Serum Concentrations Of Metabolites Research Articles

“Kynurenine switch” and obesity

Aim. To assess the concentrations of bacterial and eukaryotic metabolites mainly involved in indole, kynurenine, and serotonin pathways of tryptophan metabolism in a cohort of patients with obesity. Materials and methods. Using high-performance liquid chromatography with mass spectrometric detection, the concentrations of several serum metabolites, such as kynurenine, kynurenic acid, anthranilic acid, xanthurenic acid, quinolinic acid, 5-hydroxyindole-3-acetate, tryptamine, serotonin, indole-3-lactate, indole-3-acetate, indole-3- butyrate, indole-3-carboxaldehyde, indole-3-acrylate, and indole-3-propionate, were analyzed in a cohort of obese patients compared with healthy volunteers.Results. It was found that serum levels of tryptophan metabolites of microbial and eukaryotic origin were significantly increased in obese patients. Therefore, the concentration of kynurenine in the blood serum in obese patients was 2,413 ± 855 nmol / l, while in healthy volunteers of the same age group, the level of kynurenine in the blood serum was 2,122 ± 863 nmol / l. In obese patients, two acids formed due to kynurenine metabolism; the concentrations of kynurenic and quinolinic acids were increased in the blood serum. The concentration of kynurenic acid in the blood serum in obese patients was 21.1 ± 9.26 nmol / l, and in healthy patients, it was 16.8 ± 8.37 nmol / l. At the same time, the level of quinolinic acid in the blood serum in obese patients was 73.1 ± 54.4 nmol / l and in healthy volunteers – 56.8 ± 34.1 nmol / l. Normally, the level of quinolinic acid is 3.4 times higher than the concentration of kynurenic acid, and in case of obesity, there is a comparable increase in these acids in the blood serum.From indole derivatives, mainly of microbial origin, the concentrations of indole-3-lactate, indole-3-butyrate, and indole-3-acetate were significantly increased in the blood serum of obese patients. In obese patients, the serum concentration of 5-hydroxyindole-3-acetate was elevated to 74.6 ± 75.8 nmol / l (in healthy volunteers – 59.4 ± 36.6 nmol / l); indole-3-lactate – to 523 ± 251 nmol / l (in healthy volunteers – 433 ± 208 nmol / l); indole-3-acetate – to 1,633 ± 1,166 nmol / l (in healthy volunteers – 1,186 ± 826 nmol / l); and indole-3-butyrate – to 4.61 ± 3.31 nmol / l (in healthy volunteers – 3.85 ± 2.51 nmol / l).Conclusion. In case of obesity, the utilization of tryptophan was intensified by both the microbiota population and the macroorganism. It was found that obese patients had higher concentrations of kynurenine, quinolinic and kynurenic acids, indole-3-acetate, indole-3-lactate, indole-3-butyrate, and 5-hydroxyindole-3-acetate. Apparently, against the background of increased production of proinflammatory cytokines by adipocytes in obese patients, the “kynurenine switch” was activated which contributed to subsequent overproduction of tryptophan metabolites involved in the immune function of the macroorganism.

P4‐7: Associations between long‐term exposure to PM10 and NO2 and serum metabolite concentration in CODA cohort

P4‐7: Associations between long‐term exposure to PM10 and NO2 and serum metabolite concentration in CODA cohort

Prevotella copri increases fat accumulation in pigs fed with formula diets

BackgroundExcessive fat accumulation of pigs is undesirable, as it severely affects economic returns in the modern pig industry. Studies in humans and mice have examined the role of the gut microbiome in host energy metabolism. Commercial Duroc pigs are often fed formula diets with high energy and protein contents. Whether and how the gut microbiome under this type of diet regulates swine fat accumulation is largely unknown.ResultsIn the present study, we systematically investigated the correlation of gut microbiome with pig lean meat percentage (LMP) in 698 commercial Duroc pigs and found that Prevotella copri was significantly associated with fat accumulation of pigs. Fat pigs had significantly higher abundance of P. copri in the gut. High abundance of P. copri was correlated with increased concentrations of serum metabolites associated with obesity, e.g., lipopolysaccharides, branched chain amino acids, aromatic amino acids, and the metabolites of arachidonic acid. Host intestinal barrier permeability and chronic inflammation response were increased. A gavage experiment using germ-free mice confirmed that the P. copri isolated from experimental pigs was a causal species increasing host fat accumulation and altering serum metabolites. Colon, adipose tissue, and muscle transcriptomes in P. copri-gavaged mice indicated that P. copri colonization activated host chronic inflammatory responses through the TLR4 and mTOR signaling pathways and significantly upregulated the expression of the genes related to lipogenesis and fat accumulation, but attenuated the genes associated with lipolysis, lipid transport, and muscle growth.ConclusionsTaken together, the results proposed that P. copri in the gut microbial communities of pigs fed with commercial formula diets activates host chronic inflammatory responses by the metabolites through the TLR4 and mTOR signaling pathways, and increases host fat deposition significantly. The results provide fundamental knowledge for reducing fat accumulation in pigs through regulating the gut microbial composition.8i_LmNceHRNfJRY2rW1K4pVideo

