Stratification of Sepsis Patients on Admission into the Intensive Care Unit According to Differential Plasma Metabolic Phenotypes.

Delayed diagnosis of patients with sepsis or septic shock is associated with increased mortality and morbidity. UPLC-MS and NMR spectroscopy were used to measure panels of lipoproteins, lipids, biogenic amines, amino acids, and tryptophan pathway metabolites in blood plasma samples collected from 152 patients within 48 h of admission into the Intensive Care Unit (ICU) where 62 patients had no sepsis, 71 patients had sepsis, and 19 patients had septic shock. Patients with sepsis or septic shock had higher concentrations of neopterin and lower levels of HDL cholesterol and phospholipid particles in comparison to nonsepsis patients. Septic shock could be differentiated from sepsis patients based on different concentrations of 10 lipids, including significantly lower concentrations of five phosphatidylcholine species, three cholesterol esters, one dihydroceramide, and one phosphatidylethanolamine. The Supramolecular Phospholipid Composite (SPC) was reduced in all ICU patients, while the composite markers of acute phase glycoproteins were increased in the sepsis and septic shock patients within 48 h admission into ICU. We show that the plasma metabolic phenotype obtained within 48 h of ICU admission is diagnostic for the presence of sepsis and that septic shock can be differentiated from sepsis based on the lipid profile.

Exploring the Interactions between Obesity and Diabetes: Implications for Understanding Metabolic Dysregulation in a Saudi Arabian Adult Population.

The rising prevalence of obesity in Saudi Arabia is a major contributor to the nation's high levels of cardiometabolic diseases such as type 2 diabetes. To assess the impact of obesity on the diabetic metabolic phenotype presented in young Saudi Arabian adults, participants (n = 289, aged 18-40 years) were recruited and stratified into four groups: healthy weight (BMI 18.5-24.99 kg/m2) with (n = 57) and without diabetes (n = 58) or overweight/obese (BMI > 24.99 kg/m2) with (n = 102) and without diabetes (n = 72). Distinct plasma metabolic phenotypes associated with high BMI and diabetes were identified using nuclear magnetic resonance spectroscopy and ultraperformance liquid chromatography mass spectrometry. Increased plasma glucose and dysregulated lipoproteins were characteristics of obesity in individuals with and without diabetes, but the obesity-associated lipoprotein phenotype was partially masked in individuals with diabetes. Although there was little difference between diabetics and nondiabetics in the global plasma LDL cholesterol and phospholipid concentration, the distribution of lipoprotein particles was altered in diabetics with a shift toward denser and more atherogenic LDL5 and LDL6 particles, which was amplified in the presence of obesity. Further investigation is warranted in larger Middle Eastern populations to explore the dysregulation of metabolism driven by interactions between obesity and diabetes in young adults.

Cross-Validation of Metabolic Phenotypes in SARS-CoV-2 Infected Subpopulations Using Targeted Liquid Chromatography-Mass Spectrometry (LC-MS).

To ensure biological validity in metabolic phenotyping, findings must be replicated in independent sample sets. Targeted workflows have long been heralded as ideal platforms for such validation due to their robust quantitative capability. We evaluated the capability of liquid chromatography-mass spectrometry (LC-MS) assays targeting organic acids and bile acids to validate metabolic phenotypes of SARS-CoV-2 infection. Two independent sample sets were collected: (1) Australia: plasma, SARS-CoV-2 positive (n = 20), noninfected healthy controls (n = 22) and COVID-19 disease-like symptoms but negative for SARS-CoV-2 infection (n = 22). (2) Spain: serum, SARS-CoV-2 positive (n = 33) and noninfected healthy controls (n = 39). Multivariate modeling using orthogonal projections to latent structures discriminant analyses (OPLS-DA) classified healthy controls from SARS-CoV-2 positive (Australia; R2 = 0.17, ROC-AUC = 1; Spain R2 = 0.20, ROC-AUC = 1). Univariate analyses revealed 23 significantly different (p < 0.05) metabolites between healthy controls and SARS-CoV-2 positive individuals across both cohorts. Significant metabolites revealed consistent perturbations in cellular energy metabolism (pyruvic acid, and 2-oxoglutaric acid), oxidative stress (lactic acid, 2-hydroxybutyric acid), hypoxia (2-hydroxyglutaric acid, 5-aminolevulinic acid), liver activity (primary bile acids), and host-gut microbial cometabolism (hippuric acid, phenylpropionic acid, indole-3-propionic acid). These data support targeted LC-MS metabolic phenotyping workflows for biological validation in independent sample sets.

Nonsevere Burn Induces a Prolonged Systemic Metabolic Phenotype Indicative of a Persistent Inflammatory Response Postinjury.

