RESUMO
A potential source of novel biomarkers for mTBI is the kynurenine pathway (KP), a metabolic pathway of tryptophan (Trp), that is up-regulated by neuroinflammation and stress. Considering that metabolites of the KP (kynurenines) are implicated in various neuropsychiatric diseases, exploration of this pathway could potentially bridge the gap between physiological and psychological factors in the recovery process after mTBI. This study, therefore, set out to characterize the KP after mTBI and to examine associations with long-term outcome. Patients were prospectively recruited at the emergency department (ED), and blood samples were obtained in the acute phase (<24 h; N = 256) and at 1-month follow-up (N = 146). A comparison group of healthy controls (HC; N = 32) was studied at both timepoints. Trp, kynurenines, and interleukin (IL)-6 and IL-10 were quantified in plasma. Clinical outcome was measured at six months post-injury. Trp, xanthurenic acid (XA), and picolinic acid (PA) were significantly reduced in patients with mTBI relative to HC, corrected for age and sex. For Trp (d = -0.57 vs. d = -0.29) and XA (d = -0.98 vs. d = -0.32), larger effects sizes were observed during the acute phase compared to one-month follow-up, while for PA (d = -0.49 vs. d = -0.52) effect sizes remained consistent. Findings for other kynurenines (e.g., kynurenine, kynurenic acid, and quinolinic acid) were non-significant after correction for multiple testing. Within the mTBI group, lower acute Trp levels were significantly related to incomplete functional recovery and higher depression scores at 6 months post-injury. No significant relationships were found for Trp, XA, and PA with IL-6 or IL-10 concentrations. In conclusion, our findings indicate that perturbations of the plasma KP in the hyperacute phase of mTBI and 1 month later are limited to the precursor Trp, and glutamate system modulating kynurenines XA and PA. Correlations between acute reductions of Trp and unfavorable outcomes may suggest a potential substrate for pharmacological intervention.
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OBJECTIVES: Trace amines are powerful neuromodulators influencing the release and reuptake of catecholamines. These low concentrated endogenous amines impact mood, cognition, and hormone regulation. Dysregulation of trace amines have been associated with a variety of diseases, such as schizophrenia, Parkinson's disease, migraine, depression and more. Succesfull simultaneous quantification of trace amines, their precursors and metabolites would benefit both research and patient care. Since these compounds have various functional groups and are present in biological matrices with large concentration difference, their simultaneous quantification is an analytical challenge. Our goal was to develop a highly sensitive LC-MS/MS assay to simultaneously quantify trace amines, their precursors and metabolites in plasma. METHODS: Our method is based on a simple two-step in-matrix derivatization protocol: propionic anhydride (PA) and 3-Ethyl-1-[3-(dimethylamino)propyl]carbodiimide (EDC) in combination with 2,2,2-trifluoroethylamine (TFEA) followed by online solid phase extraction combined with LC-MS/MS. Fifteen metabolites can be measured simultaneously, three precursors, eight trace amines and four metabolites. Validation of this method was performed according to international validation guidelines. The pre-analytical stability of trace amines was assessed. RESULTS: This novel method was successful in quantifying trace amines, their precursors, and metabolites in plasma. Using just 50 µl human plasma, we were able to accomplish limit of quantification for 2-phenylethylamine and N-methyl-phenylethylamine of 0.2 nmol/L and 0.1 nmol/L for tyramine and n-methyltyramine. Inter-and intra-assay imprecision was < 15 % for all analytes. Stability assessment showed susceptibility of certain trace amines e.g. 2-phenylethylamine and N-methyl-phenylethylamine to enzymatic degradation in plasma. The addition of the monoamine oxidase inhibitor pargyline to plasma prevented this enzymatic degradation. CONCLUSIONS: We developed a novel LC-MS/MS method that1) uses a new double derivatization technique, 2) is automated with online SPE, 3) uses far less sample volume then previous methods and 4) detects more components in the same sample (eight trace amines, three precursors, and four metabolites) with high specificity and selectivity. Furthermore, addition of MAO A/B inhibitor prevents degradation and guarantees more accurate quantification of trace amines.