O-234 Using serum metabolomics to identify biomarkers of viable early, intrauterine pregnancy: an untargeted 1H NMR-based approach

Abstract Study question Which metabolites are associated with a viable intrauterine pregnancy (VIUP) when compared to other early pregnancy outcomes (failed intrauterine and ectopic pregnancies)? Summary answer Serum levels of four metabolites (phenylalanine, alanine, glutamate and glutamine) were significantly altered in VIUPs compared to other early pregnancy outcomes. What is known already Around 10% of all intrauterine pregnancies are lost in the first trimester. A further 1-2% of pregnancies are located outside the endometrial cavity; these ectopic pregnancies are the leading cause of maternal mortality in the first trimester of gestation. Early miscarriages may also cause significant morbidity when bleeding or infection occurs. The symptoms of miscarriages and ectopic pregnancy are often similar (pain and bleeding), however, such symptoms are also common in VIUPs. To date, no biomarkers have been identified to differentiate VIUPs from non-viable and ectopic pregnancies. Study design, size, duration This is a prospective cohort study that included 332 pregnant women at less than ten weeks of gestation, who attended the early pregnancy assessment unit (EPAU) at Liverpool Women’s Hospital with pain and/or bleeding. Participants/materials, setting, methods Blood samples were collected from the 332 pregnant women prior to final clinical diagnosis of pregnancy outcome. Serum samples were subjected to NMR metabolomics profiling (14 spectra that did not meet the recommended minimum reporting standards were removed from subsequent analysis). 1D 1H-NMR spectra were acquired at 37 °C on a 700 MHz spectrometer. Relative metabolite abundances underwent statistical analysis using MetaboAnalyst 5.0 (p-value FDR adjusted). Main results and the role of chance Final pregnancy outcomes were as follows: one hydatidiform mole (0.3%), 48 ectopic pregnancies (14.4%), three pregnancies of unknown location (PULs, 0.9%), 78 failed pregnancies of unknown location (FPULs, 23.4%), 47 miscarriages (14.1%), two vanishing twin pregnancies (0.6%) and 153 VIUPs (45.8%). Due to small sample numbers, the hydatidiform mole, PULs and vanishing twin pregnancies were excluded from further analysis. To compare VIUPs to other pregnancy outcomes, ectopic pregnancies, FPULs and miscarriages were grouped together. Univariate analysis of serum metabolite concentrations identified four metabolites (phenylalanine, alanine, glutamate and glutamine) as significantly different in VIUPs compared to other pregnancy outcomes. Multivariate partial least squared discriminant analysis provided only weak correlation between the serum metabolome and pregnancy outcome. In summary, we have identified differences in the metabolome of women with VIUPs compared to other common pregnancy outcomes, which may provide diagnostic utility. Limitations, reasons for caution In this study, women with VIUPs presented with pain and/or bleeding. The presence of symptoms may influence the metabolome of this group versus VIUPs without symptoms, thus limiting the translation of our findings. Furthermore, environmental factors were not controlled (e.g. fasting status), making it likely that cohort heterogeneity was enhanced. Wider implications of the findings This study identifies a metabolite profile associated with VIUPs. These findings may be useful in the development of a diagnostic test to confirm VIUPs and thus exclude potentially life-threatening pregnancy outcomes. Such a test would be invaluable in clinical emergencies. Trial registration number NA

A Cross-Platform Metabolomics Comparison Identifies Serum Metabolite Signatures of Liver Fibrosis Progression in Chronic Hepatitis C Patients.

Metabolomics offers new insights into disease mechanisms that is enhanced when adopting orthogonal instrumental platforms to expand metabolome coverage, while also reducing false discoveries by independent replication. Herein, we report the first inter-method comparison when using multisegment injection-capillary electrophoresis-mass spectrometry (MSI-CE-MS) and nuclear magnetic resonance (NMR) spectroscopy for characterizing the serum metabolome of patients with liver fibrosis in chronic hepatitis C virus (HCV) infection (n = 20) and non-HCV controls (n = 14). In this study, 60 and 30 serum metabolites were detected frequently (>75%) with good technical precision (median CV < 10%) from serum filtrate samples (n = 34) when using standardized protocols for MSI-CE-MS and NMR, respectively. Also, 20 serum metabolite concentrations were consistently measured by both methods over a 500-fold concentration range with an overall mean bias of 9.5% (n = 660). Multivariate and univariate statistical analyses independently confirmed that serum choline and histidine were consistently elevated (p < 0.05) in HCV patients with late-stage (F2-F4) as compared to early-stage (F0-F1) liver fibrosis. Overall, the ratio of serum choline to uric acid provided optimal differentiation of liver disease severity (AUC = 0.848, p = 0.00766) using a receiver operating characteristic curve, which was positively correlated with liver stiffness measurements by ultrasound imaging (r = 0.606, p = 0.0047). Moreover, serum 5-oxo-proline concentrations were higher in HCV patients as compared to non-HCV controls (F = 4.29, p = 0.0240) after adjustment for covariates (age, sex, BMI), indicative of elevated oxidative stress from glutathione depletion with the onset and progression of liver fibrosis. Both instrumental techniques enable rapid yet reliable quantification of serum metabolites in large-scale metabolomic studies with good overlap for biomarker replication. Advantages of MSI-CE-MS include greater metabolome coverage, lower operating costs, and smaller sample volume requirements, whereas NMR offers a robust platform supported by automated spectral and data processing software.