Globally, burns are a significant cause of injury that can cause substantial acute trauma as well as lead to increased incidence of chronic comorbidity and disease. To date, research has primarily focused on the systemic response to severe injury, with little in the literature reported on the impact of nonsevere injuries (<15% total burn surface area; TBSA). To elucidate the metabolic consequences of a nonsevere burn injury, longitudinal plasma was collected from adults (n = 35) who presented at hospital with a nonsevere burn injury at admission, and at 6 week follow up. A cross-sectional baseline sample was also collected from nonburn control participants (n = 14). Samples underwent multiplatform metabolic phenotyping using 1H nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry to quantify 112 lipoprotein and glycoprotein signatures and 852 lipid species from across 20 subclasses. Multivariate data modeling (orthogonal projections to latent structures-discriminate analysis; OPLS-DA) revealed alterations in lipoprotein and lipid metabolism when comparing the baseline control to hospital admission samples, with the phenotypic signature found to be sustained at follow up. Univariate (Mann-Whitney U) testing and OPLS-DA indicated specific increases in GlycB (p-value < 1.0e-4), low density lipoprotein-2 subfractions (variable importance in projection score; VIP > 6.83e-1) and monoacyglyceride (20:4) (p-value < 1.0e-4) and decreases in circulating anti-inflammatory high-density lipoprotein-4 subfractions (VIP > 7.75e-1), phosphatidylcholines, phosphatidylglycerols, phosphatidylinositols, and phosphatidylserines. The results indicate a persistent systemic metabolic phenotype that occurs even in cases of a nonsevere burn injury. The phenotype is indicative of an acute inflammatory profile that continues to be sustained postinjury, suggesting an impact on systems health beyond the site of injury. The phenotypes contained metabolic signatures consistent with chronic inflammatory states reported to have an elevated incidence postburn injury. Such phenotypic signatures may provide patient stratification opportunities, to identify individual responses to injury, personalize intervention strategies, and improve acute care, reducing the risk of chronic comorbidity.

Integrative Molecular Structure Elucidation and Construction of an Extended Metabolic Pathway Associated with an Ancient Innate Immune Response in COVID-19 Patients.

We present compelling evidence for the existence of an extended innate viperin-dependent pathway, which provides crucial evidence for an adaptive response to viral agents, such as SARS-CoV-2. We show the in vivo biosynthesis of a family of novel endogenous cytosine metabolites with potential antiviral activities. Two-dimensional nuclear magnetic resonance (NMR) spectroscopy revealed a characteristic spin-system motif, indicating the presence of an extended panel of urinary metabolites during the acute viral replication phase. Mass spectrometry additionally enabled the characterization and quantification of the most abundant serum metabolites, showing the potential diagnostic value of the compounds for viral infections. In total, we unveiled ten nucleoside (cytosine- and uracil-based) analogue structures, eight of which were previously unknown in humans allowing us to propose a new extended viperin pathway for the innate production of antiviral compounds. The molecular structures of the nucleoside analogues and their correlation with an array of serum cytokines, including IFN-α2, IFN-γ, and IL-10, suggest an association with the viperin enzyme contributing to an ancient endogenous innate immune defense mechanism against viral infection.

Rapid and Self-Administrable Capillary Blood Microsampling Demonstrates Statistical Equivalence with Standard Venous Collections in NMR-Based Lipoprotein Analysis.

We investigated plasma and serum blood derivatives from capillary blood microsamples (500 µL, MiniCollect tubes) and corresponding venous blood (10 mL vacutainers). Samples from 20 healthy participants were analyzed by 1H NMR, and 112 lipoprotein subfraction parameters; 3 supramolecular phospholipid composite (SPC) parameters from SPC1, SPC2, and SPC3 subfractions; 2 N-acetyl signals from α-1-acid glycoprotein (Glyc), GlycA, and GlycB; and 3 calculated parameters, SPC (total), SPC3/SPC2, and Glyc (total) were assessed. Using linear regression between capillary and venous collection sites, we explained that agreement (Adj. R2 ≥ 0.8, p < 0.001) was witnessed for 86% of plasma parameters (103/120) and 88% of serum parameters (106/120), indicating that capillary lipoprotein, SPC, and Glyc concentrations follow changes in venous concentrations. These results indicate that capillary blood microsamples are suitable for sampling in remote areas and for high-frequency longitudinal sampling of the majority of lipoproteins, SPCs, and Glycs.

Meta-analysis of the Microbial Diversity Cultured in Bioreactors Simulating the Gut Microbiome.

