Metabolites In Plasma Of Patients Research Articles

Study the local metabolic changes of aneurysms through microcatheter sampling

Intracranial aneurysm is the primary cause of nontraumatic subarachnoid hemorrhage. To assess aneurysm metabolism, we present a method of intra-operatively collecting blood samples from the aneurysm neck, as well as the proximal and distal responsible vessels, using microcatheters. Through these paired comparisons, we can eliminate the interpatient variation usually observed in plasma samples taken from the peripheral vein. We utilized 39 plasma samples from 13 intracranial patients to characterize the metabolite profiles using untargeted liquid chromatography-mass spectrometry. Our findings revealed that l-tyrosine is upregulated at relatively high levels at the aneurysm neck than the proximal and distal aneurysm, whereas phenylpyruvic acid, l-cystine, and l-ornithine are downregulated. Based on this, there was also a significant decrease in arginine within small aneurysm of the internal carotid artery. The 6-month follow-up indicated that patients who experienced good recovery had lower levels of biliverdin, bilirubin, and metabolites of coenzyme Q within the aneurysm. In conclusion, our investigation provides a comprehensive overview of plasma metabolites in patients with intracranial aneurysms, shedding light on potential pathogenetic mechanisms in unruptured intracranial aneurysms. Moreover, the study proposes innovative ideas for establishing postoperative follow-up timelines for flow diverter devices.

Genetic assessment of the causal effect of plasma metabolites and metabolic pathways on delirium

PurposeDelirium has a significant impact on millions of people globally and is closely linked to an unfavorable prognosis for complications. Observational studies suggest that plasma metabolites may play significant roles as markers and effectors of delirium, but causal relationships have not yet been elucidated. Based on the most recent genome-wide association study (GWAS) data, our study aims to present novel insights into the genetic relationship between delirium and plasma metabolites. This investigation offers potential clues for utilizing plasma metabolites as predictors of delirium development.MethodsWe performed a thorough Mendelian randomization (MR) analysis to investigate the causal relationship between 1,091 individual metabolites and 309 metabolite ratios in plasma with respect to delirium. Inverse-variance weighting (IVW) was employed as the primary estimation method, while MR-Egger and weighed median methods were utilized to assess the robustness of the results. Sensitivity analyses encompassing the MR Pleiotropy RESidual Sum and Outlier (MR-PRESSO) test, Cochran Q test, leave-one-out analysis and MR Egger intercept analysis were also undertaken. Additionally, the MR Steiger test was performed to explore any potential reverse causal effect of metabolites on delirium.ResultsA total of 63 types of plasma metabolites associated with delirium were detected using the IVW method (p < 0.05). Among the known metabolites, our analysis revealed that two specific metabolites (1-palmitoyl-2-palmitoleoyl-gpc (16:0/16:1) and homovanillate) and a metabolite ratio (phosphate to oleoyl-linoleoyl-glycerol (18:1 to 18:2)) demonstrated a consistent and significant causal relationship with delirium across all analysis methods. Finally, no evidence of pleiotropy was detected in our analysis.ConclusionsOur study has revealed a causal association between blood metabolites and the risk of delirium. homovanillate is known to be associated with immunity and redox, 1-palmitoyl-2-palmitoleoyl-gpc (16:0/16:1) and metabolite ratio (phosphate to oleoyl-linoleoyl-glycerol (18:1 to 18:2)) may play a role in lipid regulation. These findings may provide fresh insights into the identification of suitable diagnostic markers and potential treatment strategies focused on specific plasma metabolites in patients with delirium. However, further experiments are required to gain a comprehensive understanding of the underlying biological mechanisms involved.

Plasma metabolic profiling of patients with tetralogy of fallot

BackgroundTetralogy of Fallot (TOF) is a common congenital heart disease with high mortality. However, the medical imageology and liquidbiopsy techniques present certain limitations. Thus, this study investigated the plasma metabolic profiles to distinguish key metabolites for early diagnosis of TOF. MethodsIn total, 69 patients with TOF and 43 normal controls were enrolled for targeted metabolomics based on liquid chromatography-tandem mass spectroscopy (LC-MS/MS). Absolute quantification of metabolites was performed using our standard database. The differentially expressed metabolites (DEMs) were screened by fold change (FC), VIP value and pearson correlation coefficient of OPLS-DA model. Receiver operating characteristic curve (ROC) was used to evaluate predictive ability of DEMs. ResultsDifferent metabolic profiles were presented between TOF and Normal.The pathway analysis showed that significantly changed metabolites were enriched in nicotinamide and purine metabolism. Many intermediatesproductof purine and amido acid were higher in TOF than in Normal group, while energy substrates and electron carriers were lower in TOF than in Normal group. ROC analysis revealed a high diagnostic value of plasma FAD for differentiating TOF from Normal (AUC = 1). ConclusionOur study quantitatively characterized plasma metabolites in patients with TOF and may help to develop reliable biomarkers that contribute to the early TOF screening.