N-6 Polyunsaturated fatty acids in the feeding of late gestation hair ewes: the effects on thermoregulation, growth, and metabolism of heat-stressed growing lambs.

The objective was to determine the effects of feeding soybean oil (SBO), an ingredient rich in n-6 polyunsaturated fatty acids (PUFA), to late gestation hair ewes on physiological responses, feedlot performance, and serum metabolite and electrolyte concentrations of their growing ewe lambs under outdoor heat stress conditions. Twenty-four Dorper × Pelibuey ewe lambs weaned (body weight = 21.5 ± 0.2 kg, age= 2 months, and multiple birth) born from ewes fed 0, 30, or 60 mg of SBO/kg dry matter (DM) during late gestation were selected (n = 8/treatment) to conduct a 30-day feeding trial during the summer season of a desert region (temperature = 34 °C and temperature-humidity index = 35 units). While rectal temperature was unaffected in any daytime, respiratory rate in the afternoon quadratically increased (P = 0.05) as the SBO levels increased from 0 to 60 mg/kg DM in the maternal diet. Final weight, average daily gain, and feed efficiency linearly increased (P = 0.04) with increasing levels of SBO. Body surface temperatures and serum concentration of glucose, cholesterol, triglyceride, total protein, urea, sodium, potassium, and chlorine did not vary by the SBO inclusion in the maternal diet. In conclusion, feeding late gestation hair ewes with source rich in n-6 PUFA appears to be an effective maternal nutritional strategy to improve post-weaning growth without compromising the thermoregulatory ability of their growing offspring under a heat stress environment.

Relationships among ß-hydroxybutyrate, calcium and non-esterified fatty acids in blood with milk yield losses at early lactation

The objectives were to study the associations of concentrations of β-hydroxybutyrate acid (BHBA), calcium (Ca2+), and non-esterified fatty acids (NEFA) in blood serum 7 d prepartum with losses in milk yield (MY) and metabolic dysfunctions at seven and 14 d of lactation. Three hundred and thirty-six (336) Holstein-Friesian (780 ± 36 kg BW; which had lactated more than twice) were sampled by coccygeal venipuncture, 7 d before, and 7 and 14 d after parturition. For each sample and metabolite serum concentrations were stratified in thresholds and related to MY. When BHBA levels were high 7 d before parturition and were related to MY at d-7 postpartum, it was observed that 11.00 % of the cows lost 0.370 kg d-1 of milk. In contrast, no relationship was observed between BHBA prepartum and MY on d-14 of lactation. It was not observed any association between high NEFA and low Ca2+ levels prepartum and MY. NEFA concentrations ≥ 0.5 mmol L-1 on d-7 before calving were 7.6 more susceptible for lameness incidence (P≤ 0.01), and when BHBA ≥ 0.8 mmol L-1 cows were 2.4 times more likely to develop ketosis (P≤0.05) in the first 60 d in milk. In brief, data indicate that a high proportion of cows are above the thresholds of β-hydroxybutyrate and non-esterified fatty acids, and are also deficient in calcium, when determined one week before parturition. The risk thresholds for each metabolite were not associated with the amount of milk lost at d-14 after calving.

Metabolomic Study of High-Fat Diet-Induced Obese (DIO) and DIO Plus CCl4-Induced NASH Mice and the Effect of Obeticholic Acid

The pathophysiology of nonalcoholic fatty liver disease (NAFLD) is a complex process involving metabolic and inflammatory changes in livers and other organs, but the pathogenesis is still not well clarified. Two mouse models were established to study metabolic alteration of nonalcoholic fatty liver and nonalcoholic steatohepatitis, respectively. The concentrations of metabolites in serum, liver and intestine content were measured by the AbsoluteIDQ® p180 Kit (Biocrates Life Sciences, Innsbruck, Austria). Multivariate statistical methods, pathway analysis, enrichment analysis and correlation analysis were performed to analyze metabolomic data. The metabolic characteristics of liver, serum and intestine content could be distinctly distinguished from each group, indicating the occurrence of metabolic disturbance. Among them, metabolic alteration of liver and intestine content was more significant. Based on the metabolic data of liver, 19 differential metabolites were discovered between DIO and control, 12 between DIO-CCl4 and DIO, and 47 between DIO-CCl4 and normal. These metabolites were mainly associated with aminoacyl-tRNA biosynthesis, nitrogen metabolism, lipid metabolism, glyoxylate and dicarboxylate metabolism, and amino metabolism. Further study revealed that the intervention of obeticholic acid (OCA) could partly reverse the damage of CCl4. The correlation analysis of metabolite levels and clinical parameters showed that phosphatidylcholines were negatively associated with serum alanine aminotransferase, aspartate aminotransferase, NAFLD activity score, and fibrosis score, while lysophosphatidylcholines, sphingomyelins, amino acids, and acylcarnitines shared the reverse pattern. Our study investigated metabolic alteration among control, NAFLD model, and OCA treatment groups, providing preclinical information to understand the mechanism of NAFLD and amelioration of OCA.