Assuntos
Aminas , Espectrometria de Massas em Tandem , Humanos , Espectrometria de Massas em Tandem/métodos , Reprodutibilidade dos Testes , Aminas/sangue , Cromatografia Líquida/métodos , Limite de Detecção , Modelos Lineares , Extração em Fase Sólida/métodosRESUMO
OBJECTIVES: Despite distinct clinical profiles, amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) patients share a remarkable portion of pathological features, with a substantial percentage of patients displaying a mixed disease phenotype. Kynurenine metabolism seems to play a role in dementia-associated neuroinflammation and has been linked to both diseases. We aimed to explore dissimilarities in kynurenine pathway metabolites in these early onset neurodegenerative disorders in a brain-region-specific manner. METHODS: Using liquid chromatography mass spectrometry (LC-MS/MS), kynurenine metabolite levels were determined in the brain samples of 98 healthy control subjects (n = 20) and patients with early onset Alzheimer's disease (EOAD) (n = 23), ALS (n = 20), FTD (n = 24) or a mixed FTD-ALS (n = 11) disease profile. RESULTS: Overall, the kynurenine pathway metabolite levels were significantly lower in patients with ALS compared to FTD, EOAD and control subjects in the frontal cortex, substantia nigra, hippocampus and neostriatum. Anthranilic acid levels and kynurenine-to-tryptophan ratios were consistently lower in all investigated brain regions in ALS compared to the other diagnostic groups. CONCLUSIONS: These results suggest that the contribution of kynurenine metabolism in neuroinflammation is lower in ALS than in FTD or EOAD and may also be traced back to differences in the age of onset between these disorders. Further research is necessary to confirm the potential of the kynurenine system as a therapeutic target in these early onset neurodegenerative disorders.
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Activation of the kynurenine pathway (KP) has been reported in patients with pulmonary arterial hypertension (PAH) undergoing PAH therapy. We aimed to determine KP-metabolism in treatment-naïve PAH patients, investigate its prognostic values, evaluate the effect of PAH therapy on KP-metabolites and identify cytokines responsible for altered KP-metabolism. KP-metabolite levels were determined in plasma from PAH patients (median follow-up 42 months) and in rats with monocrotaline- and Sugen/hypoxia-induced PH. Blood sampling of PAH patients was performed at the time of diagnosis, six months and one year after PAH therapy. KP activation with lower tryptophan, higher kynurenine (Kyn), 3-hydroxykynurenine (3-HK), quinolinic acid (QA), kynurenic acid (KA), and anthranilic acid was observed in treatment-naïve PAH patients compared with controls. A similar KP-metabolite profile was observed in monocrotaline, but not Sugen/hypoxia-induced PAH. Human lung primary cells (microvascular endothelial cells, pulmonary artery smooth muscle cells, and fibroblasts) were exposed to different cytokines in vitro. Following exposure to interleukin-6 (IL-6)/IL-6 receptor α (IL-6Rα) complex, all cell types exhibit a similar KP-metabolite profile as observed in PAH patients. PAH therapy partially normalized this profile in survivors after one year. Increased KP-metabolites correlated with higher pulmonary vascular resistance, shorter six-minute walking distance, and worse functional class. High levels of Kyn, 3-HK, QA, and KA measured at the latest time-point were associated with worse long-term survival. KP-metabolism was activated in treatment-naïve PAH patients, likely mediated through IL-6/IL-6Rα signaling. KP-metabolites predict response to PAH therapy and survival of PAH patients.
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Interleucina-6 , Cinurenina , Hipertensão Arterial Pulmonar , Receptores de Interleucina-6 , Animais , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Humanos , Hipóxia/metabolismo , Interleucina-6/metabolismo , Ácido Cinurênico/metabolismo , Cinurenina/metabolismo , Monocrotalina , Hipertensão Arterial Pulmonar/metabolismo , Hipertensão Arterial Pulmonar/patologia , Ácido Quinolínico/metabolismo , Ratos , Receptores de Interleucina-6/metabolismoRESUMO
Bile acids (BAs) are implicated in the etiology of obesity-related conditions such as non-alcoholic fatty liver disease. Differently structured BA species display variable signaling activities via farnesoid X receptor (FXR) and Takeda G protein-coupled BA receptor 1 (TGR5). This study profiles plasma and fecal BAs and plasma 7α-hydroxy-4-cholesten-3-one (C4) in 297 persons with obesity, identifies underlying genetic and microbial determinants, and establishes BA correlations with liver fat and plasma lipid parameters. We identify 27 genetic associations (p < 5 × 10-8) and 439 microbial correlations (FDR < 0.05) for 50 BA entities. Additionally, we report 111 correlations between BA and 88 lipid parameters (FDR < 0.05), mainly for C4 reflecting hepatic BA synthesis. Inter-individual variability in the plasma BA profile does not reflect hepatic BA synthetic pathways, but rather transport and metabolism within the enterohepatic circulation. Our study reveals genetic and microbial determinants of BAs in obesity and their relationship to disease-relevant lipid parameters that are important for the design of personalized therapies targeting BA-signaling pathways.