Metabolic phenotyping in phenylketonuria reveals disease clustering independently of metabolic control.

Phenylketonuria (PKU, MIM #261600) is one of the most common inborn errors of metabolism (IEM) with an incidence of 1:10000 in the European population. PKU is caused by autosomal recessive mutations in phenylalanine hydroxylase (PAH) and manifests with elevation of phenylalanine (Phe) in plasma and urine. Untreated PKU manifests with intellectual disability including seizures, microcephaly and behavioral abnormalities. Early treatment and good compliance result in a normal intellectual outcome in many but not in all patients. This study examined plasma metabolites in patients with PKU (n = 27), hyperphenylalaninemia (HPA, n = 1) and healthy controls (n = 32) by LC- MS/MS. We hypothesized that PKU patients would exhibit a distinct "submetabolome" compared to that of healthy controls. We further hypothesized that the submetabolome of PKU patients with good metabolic control would resemble that of healthy controls. Results from this study show: (i) Distinct clustering of healthy controls and PKU patients based on polar metabolite profiling, (ii) Increased and decreased concentrations of metabolites within and afar from the Phe pathway in treated patients, and (iii) A specific PKU-submetabolome independently of metabolic control assessed by Phe in plasma. We examined the relationship between PKU metabolic control and extended metabolite profiles in plasma. The PKU submetabolome characterized in this study represents the combined effects of dietary adherence, adjustments in metabolic pathways to compensate for defective Phe processing, as well as metabolic derangements that could not be corrected with dietary management even in patients classified as having good metabolic control. New therapeutic targets may be uncovered to approximate the PKU submetabolome to that of healthy controls and prevent long-term organ damage.

Purine metabolites promote ectopic new bone formation in ankylosing spondylitis

Ankylosing spondylitis (AS) is a chronic inflammatory rheumatic disease that mainly affects the axial skeleton, whose typical features are inflammatory back pain, bone structural damage and pathological new bone formation. The pathology of ectopic new bone formation is still little known. In this study, we found increased purine metabolites in plasma of patients with AS. Similarly, metabolome analysis indicated increased purine metabolites in both serum of CD4-Cre; Ptpn11fl/fl and SHP2-deficient chondrocytes. SHP2-deficient chondrocytes promoted the growth of wild type chondrocytes and differentiation of osteoblasts in CD4-Cre; Ptpn11fl/fl mice, which spontaneously developed AS-like bone disease. Purine metabolites, along with PTHrP derived from SHP2-deficient chondrocytes, accelerated the growth of chondrocytes and ectopic new bone formation through PKA/CREB signaling. Moreover, Suramin, a purinergic receptor antagonist, suppressed pathological new bone formation in AS-like bone disease. Overall, these results highlight the potential role of targeting purinergic signaling in retarding ectopic new bone formation in AS.

Endocannabinoid signalling/cannabinoid receptor 2 is involved in icariin-mediated protective effects against bleomycin-induced pulmonary fibrosis