Association of Serum Metabolites With Frailty in Community-Dwelling Older Adults: The Framingham Offspring Study

Abstract Objectives Metabolomic markers may provide deeper insight into the underlying biology of frailty. Yet, little data exist characterizing metabolomic correlates of frailty. The objective of this cross-sectional study was to determine the association of serum metabolites with frailty. Methods We evaluated 2187 participants from the Framingham Offspring Study (mean age; 62y; 53% women) with metabolomic and frailty data. Metabolites (n = 217) were quantified using liquid chromatography/tandem mass spectrometry (1990–95). Frailty was assessed using Fried's frailty phenotype (1998–01). Multivariable logistic regression models were used to relate individual metabolites with odds of frailty. In 1,815 participants, we created a metabolomic signature: sum of (serum metabolite concentration*β coefficient). Metabolites were included based on the strength of their individual association with frailty (FDR ≤ 0.05). Logistic regression was used to relate the metabolomic signature with odds of frailty. C-statistics determined if the metabolomic signature improved the prediction of frailty beyond a standard model. Models were adjusted for age, sex, BMI, and smoking. Results Adjusting for age and sex, 24 metabolites were significantly associated with frailty (22 positive and 2 negative; all FDR ≤ 0.05). A majority of metabolites were lipids (n = 19) of the phosphatidylcholine (n = 7) and triacylglycerol (n = 10) sub-classes with high degree of saturation (n = 13 with &amp;lt;2 carbon double-bonds). Additional metabolites have roles in amino acid and cellular metabolism (lactate, asparagine, phosphocreatine, glycerol, and isocitrate). In an age and sex-adjusted model, a weighted signature of the 24 metabolites was associated with increased odds of frailty (OR: 2.0, CI: 1.5, 2.7). C-statistics to predict frailty improved beyond a standard model by the addition of the weighted metabolomic signature (C-statistics: 0.70 vs 0.61). Results were similar with further adjustment for BMI and smoking. Conclusions Frailty was associated with 24 distinct metabolites and with a metabolomic signature in this study of older adults. These findings may inform future investigations on the progression of frailty and warrant replication in independent populations. Funding Sources AHA and Boston Pepper Center OAIC

Genetic and Metabolite Variability in One-Carbon Metabolism Applied to an Insulin Resistance Model in Patients With Schizophrenia Receiving Atypical Antipsychotics.

Background: Patients with schizophrenia are at high risk of pre-mature mortality due to cardiovascular disease (CVD). Our group has completed studies in pharmacogenomics and metabolomics that have independently identified perturbations in one-carbon metabolism as associated with risk factors for CVD in this patient population. Therefore, this study aimed to use genetic and metabolomic data to determine the relationship between folate pharmacogenomics, one-carbon metabolites, and insulin resistance as measured using the homeostatic model assessment for insulin resistance (HOMA-IR) as a marker of CVD.Methods: Participants in this pilot analysis were on a stable atypical antipsychotic regimen for at least 6 months, with no diabetes diagnosis or use of antidiabetic medications. Participant samples were genotyped for MTHFR variants rs1801131 (MTHFR A1298C) and rs1801133 (MTHFR C677T). Serum metabolite concentrations were obtained with NMR. A least squares regression model was used to predict log(HOMA-IR) values based on the following independent variables: serum glutamate, glycine, betaine, serine, and threonine concentrations, and carrier status of the variant alleles for the selected genotypes.Results: A total of 67 participants were included, with a median age of 47 years old (IQR 42–52), 39% were female, and the median BMI was 30.3 (IQR 26.3–37.1). Overall, the model demonstrated an ability to predict log(HOMA-IR) values with an adjusted R2 of 0.44 and a p-value of < 0.001. Glutamate, threonine, and carrier status of the MTHFR 1298 C or MTHFR 677 T allele were positively correlated with log(HOMA-IR), whereas glycine, serine, and betaine concentrations trended inversely with log(HOMA-IR). All factors included in this final model were considered as having a possible effect on predicting log(HOMA-IR) as measured with a p-value < 0.1.Conclusions: Presence of pharmacogenomic variants that decrease the functional capacity of the MTHFR enzyme are associated with increased risk for cardiovascular disease, as measured in this instance by log(HOMA-IR). Furthermore, serine, glycine, and betaine concentrations trended inversely with HOMA-IR, suggesting that increased presence of methyl-donating groups is associated with lower measures of insulin resistance. Ultimately, these results will need to be replicated in a significantly larger population.

