Integrated metagenomics and metabolomics analysis illustrates the systemic impact of the gut microbiota on host metabolism after bariatric surgery.

AIM: To explore how bariatric surgery (BS) modified the obesity-associated gut microbiome, the host metabolome, and their interactions in obese Korean patients. MATERIALS AND METHODS: Stool and fasting blood samples were obtained before and 1, 3, 6, and 12 months after BS from 52 patients enrolled in the Korean Obesity Surgical Treatment Study. We analysed the gut microbiome by 16S rRNA gene sequencing and the serum metabolome, including bile acids, by nuclear magnetic resonance spectroscopy and ultrahigh-performance liquid chromatography/triple quadrupole mass spectrometry. RESULTS: Stool metagenomics showed that 27 microbiota were enriched and 14 microbiota were reduced after BS, whereas the abundances and diversity of observed features were increased. The levels of branched-chain amino acids and metabolites of energy metabolism in serum were decreased after surgery, whereas the levels of metabolites related to microbial metabolism, including dimethyl sulphone, glycine, and secondary bile acids, were increased in the serum samples. In addition, we found notable mutual associations among metabolites and gut microbiome changes attributed to BS. CONCLUSIONS: Changes in the gut microbiome community and systemic levels of amino acids and sugars were directly derived from anatomical changes in the gastrointestinal tract after BS. We hypothesized that the observed increases in microbiome-related serum metabolites were a result of complex and indirect changes derived from BS. Ethnic-specific environmental or genetic factors could affect Korean-specific postmetabolic modification in obese patients who undergo BS.

Comparable Plasma Lipid Changes in Patients with High-Grade Cervical Intraepithelial Neoplasia and Patients with Cervical Cancer.

Cervical cancer is the fourth most prevalent cancer among women worldwide and usually develops from cervical intraepithelial neoplasia (CIN). In the present study, we compared alterations in lipids associated with high-grade CIN and cervical cancer with those associated with a normal status and low-grade CIN by performing global lipid profiling on plasma (66 healthy controls and 55 patients with CIN1, 44 with CIN2/3, and 60 with cervical cancer) using ultraperformance liquid chromatography/quadrupole time-of-flight mass spectrometry. We identified 246 lipids and found 31 lipids with similar alterations in both high-grade CIN and cervical cancer. Among these 31 lipids, four lipid classes (phosphatidylcholine, phosphatidylethanolamine, diglyceride, and free fatty acids) were identified as the major lipid classes with significant differences in the patients with CIN2/3 and cervical cancer compared to the healthy controls and the patients with CIN1. Lipid metabolites belonging to the same classes were positively correlated with each other. High-grade CIN and cervical cancer induce comparable changes in lipid levels, which are closely related to the development of cervical tumors. These results suggest that lipid profiling is a useful method for monitoring progression to cervical cancer.

Circulating lipidomic alterations in obese and non-obese subjects with non-alcoholic fatty liver disease.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) affects obese and non-obese individuals. However, mechanisms underlying non-obese non-alcoholic steatohepatitis (NASH) remain unclear. AIMS: To attempt to identify metabolic perturbations associated with non-obese and obese NAFLD using a lipidomics approach. METHODS: A cross-sectional analysis of 361 subjects with biopsy-proven NAFLD (157 NAFL and 138 NASH) and healthy controls (n = 66) was performed. Individuals were categorised as obese or non-obese based on the Asian cut-off for body mass index. Circulating lipidomic profiling of sera was performed based on the histological severity of NAFLD. Circulating lipidomic alterations were validated with an independent validation set (154 NAFLD subjects [93 NAFL and 61 NASH] and 21 healthy controls). RESULTS: Saturated sphingomyelin (SM) species were significantly associated with visceral adiposity in non-obese NAFLD (SM d38:0; P < 0.001) but not in obese NAFLD. Additionally, SM levels were significantly associated with systemic and adipose tissue insulin resistance (SM d38:0; P = 0.002 and <0.001, respectively). Five potential lipid metabolites for non-obese subjects and seven potential lipids for obese subjects were selected to predict NAFLD and NASH. These lipid combinations showed good diagnostic performance for non-obese (area under the curve [AUC] for NAFLD/NASH = 0.916/0.813) and obese (AUC for NAFLD/NASH = 0.967/0.812) subjects. Moreover, distinctly altered patterns of diacylglycerol (DAG), triacylglycerol (TAG) and SM levels were confirmed in the validation set depending on the histological severity of NAFLD. CONCLUSION: Non-obese and obese NAFLD subjects exhibit unique circulating lipidomic signatures, including DAGs, TAGs and SMs. These lipid combinations may be useful biomarkers for non-obese and obese NAFLD patients.