BackgroundIdiopathic pulmonary fibrosis (IPF) is a progressive fibrotic disease of unknown aetiology with limited effective treatment options. It is important to explore novel therapeutic targets and develop potential drugs for IPF. PurposeThe aim of the present study was to analyse nontargeted plasma metabolites in patients with IPF and investigate whether cannabinoid receptor (CB2) activation mediates the antifibrotic effect of icariin (ICA). MethodsWe used an untargeted metabolomics method to detect the global metabolic profiles in the plasma of stable IPF patients and patients with stable chronic obstructive pulmonary disease (COPD), as well as healthy subjects. The untargeted liquid chromatography-mass spectrometry (LC-MS) analysis revealed that IPF showed differential metabolites and perturbed signalling pathways. ICA is pharmacologically bioactive and possesses extensive therapeutic capacities such as osteoprotective, neuroprotective, cardiovascular protective, anti-cancer, anti-inflammation and reproductive function. Therefore, ICA was administered to a pulmonary fibrosis rat model for 4 weeks and then the effect of ICA on pulmonary fibrosis was examined by dissection and histology. ResultsThe metabolites in the plasma were determined by untargeted LC-MS. An unsupervised principal component analysis (PCA) was used to observe the distribution of each sample, and a supervised partial least squares-discriminant analysis (PLS-DA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) results showed that there was significant separation between any two groups. ROC curve analyses revealed that 8 metabolites with high AUCs above 0.7 between the three groups of plasma samples. Pathway enrichment analysis revealed that 3 metabolites are involved in retrograde endocannabinoid signalling. Meanwhile, Retrograde endocannabinoid signalling was identified significantly different in IPF group from other groups by Kyoto encyclopedia of Genes and Genomes (KEGG) pathway analysis, and then we further confirmed the endocannabinoid signalling by detecting the expression of the main receptors in bleomycin-induced pulmonary fibrosis, COPD rat model and normal rats. Consistent with previous studies, we found that the elevation of CB1 and CB2 in the lung tissues could be a signature of the pulmonary fibrosis rat model. Importantly, ICA may alleviate bleomycin-induced lung injury by decreasing CB1 and CB2 expression in the bleomycin-induced rat model. ConclusionTaken together, we measured the global metabolic profile of IPF patients and identified CB2 as a novel potential target. ICA treatment demonstrated outstanding therapeutic effects on bleomycin-induced pulmonary fibrosis and targeting on CB2 may be the main underlying mechanism. ICA is a promising drug candidate to cure pulmonary fibrosis and mediate antagonists of the CB2 receptor.

Non-targeted metabonomic analysis of plasma in patients with atherosclerosis by liquid chromatography-mass spectrometry.

BackgroundThis study sought to analyze non-targeted plasma metabolites in patients with atherosclerosis (AS).MethodsThe plasma of patients with AS (the patient group) and the plasma of age-matched and gender-matched healthy individuals (the control group) at the Taihe Hospital was collected. One hundred patients were included in the study (60 in the patient group and 40 in the control group). Fasting venous plasma was collected in the morning. The metabolites in the plasma were examined by liquid chromatography-mass spectrometry (LC-MS). An unsupervised principal component analysis (PCA) was conducted to observe the overall distribution of each sample and the stability of the analysis process. Next, a supervised partial least squares-discriminant analysis (PLS-DA) and an orthogonal partial least squares-discriminant analysis (OPLS-DA) were conducted to examine the overall differences among the metabolic profiles of the groups and identify different metabolites in the groups. Pathway enrichment was analyzed using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database.ResultsIn total, 1,126 different metabolites were detected in the patient and control groups. Compared to the control group, 411 species decreased, and 715 species increased in the patient group. There were 61 different metabolites with a variable weight in the projection (VIP) >1 and a P<0.05. There were 34 types of lipid metabolites, 10 types of carbon and oxygen compounds, 8 types of organic acids and derivatives, 4 types of organoheterocyclic compounds, 3 types of nitrogen-containing organic compounds, and 2 types of nucleotides and analogs. Compared to the control group, 47 species decreased, and 14 species increased in the patient group. The following 9 metabolites had the most significant differences (|log2fold change| >1; P<0.05): 2-tetradecanone, pantothenol, all-trans-13,14-dihydroretinol, linoleoyl ethanolamide, N-oleoylethanolamine, 4-methyl-2-pentenal, Cer (d18:1/14:0), chenodeoxycholic acid glycine conjugate, and 5-acetamidovalerate. The enrichment analysis results of the 61 different metabolite pathways identified 17 metabolic pathways with significant differences (P<0.05), including the choline metabolism, lipid metabolism, autophagy, amino acid metabolism, vitamin digestion, and absorption pathways.ConclusionsThere are significant differences in non-targeted plasma metabolites between patients with AS and healthy individuals. The above-mentioned 9 most significantly different metabolites may be potential markers of AS.

A combination of Q-TOF LC/MS and LC-MS/MS based metabolomics in pediatric-onset multiple sclerosis demonstrates potential biomarkers for unclassified patients.