Alterations of Serum Metabolites and Fecal Microbiota Involved in Ewe Follicular Cyst.

While the interactions of the gut microbiome and blood metabolome have been widely studied in polycystic ovary disease in women, follicular cysts of ewes have been scarcely investigated using these methods. In this study, the fecal microbiome and serum metabolome were used to compare between ewes diagnosed with ovarian cystic follicles and ewes with normal follicles, to investigate alterations of the fecal bacterial community composition and metabolic parameters in relation to follicular cystogenesis. Ewes from the same feeding and management system were diagnosed with a follicular cyst (n = 6) or confirmed to have normal follicles (n = 6) by using a B-mode ultrasound scanner. Blood serum and fresh fecal samples of all ewes were collected and analyzed. The α-diversity of fecal microbiome did not differ significantly between follicular cyst ewes and normal follicle ewes. Three genera (Bacteroides, Anaerosporobacter, and Angelakisella) were identified and their balance differentiated between follicular cyst and normal follicle ewes. Alterations of several serum metabolite concentrations, belonging to lipids and lipid-like molecules, organic acids and derivatives, organic oxygen compounds, benzenoids, phenylpropanoids and polyketides, and organoheterocyclic compounds, were associated with the presence of a follicular cyst. Correlation analysis between fecal bacterial communities and serum metabolites indicated a positive correlation between Anaerosporobacter and several fatty acids, and a negative correlation between Bacteroides and L-proline. These observations provide new insights for the complex interactions of the gut microbiota and the host serum lipid profiles, and support gut microbiota as a potential strategy to treat and prevent follicular cysts in sheep.

Analysis of six pyrethroid insecticide metabolites in cord serum using a novel gas chromatography-ion trap mass spectrometry method

Pyrethroid insecticides are commonly used for residential and commercial pest control in the US. Pregnant women and their fetuses are vulnerable to pesticide exposures during critical windows of neurodevelopment. Biomonitoring for exposure requires accurate and sensitive methods to assess exposures during pregnancy. The objective of this study was to develop a sensitive analytical method to measure pyrethroid metabolite concentrations in cord serum. Six pyrethroid metabolites, cis- and trans-3-(2,2-dichlorovinyl)-2,2-dimethyl-cyclopropanecarboxylic acid (c/t-DCCA), trans-chrysanthemum dicarboxylic acid (t-CDCA), cis-3-(2,2-dibromovinyl)-2,2-dimethyl-cyclopropane carboxylic acid (c-DBCA), 4-fluoro-3-phenoxybenzoic acid (FPBA), and 3-phenoxybenzoic acid (3PBA) were extracted from cord serum by a dichloromethane liquid-liquid extraction, derivatized by 1,1,1,3,3,3-hexafluoro-2-propanol carboxylic acid esterification, and then measured by gas chromatography/ion trap mass spectrometry. Limits of detection ranged from 0.02 to 0.6 ng/mL. Sixty-three cord serum samples were collected from maternal-fetal dyads in central New Jersey to test for pyrethroid metabolites. Non-specific metabolites, 3PBA, t-DCCA, and t-CDCA, were detected most frequently, present in 29%, 6.3% and 6.3% of samples, respectively. Sensitivities were comparable or greater than other published studies assessing pyrethroid metabolites in cord blood. Comparisons with other literature-reported studies emphasize the importance of method sensitivity when assessing exposures at biologically relevant concentrations.

Effects of feeding bulls dried corn distiller’s grains plus solubles or CaSO4 on mineral and metabolite concentrations in serum and seminal plasma

Yearling Angus bulls (n = 36) were assigned one of three diets: 1) 60 % concentrate as corn (CON, 0.2 % S, 13.4 % CP; n = 12); 2) 60 % dried corn distiller’s grains plus solubles (60DDGS 0.5 % S, 22.0 % CP; n = 12); 3) CON diet + equivalent sulfur of 60DDGS as CaSO4 (SULF, 0.5 % S, 13.9 % CP; n = 12) to evaluate effects of feeding 60 % DDGS or sulfur as CaSO4 on mineral and metabolite concentrations in serum and seminal plasma. Treatment × day interactions (P < 0.03) were observed for serum Cu, Se, and Mo. For Cu at d 112, lesser (P < 0.01) concentrations were observed in bulls fed the 60DDGS compared to SULF and CON diets. There were greater (P < 0.01) concentrations of Se at d 112 in bulls fed 60DDGS than CON and SULF diets. Concentrations of Mo were greater at d 56 and 112 (P < 0.01) in bulls fed CON compared to SULF and 60DDGS diets. In seminal plasma, there were treatment × day interactions (P < 0.02) for Cu and Mo. For Cu, at d 112, there was a lesser (P < 0.01) concentration in the bulls fed SULF compared to CON and 60DDGS diets. For Mo, there was a greater (P < 0.01) concentration in bulls fed the CON than 60DDGS and SULF diets at d 56 and 112. Changes in mineral and metabolite concentrations may have effects on bull reproductive performance when there is a relatively greater dietary sulfur content.