Understanding the intricate ecological interactions within the gut microbiome and unravelling its impact on human health is a challenging task. Bioreactors are valuable tools that have contributed to our understanding of gut microbial ecology. However, there is a lack of studies describing and comparing the microbial diversity cultivated in these models. This knowledge is crucial for refining current models to reflect the gastrointestinal microbiome accurately. In this study, we analysed the microbial diversity of 1512 samples from 18 studies available in public repositories that employed cultures performed in batches and various bioreactor models to cultivate faecal microbiota. Community structure comparison between samples using t-distributed stochastic neighbour embedding and the Hellinger distance revealed a high variation between projects. The main driver of these differences was the inter-individual variation between the donor faecal inocula. Moreover, there was no overlap in the structure of the microbial communities between studies using the same bioreactor platform. In addition, α-diversity analysis using Hill numbers showed that highly complex bioreactors did not exhibit higher diversities than simpler designs. However, analyses of five projects in which the samples from the faecal inoculum were also provided revealed an amplicon sequence variants enrichment in bioreactors compared to the inoculum. Finally, a comparative analysis of the taxonomy of the families detected in the projects and the GMRepo database revealed bacterial families exclusively found in the bioreactor models. These findings highlight the potential of bioreactors to enrich low-abundance microorganisms from faecal samples, contributing to uncovering the gut microbial "dark matter".

Microbial Indoles: Key Regulators of Organ Growth and Metabolic Function.

Gut microbes supporting body growth are known but the mechanisms are less well documented. Using the microbial tryptophan metabolite indole, known to regulate prokaryotic cell division and metabolic stress conditions, we mono-colonized germ-free (GF) mice with indole-producing wild-type Escherichia coli (E. coli) or tryptophanase-encoding tnaA knockout mutant indole-non-producing E. coli. Indole mutant E. coli mice showed multiorgan growth retardation and lower levels of glycogen, cholesterol, triglycerides, and glucose, resulting in an energy deficiency despite increased food intake. Detailed analysis revealed a malfunctioning intestine, enlarged cecum, and reduced numbers of enterochromaffin cells, correlating with a metabolic phenotype consisting of impaired gut motility, diminished digestion, and lower energy harvest. Furthermore, indole mutant mice displayed reduction in serum levels of tricarboxylic acid (TCA) cycle intermediates and lipids. In stark contrast, a massive increase in serum melatonin was observed-frequently associated with accelerated oxidative stress and mitochondrial dysfunction. This observational report discloses functional roles of microbe-derived indoles regulating multiple organ functions and extends our previous report of indole-linked regulation of adult neurogenesis. Since indoles decline by age, these results imply a correlation with age-linked organ decline and levels of indoles. Interestingly, increased levels of indole-3-acetic acid, a known indole metabolite, have been shown to correlate with younger biological age, further supporting a link between biological age and levels of microbe-derived indole metabolites. The results presented in this resource paper will be useful for the future design of food intervention studies to reduce accelerated age-linked organ decline.

Unravelling inulin molecules in food sources using a matrix-assisted laser desorption/ionization magnetic resonance mass spectrometry (MALDI-MRMS) pipeline.

Inulin, a polysaccharide characterized by a ß-2,1 fructosyl-fructose structure terminating in a glucosyl moiety, is naturally present in plant roots and tubers. Current methods provide average degrees of polymerization (DP) but lack information on the distribution and absolute concentration of each DP. To address this limitation, a reproducible (CV < 10 %) high throughput (<2 min/sample) MALDI-MRMS approach capable of characterizing and quantifying inulin molecules in plants using matched-matrix consisting of α-cyano-4-hydroxycinnamic acid butylamine salt (CHCA-BA), chicory inulin-12C and inulin-13C was developed. The method identified variation in chain lengths and concentration of inulin across various plant species. Globe artichoke hearts, yacón and elephant garlic yielded similar concentrations at 15.6 g/100 g dry weight (DW), 16.8 g/100 g DW and 17.7 g/100 g DW, respectively, for DP range between 9 and 22. In contrast, Jerusalem artichoke demonstrated the highest concentration (53.4 g/100 g DW) within the same DP ranges. Jerusalem artichoke (DPs 9-32) and globe artichoke (DPs 9-36) showed similar DP distributions, while yacón and elephant garlic displayed the narrowest and broadest DP ranges (DPs 9-19 and DPs 9-45, respectively). Additionally, qualitative measurement for all inulin across all plant samples was feasible using the peak intensities normalized to Inulin-13C, and showed that the ratio of yacón, elephant garlic and Jerusalem was approximately one, two and three times that of globe artichoke. This MALDI-MRMS approach provides comprehensive insights into the structure of inulin molecules, opening avenues for in-depth investigations into how DP and concentration of inulin influence gut health and the modulation of noncommunicable diseases, as well as shedding light on refining cultivation practices to elevate the beneficial health properties associated with specific DPs.

Diet shapes the metabolite profile in the intact human ileum, which affects PYY release.