Metabolomics has the potential to provide putative biomarkers and insights into the pathophysiology and diagnosis of pediatric multiple sclerosis (pMS), which is an inflammatory demyelinating disorder of the central nervous system with a broad spectrum of clinical manifestations. In this study, we aimed to investigate serum metabolomics in pMS to help elucidate the pathophysiology of MS. An untargeted approach was applied using the quadrupole time-of-flight liquid chromatography/mass spectrometry (Q-TOF LC/MS) method to study plasma metabolites in patients with pMS (n = 33), patients with unclassified central nervous system demyelinating diseases (n = 6), and age-matched healthy control subjects (n = 40). The patient and control groups were compared for metabolites and the normalized peak areas differed statistically (p < 0.05), showing at least a 1.25-fold change between groups. Bioinformatic tools combined with a clinical perspective were employed for the identification of the putative metabolites. In addition to the untargeted metabolomics approach, targeted LC-MS/MS metabolite analysis was employed to compare the pMS group with the control group. Significant differences between the patient and control groups were noted for tyramine, 4-hydroxyphenylacetaldehyde, sphingosine/3-dehydrosphinganine, prostaglandins/thromboxane A2, 20-hydroxy-leukotriene E4, 3α,7α,12α-trihydroxy-5β-cholestan26-al/calcitriol, pantetheine, ketoleucine/3-methyl-2-oxovaleric acid, L-arginine/D-arginine, coproporphyrinogen III, (S)-reticuline, carnosine, cytidine, and phosphoribosyl pyrophosphate. Additional tests for sphingosine 1-phosphate, sphingophosphocholines, ceramides, oxysterols, and calcitriol levels yielded significant metabolomic differences for the pMS group compared to the control group. The metabolomic data of 3/6 patients with unclassified demyelinating disorders matched the pMS group; their follow-up verified the diagnosis of pMS. In general, plasma metabolites related to sphingolipid metabolism, myelin products, inflammatory pathways, mitochondrial dysfunction, and oxidative stress were found to be altered in cases of pMS. The method applied in this study, combining untargeted analysis with a targeted approach, can be applied to larger series of cases of pMS and other demyelinating disorders for further validation.

Pre- and Post-treatment Levels of Plasma Metabolites in Patients With Bipolar Depression.

Background: Bipolar disorder (BD) is a serious mental disease with complex clinical manifestations and high recurrence rate. The purpose of this study was to detect metabolites related to the diagnosis and efficacy evaluation of bipolar depression in plasma samples by metabolomics.Methods: Thirty-one bipolar depression patients were recruited and completed 8 weeks medication and a matched group of 47 healthy controls (HCs) was recruited. Nuclear magnetic resonance spectroscopy was used to profile plasma samples of bipolar depression patients at baseline and after 8 weeks medication, and HCs. Then Multivariate statistical analysis was performed to analyze differences of plasma metabolites among the three groups.Results: We detected seven specific differential metabolites in bipolar depression. Six of the metabolites were returned to the normal levels in different degrees after 8 weeks medication, only Glycine continuously decreased in the acute and significant improvement stages of bipolar depression (VIP > 1 and p < 0.05). These differential metabolites involved several metabolic pathways.Limitations: The small sample size was one of the most prominent limitations. Each BD patient was given an individualized medication regimen according to the clinical guidelines.Conclusion: There were metabolites changes before and after 8 weeks medication. Glycine may be a characteristic marker of bipolar depression and does not change with the improvement of bipolar depression, while other 6 differential metabolites may be biomarkers associated with the pathological development or the improvement of bipolar depression. And, these differential metabolites mainly related to energy metabolism, amino acid metabolism and gut microbiota metabolism.

LC-MS/MS method for simultaneous quantification of the first-line anti-tuberculosis drugs and six primary metabolites in patient plasma: Implications for therapeutic drug monitoring

The pharmacokinetic profiling of drug substances and corresponding metabolites in the biological matrix is one of the most informative tools for the treatment efficacy assessment. Therefore, to satisfy the need for comprehensive monitoring of anti-tuberculosis drugs in human plasma, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for simultaneous quantification of first-line anti-tuberculosis drugs (ethambutol, isoniazid, pyrazinamide, and rifampicin) along with their six primary metabolites. Simple single-step protein precipitation with methanol was chosen as the most convenient sample pre-treatment method. Chromatographic separation of the ten analyte mixture was achieved within 10 minutes on a reverse-phase C8 column using mobile phase gradient mode. The multiple reaction monitoring mode (MRM) was used for analyte detection and quantification in patient samples. The chosen quantification ranges fully covered expected plasma concentrations. The method exhibited acceptable selectivity; the within- and between-run accuracy ranged from 87.2 to 113.6%, but within- and between-run precision was between 1.6 and 14.9% (at the LLOQ level CV < 20%). Although the response of the isonicotinic acid varied depending on the matrix source (CV 21.8%), validation results proved that such inconsistency does not affect the accuracy and precision of results. If stored at room temperature plasma samples should be processed within 4 h after collection, temporary storage at −20 °C up to 24 h is acceptable due to stability issues of analytes. The developed method was applied for the patient sample analysis (n = 34) receiving anti-tuberculosis treatment with the first-line drugs.