Association of metabolic markers with neutrophil function in healthy postpartum dairy cows

The objective of this study was to evaluate the associations of serum markers for systemic inflammation, liver, mineral, and energy status, and blood neutrophil counts with the function of circulating neutrophils in postpartum dairy cows. Blood samples were collected from 21 healthy Holstein cows at 5, 10, 14, and 21 d postpartum. Serum samples were used to measure concentrations of total calcium, phosphorus, magnesium, total protein, albumin, globulin, cholesterol, urea, glucose, gamma-glutamyl transferase, aspartate aminotransferase, glutamate dehydrogenase, haptoglobin (Hp), β-hydroxybutyrate, non-esterified fatty acids (NEFA), and insulin-like growth factor-1. The shift of percentage of activated neutrophils for phagocytosis (PPC) and oxidative burst (POB) and the median fluorescence intensity (MFI) for PC (MFIPC), OB (MFIOB), and endocytic and proteolytic degradation measured via DQ-ovalbumin (MFIDQ) were evaluated using flow cytometry. Mixed linear regression models were used to assess the associations of serum concentrations of metabolites and blood neutrophil counts with each neutrophil function assay outcome accounting for d postpartum, parity, and body condition score. Pearson correlation tests (r) were used to describe the relationships of metabolites and neutrophil counts with neutrophil function assays that were significant in the regression models. Greater serum concentrations of Hp were associated with lesser OB function (POB and MFIOB; r = −0.2 for both), but greater PPC and MFIDQ (r = 0.3 for both). Greater serum NEFA and cholesterol were associated with lesser (r = −0.3) and greater (r = 0.2) POB function, respectively. Blood neutrophil counts were associated with PPC (r = −0.2) and MFIDQ (r = −0.3). At 5 d postpartum, greater serum total protein was associated with greater OB function (POB and MFIOB; r = 0.4 for both). Other markers were not associated with neutrophil function. Some metabolic and inflammatory markers in healthy postpartum dairy cows were associated with circulating neutrophil function. However, these associations only explained a small proportion of the variance in neutrophil function. Serum Hp concentration was most associated with neutrophil function changes but had opposite directions of association with OB- and PC-related functions. Future studies should focus on understanding the mechanisms by which Hp and other metabolic indicators affect neutrophil function in healthy and diseased postpartum dairy cows.

Serum metabolomics study of women with different annual decline rates of anti-Müllerian hormone: an untargeted gas chromatography-mass spectrometry-based study.

Which metabolites are associated with varying rates of ovarian aging, measured as annual decline rates of anti-Müllerian hormone (AMH) concentrations? Higher serum concentrations of metabolites of phosphate, N-acetyl-d-glucosamine, branched chained amino acids (BCAAs), proline, urea and pyroglutamic acid were associated with higher odds of fast annual decline rate of AMH. Age-related rate of ovarian follicular loss varies among women, and the factors underlying such inter-individual variations are mainly unknown. The rate of ovarian aging is clinically important due to its effects on both reproduction and health of women. Metabolomics, a global investigation of metabolites in biological samples, provides an opportunity to study metabolites or metabolic pathways in relation to a physiological/pathophysiological condition. To date, no metabolomics study has been conducted regarding the differences in the rates of ovarian follicular loss. This prospective study was conducted on 186 reproductive-aged women with regular menstrual cycles and history of natural fertility, randomly selected using random case selection option in SPSS from the Tehran Lipid and Glucose Study. AMH concentrations were measured at baseline (1999-2001) and the fifth follow-up examination (2014-2017), after a median follow-up of 16 years, by immunoassay using Gen II kit. The annual decline rate of AMH was calculated by dividing the AMH decline rate by the follow-up duration (percent/year). The women were categorized based on the tertiles of the annual decline rates. Untargeted metabolomics analysis of the fasting-serum samples collected during the second follow-up examination cycle (2005-2008) was performed using gas chromatography-mass spectrometry. A combination of univariate and multivariate approaches was used to investigate the associations between metabolites and the annual decline rates of AMH. After adjusting the baseline values of age, AMH and BMI, 29 metabolites were positively correlated with the annual AMH decline rates. The comparisons among the tertiles of the annual decline rate of AMH revealed an increase in the relative abundance of 15 metabolites in the women with a fast decline (tertile 3), compared to those with a slow decline (tertile 1). There was no distinct separation between women with slow and fast decline rates while considering 41 metabolites simultaneously using the principal component analysis and the partial least-squares discriminant analysis models. The odds of fast AMH decline was increased with higher serum metabolites of phosphate, N-acetyl-d-glucosamine, BCAAs, proline, urea and pyroglutamic acid. Amino sugar and nucleotide sugar metabolism, BCAAs metabolism and aminoacyl tRNA biosynthesis were among the most significant pathways associated with the fast decline rate of AMH. Estimating the annual decline rates of AMH using the only two measures of AMH is the main limitation of the study which assumes a linear fixed reduction in AMH during the study. Since using the two-time points did not account for the variability in the decline rate of AMH, the annual decline rates estimated in this study may not accurately show the trend of the reduction in AMH. In addition, despite the longitudinal nature of the study and statistical adjustment of the participants' ages, it is difficult to distinguish the AMH-related metabolites observed in this study can accelerate ovarian aging or they are reflections of different rates of the aging process. Some metabolite features related to the decline rates of AMH have been suggested in this study; further prospective studies with multiple measurements of AMH are needed to confirm the findings of this study and to better understand the molecular process underlying variations in ovarian aging. This study, as a part of PhD thesis of Ms Nazanin Moslehi, was supported by Shahid Beheshti University of Medical Sciences (10522-4). There were no competing interests. N/A.