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Ácidos e Sais Biliares/fisiologia , Metabolismo dos Lipídeos/genética , Fígado/patologia , Síndrome Metabólica/fisiopatologia , Obesidade/fisiopatologia , Idoso , Idoso de 80 Anos ou mais , Humanos , Pessoa de Meia-IdadeRESUMO
BACKGROUND: People with HIV (PWH) taking antiretroviral therapy (ART) may experience weight gain, dyslipidemia, increased risk of non-AIDS comorbidities, and long-term alteration of the gut microbiota. Both low CD4/CD8 ratio and chronic inflammation have been associated with changes in the gut microbiota of PWH. The antidiabetic drug metformin has been shown to improve gut microbiota composition while decreasing weight and inflammation in diabetes and polycystic ovary syndrome. Nevertheless, it remains unknown whether metformin may benefit PWH receiving ART, especially those with a low CD4/CD8 ratio. METHODS: In the Lilac pilot trial, we recruited 23 nondiabetic PWH receiving ART for more than 2 years with a low CD4/CD8 ratio (<0.7). Blood and stool samples were collected during study visits at baseline, after a 12-week metformin treatment, and 12 weeks after discontinuation. Microbiota composition was analyzed by 16S rDNA gene sequencing, and markers of inflammation were assessed in plasma. RESULTS: Metformin decreased weight in PWH, and weight loss was inversely correlated with plasma levels of the satiety factor GDF-15. Furthermore, metformin changed the gut microbiota composition by increasing the abundance of anti-inflammatory bacteria such as butyrate-producing species and the protective Akkermansia muciniphila. CONCLUSIONS: Our study provides the first evidence that a 12-week metformin treatment decreased weight and favored anti-inflammatory bacteria abundance in the microbiota of nondiabetic ART-treated PWH. Larger randomized placebo-controlled clinical trials with longer metformin treatment will be needed to further investigate the role of metformin in reducing inflammation and the risk of non-AIDS comorbidities in ART-treated PWH.
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Plasma-free metanephrines and catecholamines are essential markers in the biochemical diagnosis and follow-up of neuroendocrine tumors and inborn errors of metabolism. However, their low circulating concentrations (in the nanomolar range) and poor fragmentation characteristics hinder facile simultaneous quantification by liquid chromatography and tandem mass spectrometry (LC-MS/MS). Here, we present a sensitive and simple matrix derivatization procedure using propionic anhydride that enables simultaneous quantification of unconjugated l-DOPA, catecholamines, and metanephrines in plasma by LC-MS/MS. Dilution of propionic anhydride 1:4 (v/v) in acetonitrile in combination with 50 µL of plasma resulted in the highest mass spectrometric response. In plasma, derivatization resulted in stable derivatives and increased sensitivity by a factor of 4-30 compared with a previous LC-MS/MS method for measuring plasma metanephrines in our laboratory. Furthermore, propionylation increased specificity, especially for 3-methoxytyramine, by preventing interference from antihypertensive medication (ß-blockers). The method was validated according to international guidelines and correlated with a hydrophilic interaction LC-MS/MS method for measuring plasma metanephrines (R2 > 0.99) and high-performance liquid chromatography with an electrochemical detection method for measuring plasma catecholamines (R2 > 0.85). Reference intervals for l-DOPA, catecholamines, and metanephrines in n = 115 healthy individuals were established. Our work shows that analytes in the subnanomolar range in plasma can be derivatized in situ without any preceding sample extraction. The developed method shows improved sensitivity and selectivity over existing methods and enables simultaneous quantification of several classes of amines.