The Feasibility of Studying Metabolites in PICU Multi-Organ Dysfunction Syndrome Patients over an 8-Day Course Using an Untargeted Approach.

Metabolites are generated from critical biological functions and metabolism. This pediatric study reviewed plasma metabolites in patients suffering from multi-organ dysfunction syndrome (MODS) in the pediatric intensive care unit (PICU) using an untargeted metabolomics approach. Patients meeting the criteria for MODS were screened for eligibility and consented (n = 24), and blood samples were collected at baseline, 72 h, and 8 days; control patients (n = 4) presented for routine sedation in an outpatient setting. A subset of MODS patients (n = 8) required additional support with veno-atrial extracorporeal membrane oxygenation (VA-ECMO) therapy. Metabolites from thawed blood plasma were determined from ion pairing reversed-phase liquid chromatography–mass spectrometry (LC-MS) analysis. Chromatographic peak alignment, identification, relative quantitation, and statistical and bioinformatics evaluation were performed using MAVEN and MetaboAnalyst 4.0. Metabolite analysis revealed 115 peaks per sample. From the partial least squares-discriminant analysis (PLS-DA) with variance of importance (VIP) scores above ≥2.0, 7 dynamic metabolites emerged over the three time points: tauro-chenodeoxycholic acid (TCDCA), hexose, p-hydroxybenzoate, hydroxyphenylacetic acid (HPLA), 2_3-dihydroxybenzoic acid, 2-keto-isovalerate, and deoxyribose phosphate. After Bonferroni adjustment for repeated measures, hexose and p-hydroxybenzoate were significant at one time point or more. Kendall’s tau-b test was used for internal validation of creatinine. Metabolites may be benign or significant in describing a patient’s pathophysiology and require operator interpretation.

Simple Liquid Chromatography-Tandem Mass Spectrometry Method for Quantitation of Total and Free Aprepitant and Its Active N-Dealkylated Metabolites in Human Plasma.

Aprepitant, an antiemetic selective neurokinin-1 receptor antagonist, is primarily metabolized to the active N-dealkylated form (ND-AP) and then converted to its carbonyl form (ND-CAP) in humans. This study developed a simple liquid chromatography-tandem mass spectrometry method using electrospray ionization for the quantitation of plasma total and free aprepitant and its N-dealkylated metabolites and used them to analyze patient plasma. Free aprepitant and ND-AP in plasma were fractionated using centrifugal ultrafiltration. The analytes in plasma or their ultrafiltered specimens treated with triethylamine/acetonitrile were isocratically separated using a 3-μm octadecylsilyl column with a total run time of 10 minutes and scanned using positive ion electrospray ionization. The calibration curves of total aprepitant, ND-AP, and ND-CAP were prepared at concentration ranges of 50-2500, 20-1000, and 5-250 ng/mL, respectively, whereas that of free aprepitant and ND-AP were at a concentration range of 2-150 ng/mL. The intraassay and interassay accuracy and imprecision values were 93.5%-107.7% and 94.6%-103.3%, and 2.1%-7.5% and 1.0%-8.9%, respectively. Aprepitant and its metabolites did not exhibit any matrix effects or instabilities in the plasma specimens. In cancer patients receiving oral aprepitant, the plasma concentration ranges of total aprepitant, ND-AP, and ND-CAP, and free aprepitant and ND-AP were 137-2170, 104-928, 22.4-97.6, 8.11-60.0, and 3.53-56.0 ng/mL, respectively. The median plasma free fraction proportion of aprepitant and ND-AP was 4.14% and 4.90%, respectively. The present developed method showed an acceptable analytical performance and can be used to evaluate total and free aprepitant and its N-dealkylated metabolites in patient plasma.

Effects of a Vegetarian Diet on Cardiometabolic Risk Factors, Gut Microbiota, and Plasma Metabolome in Subjects With Ischemic Heart Disease: ARandomized, Crossover Study.