PSIII-6 Effects of repeated freeze and thaw cycles on serum and plasma metabolite concentrations in beef cattle

Abstract The objective of this study was to determine whether repeated freeze and thaw cycles alter bovine circulating metabolite concentrations, which serve as indicators of nutrient utilization. In research settings, it is common to thaw and refreeze samples when confirming results or performing further analyses; thus, it is important to determine if these actions alter metabolites. Jugular blood samples were collected from beef cows (n = 14) and fall-born calves (n = 14) at weaning. Samples were processed into multiple aliquots and stored at -20°C until analysis. Samples were thawed at 4°C for 8 to 10 h on each day of analysis and refrozen for at least 48 h between cycles. Concentrations of serum non-esterified fatty acids (NEFA), serum urea N, plasma triglycerides, and plasma glucose were analyzed using commercially available kits on a UV-visible light plate reader. Data were analyzed with sample type (cow vs. calf), thaw (cycles 1-4), and their interaction in a mixed model. Additionally, R2 were determined among freeze-thaw cycles within each metabolite. Thaw cycle impacted glucose concentrations (P = 0.03); other metabolites were unaffected (P &amp;gt; 0.25). Thaw cycle and its interaction with sample type did not affect metabolite concentrations (P &amp;gt; 0.33). Sample type did not affect triglyceride and BUN concentrations (P &amp;gt; 0.34). There was a type effect (P &amp;lt; 0.0001) for NEFA and glucose; NEFA were greater in cows and glucose was greater in calves. There was a strong relationship between urea N thaw cycles 1 and 2 (R2 = 0.88), which was also apparent between all other urea N, triglycerides, NEFA, and glucose cycles (R2 ≥ 0.79, 0.96, 0.91, and 0.61 respectively). These data indicate that repetitive freezing and thawing of bovine serum and plasma samples has little effect on non-esterified fatty acids, triglycerides, and urea N, but may impact glucose concentrations.

Abstract 13929: Methadone Lengthens Both Early and Late Ventricular Repolarization in Patients Undergoing Maintenance Therapy in an Urban Narcotic Treatment Center

Introduction: Methadone, a racemic mixture of S - and R -enantiomers, prolongs the QT interval and causes torsades de pointes. Methadone has been shown to lengthen late ventricular repolarization, represented by the T peak-Tend interval, which is modulated primarily by I Kr and I Ks . However, it is unknown whether methadone prolongs early ventricular repolarization, represented by the heart rate-corrected J-T peak (J-T peak c) interval, which is regulated primarily by L-type calcium and late sodium currents. In addition, the relationship between stereospecific serum methadone and metabolite concentrations and effects on J-T peak c and T peak -T end intervals are unknown. Hypothesis: 1) Methadone lengthens both J-T peak c and T peak -T end , and 2) Methadone effects on J-T peak c and T peak -T end correlate with serum S - and R -methadone and S - and R -metabolite [ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP)] concentrations. Methods: Patients (n=81) on steady state maintenance methadone therapy at an urban narcotic treatment center who had undergone a pre-methadone 12-lead ECG within the previous 6 months were identified and asked to return for a followup ECG during steady-state methadone therapy. In a subset (n=41), blood was obtained to determine serum S - and R -methadone and S - and R -EDDP concentrations. Results: Compared to baseline, methadone lengthened mean ± SD J-T peak c (225 ± 26 vs 232 ± 28 ms, p=0.02) as well as T peak - T end (80 ±14 vs 90 ± 16 ms, p&lt;0.00001). Serum concentrations of S -methadone correlated with J-T peak c [R 2 = 0.41 (95% CI 0.11-0.65), p=0.007]. Serum R -methadone concentrations also correlated with J-T peak c [R 2 = 0.47 (0.17-0.68), p=0.002]. In addition, serum concentrations of S - and R -EDDP correlated with J-T peak c [R 2 = 0.39 (0.09-0.63), p=0.01] and [R 2 = 0.38 (0.07-0.62), p=0.02], respectively. There were no significant correlations between serum S - and R -methadone or S- and R-EDDP concentrations with T peak - T end intervals. Conclusions: Methadone lengthens both early and late ventricular repolarization, suggesting that methadone’s ion channel effects may extend beyond inhibition of I Kr . Serum S - and R -methadone and S -and R -EDDP concentrations are associated with effects on J-T peak c intervals.