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Catecolaminas/sangue , Metanefrina/sangue , Espectrometria de Massas em Tandem/métodos , Catecolaminas/isolamento & purificação , Catecolaminas/normas , Cromatografia Líquida de Alta Pressão/normas , Dopamina/análogos & derivados , Dopamina/sangue , Dopamina/isolamento & purificação , Dopamina/normas , Humanos , Levodopa/sangue , Levodopa/isolamento & purificação , Levodopa/normas , Limite de Detecção , Metanefrina/isolamento & purificação , Metanefrina/normas , Valores de Referência , Extração em Fase Sólida , Espectrometria de Massas em Tandem/normasRESUMO
Exploring the neurochemical continuum between frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) with respect to monoamines and kynurenines in cerebrospinal fluid (CSF) and serum, may be useful to identify possible new research/therapeutic targets. Hence, we analysed monoamines and kynurenines in CSF and serum derived from patients with FTD (n = 39), ALS (n = 23), FTD-ALS (n = 4) and age-matched control subjects (n = 26), using reversed-phase ultra-high performance liquid chromatography (RP-UHPLC) with electrochemical detection (ECD) and liquid chromatography tandem mass spectrometry, respectively. We noted a shared dopaminergic disturbance in FTD and ALS when compared to CONTR, with significantly increased serum DA levels and decreased DOPAC concentrations, as well as decreased DOPAC/DA ratios in both disease groups. In CSF, significantly reduced DOPAC concentrations in FTD and ALS were observed as well. Here, a significant increase in DA levels and decrease in DOPAC/DA ratios was only found in FTD relative to CONTR. With respect to the kynurenine pathway (KP), we only found decreased HK/XA ratios, indicative for vitamin B6 status, in serum of ALS subjects compared to FTD. The dopaminergic commonalities observed in FTD and ALS might relate to a disturbance of dopaminergic nerve terminals in projection areas of the substantia nigra and/or ventral tegmental area, although these findings should first be confirmed in brain tissue. Lastly, based on the results of this work, the KP does not hold promise as a research/therapeutic target in FTD and ALS.
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Esclerose Lateral Amiotrófica/metabolismo , Monoaminas Biogênicas/metabolismo , Demência Frontotemporal/metabolismo , Cinurenina/metabolismo , Idoso , Esclerose Lateral Amiotrófica/sangue , Esclerose Lateral Amiotrófica/líquido cefalorraquidiano , Monoaminas Biogênicas/sangue , Monoaminas Biogênicas/líquido cefalorraquidiano , Biomarcadores/sangue , Biomarcadores/líquido cefalorraquidiano , Feminino , Demência Frontotemporal/sangue , Demência Frontotemporal/líquido cefalorraquidiano , Humanos , Cinurenina/sangue , Cinurenina/líquido cefalorraquidiano , Masculino , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
The kynurenine (Kyn) pathway, which regulates neuroinflammation and N-methyl-d-aspartate receptor activation, is implicated in Parkinson's disease (PD) and Alzheimer's disease (AD). Age-related changes in Kyn metabolism and altered cerebral Kyn uptake along large neutral amino acid transporters, could contribute to these diseases. To gain further insight into the role and prognostic potential of the Kyn pathway in PD and AD, we investigated systemic and cerebral Kyn metabolite production and estimations of their transporter-mediated uptake in the brain. Kyn metabolites and large neutral amino acids were retrospectively measured in serum and cerebrospinal fluid (CSF) of clinically well-characterized PD patients (n = 33), AD patients (n = 33), and age-matched controls (n = 39) using solid-phase extraction-liquid chromatographic-tandem mass spectrometry. Aging was disease independently associated with increased Kyn, kynurenic acid and quinolinic acid in serum and CSF. Concentrations of kynurenic acid were reduced in CSF of PD and AD patients (p = 0.001; p = 0.002) but estimations of Kyn brain uptake did not differ between diseased and controls. Furthermore, serum Kyn and quinolinic acid levels strongly correlated with their respective content in CSF and Kyn in serum negatively correlated with AD disease severity (p = 0.002). Kyn metabolites accumulated with aging in serum and CSF similarly in PD patients, AD patients, and control subjects. In contrast, kynurenic acid was strongly reduced in CSF of PD and AD patients. Differential transporter-mediated Kyn uptake is unlikely to majorly contribute to these cerebral Kyn pathway disturbances. We hypothesize that the combination of age- and disease-specific changes in cerebral Kyn pathway activity could contribute to reduced neurogenesis and increased excitotoxicity in neurodegenerative disease.