BackgroundA vegetarian diet (VD) may reduce future cardiovascular risk in patients with ischemic heart disease.Methods and ResultsA randomized crossover study was conducted in subjects with ischemic heart disease, assigned to 4‐week intervention periods of isocaloric VD and meat diet (MD) with individually designed diet plans, separated by a 4‐week washout period. The primary outcome was difference in oxidized low‐density lipoprotein cholesterol (LDL‐C) between diets. Secondary outcomes were differences in cardiometabolic risk factors, quality of life, gut microbiota, fecal short‐chain and branched‐chain fatty acids, and plasma metabolome. Of 150 eligible patients, 31 (21%) agreed to participate, and 27 (87%) participants completed the study. Mean oxidized LDL‐C (−2.73 U/L), total cholesterol (−5.03 mg/dL), LDL‐C (−3.87 mg/dL), and body weight (−0.67 kg) were significantly lower with the VD than with the MD. Differences between VD and MD were observed in the relative abundance of several microbe genera within the families Ruminococcaceae, Lachnospiraceae, and Akkermansiaceae. Plasma metabolites, including l‐carnitine, acylcarnitine metabolites, and phospholipids, differed in subjects consuming VD and MD. The effect on oxidized LDL‐C in response to the VD was associated with a baseline gut microbiota composition dominated by several genera of Ruminococcaceae.ConclusionsThe VD in conjunction with optimal medical therapy reduced levels of oxidized LDL‐C, improved cardiometabolic risk factors, and altered the relative abundance of gut microbes and plasma metabolites in patients with ischemic heart disease. Our results suggest that composition of the gut microbiota at baseline may be related to the reduction of oxidized LDL‐C observed with the VD.RegistrationURL: https://www.clini​caltr​ials.gov; Unique identifier: NCT02942628.

Disrupted Folate Metabolism with Anesthesia Leads to Myelination Deficits Mediated by Epigenetic Regulation of ERMN

Objective: Anesthetics during early life stages may impair cognitive and emotional functions, of which the underlying molecular mechanisms remain largely unknown. Methods: Here we performed the unbiased transcriptome profiling in prefrontal cortex of both non-human primates and mice after anesthesia. We also constructed a brain blood barrier (BBB)-crossing AAV-PHP.EB vector to harbor GFP or ERMN expression cassette. Findings: We found that the thymidylate synthase gene (TYMS), a critical gene implicated in folate metabolism, was significantly downregulated after multiple anesthesia in both non-human primates and mice. Hereby, we examined the metabolites in plasma of patients with anesthesia and surprisingly found that blood folate levels significantly decreased after surgeries under general anesthesia in children. Combined with transcriptome and genome-wide DNA methylation analysis, we identified that ERMN was the primary target of the disrupted folate metabolism. We further showed that myelination in brain during early postnatal stage was indeed compromised by the anesthesia in mice, which was fully rescued by systematic administration of folic acid or expression of ERMN in the brain through brain-specific delivery of adeno-associated virus. Notably, administration of folic acid and ERMN remarkably alleviated the cognitive impairment of mice caused by the anesthesia during early postnatal stages. Interpretation: This study indicates that anesthesia during early the postnatal period leads to disrupted folate metabolism and subsequently defects in myelination in the young brain, and ERMN is the key target gene affected by anesthesia via epigenetic mechanisms. Clinical Trial Registration: NCT03746340 Funding Statement: This work was mainly supported by NSFC Grants (#81571028, # 81771132, #31625013, #91732302) Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: The study protocol was approved by the Human Research Ethics Committee of Shanghai 9th People’s Hospital and Shanghai Jiao Tong University in Shanghai, P. R. China SH9H-2018-T53-2. The animal studies were performed according to the guidelines and regulations of the Institute of Laboratory Animal Science, Peking Union Medical College and Chinese Academy of Medical Science (Peking, China). Efforts were made to minimize the number of animals in the studies. The use of rhesus macaque in research at Institute of Laboratory Animal Science (Peking, China) was approved by Institutional Animal Care and Use Committee (Protocol number #XC17001).

Targeted metabolomic analysis of plasma metabolites in patients with coronary heart disease in southern China.

