of study PN) and again after 7 d of study PN. Data were analyzed for the total study cohort and within the pancreatic surgery and non-pancreatic (cardiac, vascular, and colonic) surgery patient subgroups. RESULTS: Mean plasma antioxidant concentrations were within or slightly below the normal ranges at baseline. However, a larger percentage of patients demonstrated below-normal baseline plasma concentrations of GSH (59%), vitamin C (59%), and zinc (68%), respectively. A smaller percentage of patients exhibited below-normal plasma alpha-tocopherol levels (21%). Study PN significantly improved plasma zinc levels in the entire study group and in each surgical subgroup. Gln-PN significantly improved the change in plasma levels of reduced GSH from baseline to day 7 in the non-pancreatic surgery patients (PN -0.27 microM versus Gln-PN +0.26 microM, P < 0.03). CONCLUSION: Low plasma levels of key antioxidants were common in this group of patients in the SICU despite administration of PN containing conventional micronutrients. Compared with standard PN, Gln-supplemented PN improved plasma GSH levels in patients in the SICU after cardiac, vascular, or colonic operations.

Diurnal variation in glutathione and cysteine redox states in human plasma.

BACKGROUND: Plasma glutathione/glutathione disulfide (GSH/GSSG) and cysteine/cystine (Cys/CySS) couples are oxidized in humans in association with oxidative stress and cardiovascular disease risk. Animal studies show that both pools undergo diurnal variations associated with dietary intake of sulfur amino acids. OBJECTIVE: The objective of this study was to determine whether the redox state of GSH, Cys, GSH/GSSG, or Cys/CySS undergoes diurnal variation in healthy adults. DESIGN: Plasma samples were collected every hour for 24 h from 63 persons aged 18-86 y who were consuming normal food (protein, 0.8 g kg(-1) d(-1); sulfur amino acids, 20 mg kg(-1) d(-1)) at standardized mealtimes. Measurements of Cys, CySS, GSH, and GSSG were used with the Nernst equation to calculate the redox states. RESULTS: Plasma Cys and GSH concentrations varied with the time of day. The highest values for plasma Cys occurred approximately 3 h after meals. Glutathione was maximal 6 h after peak plasma Cys. The calculated redox states of the GSH/GSSG and Cys/CySS couples varied in association with the concentrations of the thiol forms. Maximal reduction and oxidation of the Cys/CySS couple occurred at 2130 and 0630, whereas the respective values for the GSH/GSSG couple occurred at 0330 and 1330. The mean diurnal variation for Cys/CySS redox in persons aged >or=60 y was 1.8-fold that in persons aged <40 y. CONCLUSIONS: Cys/CySS and GSH/GSSG redox states in human plasma undergo diurnal variation with an increased magnitude of variation in Cys/CySS redox state in older persons. This variation could alter sensitivity to oxidative stress over a course of hours.