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Doença de Alzheimer/metabolismo , Cinurenina/metabolismo , Doença de Parkinson/metabolismo , Idoso , Envelhecimento/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Background: Low plasma concentrations of pyridoxal 5'-phosphate (PLP) are common in renal transplant recipients (RTRs) and confer increased risk of long-term mortality. To our knowledge, it is not known whether low plasma PLP concentrations have functional (i.e., intracellular) consequences and, if so, whether such consequences are associated with increased risk of mortality.Objectives: We assessed the association of plasma PLP with functional vitamin B-6 status and explored the potential association of functional vitamin B-6 status with long-term mortality in RTRs.Design: In a longitudinal cohort of 678 stable RTRs with a median follow-up of 5.3 y (IQR: 4.8-6.1 y) and 297 healthy controls, PLP, plasma 3-hydroxykynurenine (3-HK), and xanthurenic acid (XA) were analyzed via validated assays. PLP was used as direct biomarker for vitamin B-6 status, and the 3-HK:XA ratio was used as functional biomarker of vitamin B-6 status with a higher ratio reflecting worse functional vitamin B-6 status.Results: Median PLP, 3-HK, and XA concentrations were 41 nmol/L (IQR: 29-60 nmol/L), 40.1 nmol/L (IQR: 33.0-48.0 nmol/L), and 19.1 nmol/L (IQR: 14.5-24.9 nmol/L), respectively, in healthy controls compared with 29 nmol/L (IQR: 17-50 nmol/L), 61.5 nmol/L (IQR: 45.6-86.5 nmol/L), and 25.5 nmol/L (IQR: 17.2-40.0 nmol/L), respectively, in RTRs (all P < 0.001). RTRs had a higher median 3-HK:XA ratio (2.38; IQR: 1.68-3.49) than did healthy controls (2.13; IQR: 1.63-2.71) (P < 0.05). In RTRs, the 3-HK:XA ratio was inversely associated with plasma PLP (ß = -0.21, P < 0.001). Moreover, a higher 3-HK:XA ratio was independently associated with increased risk of all-cause mortality (HR per SD increment: 1.30; 95% CI: 1.13, 1.49), cancer mortality (HR per SD increment: 1.47; 95% CI: 1.12, 1.95), and infectious disease mortality (HR per SD increment: 1.50; 95% CI: 1.21, 1.86) in RTRs.Conclusions: Vitamin B-6-deficient RTRs have a worse functional vitamin B-6 status than do healthy controls and vitamin B-6-sufficient RTRs. Worse functional vitamin B-6 status in RTRs is independently associated with an increased risk of mortality particularly because of cancer and infectious disease. This trial was registered at clinicaltrials.gov as NCT02811835.
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Infecções/mortalidade , Transplante de Rim/efeitos adversos , Cinurenina/análogos & derivados , Neoplasias/mortalidade , Fosfato de Piridoxal/sangue , Deficiência de Vitamina B 6/complicações , Xanturenatos/sangue , Adulto , Idoso , Biomarcadores/sangue , Causas de Morte , Estudos de Coortes , Feminino , Humanos , Rim/cirurgia , Nefropatias/cirurgia , Cinurenina/sangue , Masculino , Pessoa de Meia-Idade , Estado Nutricional , Fatores de Risco , Vitamina B 6/sangue , Deficiência de Vitamina B 6/sangue , Complexo Vitamínico B/sangueRESUMO
BACKGROUND: Function of the hypothalamus-pituitary-adrenal (HPA) axis has been associated with several somatic and psychiatric health problems. The amount of free cortisol excreted in the urine during 24h (24-h UFC) has often been used as a proxy for HPA-axis function. Reference values for 24-h UFC and their stability in the short and long term, as well as sources of variability, are largely lacking. METHODS: This study was performed in a general population cohort. Participants collected 24-h UFC on two consecutive days (T1), and repeated this collection approximately 2 years later (T2). Cortisol in urine was measured using LC-MS/MS. Height and weight were measured at the research facilities; glomerular filtration rate was estimated using creatinine clearance. Psychological distress (General Health Questionnaire), smoking, alcohol use and exercise were measured by means of questionnaires. RESULTS: 24-h UFC stability on a day-to-day basis was 0.69 (T1, N=1192) and 0.72 (T2, N=963) (both p<0.001). Long-term stability as indicated by correlation between 2-day averages of T1 and T2 was 0.60 (N=972, p<0.001). Multivariable linear regression analysis revealed that 24-h UFC was predicted by urine volume (standardized beta 0.282 (T1, N=1556) and 0.276 (T2, N=1244); both p<0.001) and glomerular filtration rate (standardized beta 0.137 (T1) and 0.179 (T2); both p<0.001), while also sex explained a small part (standardized beta for female sex -0.057 (T1) and -0.080 (T2); both p<0.05). CONCLUSION: 24-h UFC is moderately stable both in the short and the long term. The effects of urine volume and glomerular filtration rate on 24-h UFC are much stronger than those of sex.