Coronary heart disease (CHD), one of the leading causes of death in the world, is a complex metabolic disorder due to genetic and environmental interactions. The potential mechanisms and diagnostic biomarkers for different types of coronary heart disease remain unclear. Metabolomics is increasingly considered to be a promising technology with the potential to identify metabolomic features in an attempt to distinguish the different stages of CHD.We aimed to investigate serum metabolite profiling between CHD patients and normal coronary artery (NCA) subjects and identify metabolic biomarkers associated with CHD progression in an ethnic Hakka population in southern China.Using a novel targeted metabolomics approach, we explored the metabolic characteristics of CHD patients. Blood samples from 302 patients with CHD and 59 NCA subjects were collected that analyses using targeted liquid-chromatography coupled with tandem mass spectrometry (LC-MS).A total of 361 blood samples were determined using targeted LC-MS. Plasma concentrations for trimetlylamine oxide (TMAO), choline, creatinine, and carnitine were significantly higher in patients with CHD compared to the NCA cohort. Further, we observed that the concentration of the 4 metabolites were higher than that of the NCA group in any group of CHD, which including acute myocardial infarction (AMI), unstable angina (UA), and stable angina (SA). In addition, the diagnostic model was constructed based on the metabolites identified and the ROC curve of the NCA subjects and CHD patients were performed. For choline and creatinine, the AUCs ranged from 0.720 to 0.733. For TMAO and carnitine, the AUCs ranged from 0.568 to 0.600.In conclusion, the current study illustrates the distribution of 4 metabolites between CHD patients and NCA subjects. Metabolomics analysis may yield novel predictive biomarkers that will potentially provide value for clinical diagnosis of CHD.

Tryptophan Metabolites in Plasma of Patients with Depression

There are two major pathways in tryptophan metabolism. The serotonin pathway mediates mood, anxiety, memory, cognition, and is impaired in depression. The kynurenine pathways are involved in immunity, inflammation, muscle movement, and mental health. We investigated changes in tryptophan metabolites in plasma from depressed patients. Plasma levels of serotonin were very low or undetectable in patients with monopolar depression. 5-hydroxyindole acetic acid (5-HIAA)/tryptophan ratios or kynurenine/tryptophan ratios were not different between healthy controls and depressive patients, indicating rapid degradation of serotonin into 5-HIAA. However, there were no significant changes in kynurenine levels in depressed patients. It is important to examine the roles of tryptophan metabolites in monopolar and bipolar depression since both require different treatments.

Metabolite profiling of plasma in patients with ossification of the posterior longitudinal ligament

BackgroundMetabolomics is one of the “omics” technologies, and is a comprehensive analysis of small molecule metabolites which include amino acid, nucleotides, carbohydrates and fatty acid. The purpose of the present study was to compare the differences of metabolite profiling between patients with ossification of the posterior longitudinal ligament (OPLL) and control subjects. MethodsWe analyzed plasma metabolites in patients with cervical OPLL (n = 10) and in control subjects (n = 10). Ionic metabolites were analyzed using capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS) and lipophilic metabolites were analyzed using liquid chromatograph time-of-flight mass spectrometry (LC-TOFMS). ResultsA total of 259 metabolites (144 metabolites in CE-TOFMS and 115 metabolites in LC-TOFMS) were detected. Among the 259 metabolites, six metabolites, namely acylcarnitine (AC) (14:0), palmitoylcarnitine, AC (18:2), fatty acid (FA) (24:2), thyroxine, thiaproline were significantly larger in OPLL group, even in analyzes excluding patients with diabetes mellitus and hyperlipidemia. ConclusionsWe examined the metabolite profiling in patients with OPLL for the first time and detected six metabolites showing suggestive association with disease. These results of the present study could lead to new insights into clarifying the molecular pathomechanisms of OPLL.