The human ileum contains a high density of enteroendocrine L-cells, which release the appetite-suppressing hormones glucagon-like peptide-1 (GLP-1) and peptide tyrosine tyrosine (PYY) in response to food intake. Recent evidence highlighted the potential role of food structures in PYY release, but the link between food structures, ileal metabolites, and appetite hormone release remains unclear owing to limited access to intact human ileum. In a randomized crossover trial (ISRCTN11327221; isrctn.com), we investigated the role of human ileum in GLP-1 and PYY release by giving healthy volunteers diets differing in fiber and food structure: high-fiber (intact or disrupted food structures) or low-fiber disrupted food structures. We used nasoenteric tubes to sample chyme from the intact distal ileum lumina of humans in the fasted state and every 60 min for 480 min postprandially. We demonstrate the highly dynamic, wide-ranging molecular environment of the ileum over time, with a substantial decrease in ileum bacterial numbers and bacterial metabolites after food intake. We also show that high-fiber diets, independent of food structure, increased PYY release compared with a low-fiber diet during 0 to 240 min postprandially. High-fiber diets also increased ileal stachyose, and a disrupted high-fiber diet increased certain ileal amino acids. Treatment of human ileal organoids with ileal fluids or an amino acid and stachyose mixture stimulated PYY expression in a similar profile to blood PYY concentrations, confirming the role of ileal metabolites in PYY release. Our study demonstrates the diet-induced changes over time in the metabolite environment of intact human ileum, which play a role in PYY release.

The Association of Cardiometabolic, Diet and Lifestyle Parameters With Plasma Glucagon-like Peptide-1: An IMI DIRECT Study.

CONTEXT: The role of glucagon-like peptide-1 (GLP-1) in type 2 diabetes (T2D) and obesity is not fully understood. OBJECTIVE: We investigate the association of cardiometabolic, diet, and lifestyle parameters on fasting and postprandial GLP-1 in people at risk of, or living with, T2D. METHODS: We analyzed cross-sectional data from the two Innovative Medicines Initiative (IMI) Diabetes Research on Patient Stratification (DIRECT) cohorts, cohort 1 (n = 2127) individuals at risk of diabetes; cohort 2 (n = 789) individuals with new-onset T2D. RESULTS: Our multiple regression analysis reveals that fasting total GLP-1 is associated with an insulin-resistant phenotype and observe a strong independent relationship with male sex, increased adiposity, and liver fat, particularly in the prediabetes population. In contrast, we showed that incremental GLP-1 decreases with worsening glycemia, higher adiposity, liver fat, male sex, and reduced insulin sensitivity in the prediabetes cohort. Higher fasting total GLP-1 was associated with a low intake of wholegrain, fruit, and vegetables in people with prediabetes, and with a high intake of red meat and alcohol in people with diabetes. CONCLUSION: These studies provide novel insights into the association between fasting and incremental GLP-1, metabolic traits of diabetes and obesity, and dietary intake, and raise intriguing questions regarding the relevance of fasting GLP-1 in the pathophysiology T2D.

Altered dietary behaviour during pregnancy impacts systemic metabolic phenotypes.

Rationale: Evidence suggests consumption of a Mediterranean diet (MD) can positively impact both maternal and offspring health, potentially mediated by a beneficial effect on inflammatory pathways. We aimed to apply metabolic profiling of serum and urine samples to assess differences between women who were stratified into high and low alignment to a MD throughout pregnancy and investigate the relationship of the diet to inflammatory markers. Methods: From the ORIGINS cohort, 51 pregnant women were stratified for persistent high and low alignment to a MD, based on validated MD questionnaires. 1H Nuclear Magnetic Resonance (NMR) spectroscopy was used to investigate the urine and serum metabolite profiles of these women at 36 weeks of pregnancy. The relationship between diet, metabolite profile and inflammatory status was investigated. Results: There were clear differences in both the food choice and metabolic profiles of women who self-reported concordance to a high (HMDA) and low (LMDA) Mediterranean diet, indicating that alignment with the MD was associated with a specific metabolic phenotype during pregnancy. Reduced meat intake and higher vegetable intake in the HMDA group was supported by increased levels of urinary hippurate (p = 0.044) and lower creatine (p = 0.047) levels. Serum concentrations of the NMR spectroscopic inflammatory biomarkers GlycA (p = 0.020) and GlycB (p = 0.016) were significantly lower in the HDMA group and were negatively associated with serum acetate, histidine and isoleucine (p < 0.05) suggesting a greater level of plant-based nutrients in the diet. Serum branched chain and aromatic amino acids were positively associated with the HMDA group while both urinary and serum creatine, urine creatinine and dimethylamine were positively associated with the LMDA group. Conclusion: Metabolic phenotypes of pregnant women who had a high alignment with the MD were significantly different from pregnant women who had a poor alignment with the MD. The metabolite profiles aligned with reported food intake. Differences were most significant biomarkers of systemic inflammation and selected gut-microbial metabolites. This research expands our understanding of the mechanisms driving health outcomes during the perinatal period and provides additional biomarkers for investigation in pregnant women to assess potential health risks.