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Ritmo Circadiano , Projetos de Pesquisa Epidemiológica , Hidrocortisona/urina , Coleta de Urina/métodos , Adulto , Estudos de Coortes , Estabilidade de Medicamentos , Estudos Epidemiológicos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Variações Dependentes do Observador , Preservação Biológica/normas , Projetos de Pesquisa , Tamanho da Amostra , Coleta de Urina/normasRESUMO
BACKGROUND & AIMS: Glucocorticoids, produced by the adrenal gland under control of the hypothalamic-pituitary-adrenal axis, exert their metabolic actions largely via activation of the glucocorticoid receptor (GR). Synthetic glucocorticoids are widely used as anti-inflammatory and immunosuppressive drugs but their application is hampered by adverse metabolic effects. Recently, it has been shown that GR may regulate several genes involved in murine bile acid (BA) and cholesterol metabolism, yet the physiological relevance hereof is controversial. The aim of this study is to provide a mechanistic basis for effects of prednisolone on BA and cholesterol homeostasis in mice. METHODS: Male BALB/c mice were treated with prednisolone (12.5mg/kg/day) for 7days by subcutaneous implantation of slow-release pellets, followed by extensive metabolic profiling. RESULTS: Sustained prednisolone treatment induced the expression of the apical sodium-dependent bile acid transporter (Asbt) in the ileum, which stimulated BA absorption. This resulted in elevated plasma BA levels and enhanced biliary BA secretion. Concomitantly, both biliary cholesterol and phospholipid secretion rates were increased. Enhanced BA reabsorption suppressed hepatic BA synthesis, as evident from hepatic gene expression, reduced plasma C4 levels and reduced fecal BA loss. Plasma HDL cholesterol levels were elevated in prednisolone-treated mice, which likely contributed to the stimulated flux of cholesterol from intraperitoneally injected macrophage foam cells into feces. CONCLUSIONS: Sustained prednisolone treatment increases enterohepatic recycling of BA, leading to elevated plasma levels and reduced synthesis in the absence of cholestasis. Under these conditions, prednisolone promotes macrophage-derived reverse cholesterol transport.
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Ácidos e Sais Biliares/metabolismo , Colesterol/metabolismo , Circulação Êntero-Hepática , Transportadores de Ânions Orgânicos Dependentes de Sódio/biossíntese , Prednisolona/farmacologia , Simportadores/biossíntese , Animais , Transporte Biológico , Homeostase , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
BACKGROUND & AIMS: Obesity has been associated with changes in the composition and function of the intestinal microbiota. Modulation of the microbiota by antibiotics also alters bile acid and glucose metabolism in mice. Hence, we hypothesized that short term administration of oral antibiotics in humans would affect fecal microbiota composition and subsequently bile acid and glucose metabolism. METHODS: In this single blinded randomized controlled trial, 20 male obese subjects with metabolic syndrome were randomized to 7 days of amoxicillin 500 mg t.i.d. or 7 days of vancomycin 500 mg t.i.d. At baseline and after 1 week of therapy, fecal microbiota composition (Human Intestinal Tract Chip phylogenetic microarray), fecal and plasma bile acid concentrations as well as insulin sensitivity (hyperinsulinemic euglycemic clamp using [6,6-(2)H2]-glucose tracer) were measured. RESULTS: Vancomycin reduced fecal microbial diversity with a decrease of gram-positive bacteria (mainly Firmicutes) and a compensatory increase in gram-negative bacteria (mainly Proteobacteria). Concomitantly, vancomycin decreased fecal secondary bile acids with a simultaneous postprandial increase in primary bile acids in plasma (p<0.05). Moreover, changes in fecal bile acid concentrations were predominantly associated with altered Firmicutes. Finally, administration of vancomycin decreased peripheral insulin sensitivity (p<0.05). Amoxicillin did not affect any of these parameters. CONCLUSIONS: Oral administration of vancomycin significantly impacts host physiology by decreasing intestinal microbiota diversity, bile acid dehydroxylation and peripheral insulin sensitivity in subjects with metabolic syndrome. These data show that intestinal microbiota, particularly of the Firmicutes phylum contributes to bile acid and glucose metabolism in humans. This trial is registered at the Dutch Trial Register (NTR2566).