NAD Deficiency, Congenital Malformations, and Niacin Supplementation

BackgroundCongenital malformations can be manifested as combinations of phenotypes that co-occur more often than expected by chance. In many such cases, it has proved difficult to identify a genetic cause. We sought the genetic cause of cardiac, vertebral, and renal defects, among others, in unrelated patients.MethodsWe used genomic sequencing to identify potentially pathogenic gene variants in families in which a person had multiple congenital malformations. We tested the function of the variant by using assays of in vitro enzyme activity and by quantifying metabolites in patient plasma. We engineered mouse models with similar variants using the CRISPR (clustered regularly interspaced short palindromic repeats)–Cas9 system.ResultsVariants were identified in two genes that encode enzymes of the kynurenine pathway, 3-hydroxyanthranilic acid 3,4-dioxygenase (HAAO) and kynureninase (KYNU). Three patients carried homozygous variants predicting loss-of-function changes in the HAAO or KYNU proteins (HAAO p.D162*, HAAO p.W186*, or KYNU p.V57Efs*21). Another patient carried heterozygous KYNU variants (p.Y156* and p.F349Kfs*4). The mutant enzymes had greatly reduced activity in vitro. Nicotinamide adenine dinucleotide (NAD) is synthesized de novo from tryptophan through the kynurenine pathway. The patients had reduced levels of circulating NAD. Defects similar to those in the patients developed in the embryos of Haao-null or Kynu-null mice owing to NAD deficiency. In null mice, the prevention of NAD deficiency during gestation averted defects.ConclusionsDisruption of NAD synthesis caused a deficiency of NAD and congenital malformations in humans and mice. Niacin supplementation during gestation prevented the malformations in mice. (Funded by the National Health and Medical Research Council of Australia and others.)

The metabolites in peripheral blood mononuclear cells showed greater differences between patients with impaired fasting glucose or type 2 diabetes and healthy controls than those in plasma.

To determine differences between peripheral blood mononuclear cells and the plasma metabolites in patients with impaired fasting glucose or type 2 diabetes and healthy controls. In all, 65 nononobese patients (aged 30-70 years) with impaired fasting glucose or type 2 diabetes and 65 nonobese sex-matched healthy controls were included, and fasting peripheral blood mononuclear cell and plasma metabolomes were profiled. The diabetic or impaired fasting glucose patients showed higher circulating and peripheral blood mononuclear cell lipoprotein phospholipase A2 activities, high-sensitivity C-reactive protein and tumour necrosis factor-α than controls. Compared with controls, impaired fasting glucose or diabetic subjects showed increases in 11 peripheral blood mononuclear cell metabolites: six amino acids (valine, leucine, methionine, phenylalanine, tyrosine and tryptophan), l-pyroglutamic acid, two fatty acid amides containing palmitic amide and oleamide and two lysophosphatidylcholines. In impaired fasting glucose or diabetic patients, peripheral blood mononuclear cell lipoprotein phospholipase A2 positively associated with peripheral blood mononuclear cell lysophosphatidylcholines and circulating inflammatory markers, including tumour necrosis factor-α, high-sensitivity C-reactive protein and lipoprotein phospholipase A2 activities. In plasma metabolites between patients and healthy controls, we observed significant increases in only three amino acids (proline, valine and leucine) and decreases in only five lysophosphatidylcholines. This study demonstrates significant differences in the peripheral blood mononuclear cell metabolome in patients with impaired fasting glucose or diabetes compared with healthy controls. These differences were greater than those observed in the plasma metabolome. These data suggest peripheral blood mononuclear cells as a useful tool to better understand the inflammatory pathophysiology of diabetes.

Simultaneous determination of flumatinib and its two major metabolites in plasma of chronic myelogenous leukemia patients by liquid chromatography–tandem mass spectrometry

Flumatinib is an antineoplastic tyrosine kinase inhibitor used for the treatment of chronic myelogenous leukemia (CML). Its major metabolites in the circulation are N-desmethyl flumatinib (M1) and amide hydrolysis product (M3). To investigate the pharmacokinetics of flumatinib in CML patients, a simple, specific and rapid liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of flumatinib and its two major metabolites in patient plasma. After a simple, one-step protein precipitation with methanol, flumatinib, its two metabolites, and internal standard (HHGV-E) were separated on a C18 column using an isocratic mobile phase of methanol:5mM ammonium acetate:formic acid (60:40:0.4, v/v/v). A total chromatographic run time of 4.2min was achieved. The detection was performed in multiple reaction monitoring mode, using the transitions of m/z 563→m/z 463 for flumatinib, m/z 549→m/z 463 for M1, m/z 303→m/z 175 for M3, and m/z 529→m/z 429 for HHGV-E. The method was linear over the concentration ranges of 0.400–400ng/mL for flumatinib, 0.100–100ng/mL for M1, and 0.200–200ng/mL for M3, using only 50μL of plasma. The intra- and inter-day precisions were less than 8.5% for flumatinib, 9.8% for M1, and 10.6% for M3 in terms of the relative standard deviation. The accuracy was within ±2.2% for flumatinib, ±6.0% for M1, and ±9.9% for M3 in terms of relative error. The validated method was successfully applied to clinical pharmacokinetic studies of flumatinib mesylate in CML patients following oral administration at all dosage regimens.