Urinary Metabolomic Profiling Analysis and Evaluation of the Effect of Ecklonia cava Extract Intake.

Metabolomics is a powerful tool for the investigation of interactions between diet, nutrients, and human metabolism. Ecklonia cava is an edible brown alga that is abundantly found in Korea and Japan and contains unique polyphenols referred to as phlorotannins. However, there are few metabolomics studies related to the effects of polyphenols in humans. In this study, we performed a mass spectrometry-based metabolomics analysis of urine samples from participants with a body mass index (BMI) higher than 25 kg/m2 and lower than 30 kg/m2 to investigate the effects of the intake of seapolynol isolated from E. cava. Metabolomic profiling showed that the levels of riboflavin, urocanic acid, 5-hydroxy-6-methoxyindole glucuronide, and guanidino valeric acid were significantly increased in the seapolynol intake group compared with the placebo group. A correlation analysis was performed to identify the association between the metabolites' levels and clinical characteristics related to body fat. Among the metabolites whose concentrations changed in the seapolynol intake group, riboflavin was associated with BMI, body weight, fat mass, and percent body fat. These findings suggest that the decreased body fat induced by the intake of seapolynol is related to an increase in the antioxidant effect of riboflavin.

Metabolic phenotyping of human atherosclerotic plaques: Metabolic alterations and their biological relevance in plaque-containing aorta.

BACKGROUND AND AIMS: Atherosclerosis is a chronic inflammatory disease characterized by thickening of the arterial wall. However, a limited number of studies have been conducted on metabolic profiling of human aortic tissue. METHODS: We applied liquid chromatography/mass spectrometry to perform global and targeted profiling of plaque-containing aortic tissue. The aorta samples included plaque-containing (n = 18) and control plaque-free (n = 24) aortic tissue from patients undergoing aortic surgery. RESULTS: The metabolic patterns of atherosclerotic and control vessels were significantly different. Metabolites in the purine and glutathione pathways showed dysregulation of oxidative stress in plaques, and levels of glucosylceramide, tryptophan, and kynurenine, which are related to inflammation, were also altered. Interestingly, an increased level of quinic acid was observed in plaques (p < 0.000), and we demonstrated an inhibitory effect of quinic acid on inflammatory activation and oxidative stress in macrophages. CONCLUSIONS: Our study provides insight into the disease mechanism and potential markers of atherosclerosis through comprehensive metabolic profiling of human aortic tissue samples containing plaque.

A metabolomics-driven approach reveals metabolic responses and mechanisms in the rat heart following myocardial infarction.

BACKGROUND: Myocardial infarction (MI) is caused by myocardial necrosis resulting from prolonged ischemia. However, the biological mechanisms underlying MI remain unclear. METHODS: We evaluated metabolic and lipidomic changes in rat heart tissue from sham and MI at 1h, 1day and 10day after coronary ligation, using global profiling based on metabolomics. RESULTS: A time-dependent increase or decrease in polar and lipid metabolite levels was measured. The S-adenosylmethionine (SAM) concentration and the SAM/S-adenosylhomocysteine (SAH) ratio gradually decreased in a time-dependent manner and were significantly downregulated 10days after MI. Transcriptome analysis revealed that the levels of coenzyme Q (Coq)-3 and Coq5, both of which are SAM-dependent methyltransferases, were decreased in the MI groups. These results suggested that dysregulation of SAM may be related to down regulated COQ biosynthetic pathway. In addition, short-chain (C3) and medium-chain (C4-C12) acylcarnitine levels gradually decreased, whereas long-chain acylcarnitine (C14-18) levels increased, owing to a defect in ß-oxidation during ischemia. These changes are related to energy-dependent metabolic pathways, and a subsequent decrease in adenosine triphosphate concentration was observed. CONCLUSIONS: The comprehensive integration of various omics data provides a novel means of understanding the underlying pathophysiological mechanisms of MI.