Retrospective Analysis of Pediatric and Adult Populations Using an LC-MS/MS Method for Oxcarbazepine/Eslicarbazepine Metabolite.

Therapeutic drug monitoring of anti-epileptic drugs is important to manage seizure control in patients with epilepsy. Oxcarbazepine is a second-generation anti-epileptic drug approved for use in pediatric patients, and eslicarbazepine acetate is a newer generation drug used as adjunctive therapy and monotherapy for partial-onset (focal) seizures. While several second and third generation anti-epileptic drugs have broader therapeutic efficacy in patients, these drugs can still have severe side effects and variable interpatient pharmacokinetics. Consequently, there is a need for accurate and sensitive analytical methods to support therapeutic drug monitoring. An assay improvement for a LC-MS/MS method was developed for the major metabolite of oxcarbazepine and eslicarbazepine, licarbazepine (MHD), using a 13C-labeled form of the compound as the internal standard. Additionally, retrospective data analysis was used to compare the distribution of results observed in adult vs pediatric patients. Accuracy and linearity across the analytical measuring range of 1 to 60 µg/mL was acceptable. Inter- and intra-run precision was less than 6% at 3 concentrations tested. The limit of detection was determined to be 0.5 µg/mL. Significant interference from hemolysis, icterus, lipemia, or 187 other potential interferences was not detected. The improved assay for MHD was appropriate for clinical use. Retrospective data analysis showed that pediatric and adult patients had a similar distribution of oxcarbazepine/eslicarbazepine metabolite concentrations in serum.

Common and specific mineral and metabolic features in dairy cows with clinical metritis, hypocalcaemia or ketosis

The objectives were to evaluate differences in serum concentration of metabolites, macro minerals and hepatic enzymes at pre and postpartum time-points in dairy cows diagnosed with clinical metritis, hypocalcaemia or ketosis postpartum. A total of 144 Holstein cows from 11 commercial dairy herds in Alberta, (Western Canada) were enrolled in this study. Cows with clinical metritis had lower serum concentrations of glutamate dehydrogenase (GLDH) at pre and postpartum and lower total Ca, albumin, urea, and cholesterol at postpartum when compared to control cows. Cows with hypocalcaemia had greater serum concentrations of Na, Cl, and calculated osmolarity (CalOsmo) at prepartum and lower concentration of total serum Ca, glucose, cholesterol, gamma-glutamyl transpeptidase (GGT), GLDH, total protein and albumin at postpartum. Prepartum serum concentrations of non-esterified fatty acids (NEFA), beta hydroxybutyrate (BHB), Cl, albumin/globulin ratio (A/G), Na, K and sum of Na and K were greater in ketotic cows when compared with control cows. Cows with ketosis had also greater postpartum serum concentrations of NEFA, BHB, GGT and aspartate transaminase (AST) when compared with control cows. Prepartum serum Na and Cl concentrations and CalOsmo were greater in cows diagnosed with hypocalcaemia or ketosis when compared with control cows. Furthermore, postpartum serum concentrations of total Ca, cholesterol, albumin and GLDH were significantly affected by hypocalcaemia or clinical metritis and concentrations of GGT by hypocalcaemia or ketosis. Finally, postpartum serum concentrations of haptoglobin increased in all disease groups when compared with control cows. These results suggest common metabolic features for clinical metritis, hypocalcaemia and ketosis in dairy cows in addition to the specific ones.

Hepatocyte-intrinsic type I interferon signaling reprograms metabolism and reveals a novel compensatory mechanism of the tryptophan-kynurenine pathway in viral hepatitis.

The liver is a central regulator of metabolic homeostasis and serum metabolite levels. Hepatocytes are the functional units of the liver parenchyma and not only responsible for turnover of biomolecules but also act as central immune signaling platforms. Hepatotropic viruses infect liver tissue, resulting in inflammatory responses, tissue damage and hepatitis. Combining well-established in vitro and in vivo model systems with transcriptomic analyses, we show that type I interferon signaling initiates a robust antiviral immune response in hepatocytes. Strikingly, we also identify IFN-I as both, sufficient and necessary, to induce wide-spread metabolic reprogramming in hepatocytes. IFN-I specifically rewired tryptophan metabolism and induced hepatic tryptophan oxidation to kynurenine via Tdo2, correlating with altered concentrations of serum metabolites upon viral infection. Infected Tdo2-deficient animals displayed elevated serum levels of tryptophan and, unexpectedly, also vast increases in the downstream immune-suppressive metabolite kynurenine. Thus, Tdo2-deficiency did not result in altered serum homeostasis of the tryptophan to kynurenine ratio during infection, which seemed to be independent of hepatocyte-intrinsic compensation via the IDO-axis. These data highlight that inflammation-induced reprogramming of systemic tryptophan metabolism is tightly regulated in viral hepatitis.