Integrated omics-analysis reveals Wnt-mediated NAD+ metabolic reprogramming in cancer stem-like cells.

Abnormal tumor cell metabolism is a consequence of alterations in signaling pathways that provide critical selective advantage to cancer cells. However, a systematic characterization of the metabolic and signaling pathways altered in cancer stem-like cells (CSCs) is currently lacking. Using nuclear magnetic resonance and mass spectrometry, we profiled the whole-cell metabolites of a pair of parental (P-231) and stem-like cancer cells (S-231), and then integrated with whole transcriptome profiles. We identified elevated NAAD+ in S-231 along with a coordinated increased expression of genes in Wnt/calcium signaling pathway, reflecting the correlation between metabolic reprogramming and altered signaling pathways. The expression of CD38 and ALP, upstream NAAD+ regulatory enzymes, was oppositely regulated between P- and S-231; high CD38 strongly correlated with NAADP in P-231 while high ALP with NAAD+ levels in S-231. Antagonizing Wnt activity by dnTCF4 transfection reversed the levels of NAAD+ and ALP expression in S-231. Of note, elevated NAAD+ caused a decrease of cytosolic Ca2+ levels preventing calcium-induced apoptosis in nutrient-deprived conditions. Reprograming of NAD+ metabolic pathway instigated by Wnt signaling prevented cytosolic Ca2+ overload thereby inhibiting calcium-induced apoptosis in S-231. These results suggest that "oncometabolites" resulting from cross talk between the deranged core cancer signaling pathway and metabolic network provide a selective advantage to CSCs.

Changes in one-carbon metabolism after duodenal-jejunal bypass surgery.

Bariatric surgery alleviates obesity and ameliorates glucose tolerance. Using metabolomic and proteomic profiles, we evaluated metabolic changes in serum and liver tissue after duodenal-jejunal bypass (DJB) surgery in rats fed a normal chow diet. We found that the levels of vitamin B12 in the sera of DJB rates were decreased. In the liver of DJB rats, betaine-homocysteine S-methyltransferase levels were decreased, whereas serine, cystathionine, cysteine, glutathione, cystathionine ß-synthase, glutathione S-transferase, and aldehyde dehydrogenase levels were increased. These results suggested that DJB surgery enhanced trans-sulfuration and its consecutive reactions such as detoxification and the scavenging activities of reactive oxygen species. In addition, DJB rats showed higher levels of purine metabolites such as ATP, ADP, AMP, and inosine monophosphate. Decreased guanine deaminase, as well as lower levels of hypoxanthine, indicated that DJB surgery limited the purine degradation process. In particular, the AMP/ATP ratio and phosphorylation of AMP-activated protein kinase increased after DJB surgery, which led to enhanced energy production and increased catabolic pathway activity, such as fatty acid oxidation and glucose transport. This study shows that bariatric surgery altered trans-sulfuration and purine metabolism in the liver. Characterization of these mechanisms increases our understanding of the benefits of bariatric surgery.

Discovery of substituted pyrazol-4-yl pyridazinone derivatives as novel c-Met kinase inhibitors.

A series of pyridazin-3-one substituted with morpholino-pyrimidine derivatives was synthesized and evaluated as tyrosine kinase inhibitors against c-Met enzyme, and anti-proliferative activities of Hs746T human gastric cancer cell line. Most of compounds exhibited good biological activity, while compound 10, 12a, 14a displayed excellent c-Met enzyme inhibitory activities and Hs746T cell-based activities.

Noninvasive diagnosis and evaluation of curative surgery for gastric cancer by using NMR-based metabolomic profiling.

BACKGROUND: Mass screening for gastric cancer (GC), particularly using endoscopy, may not be the most practical approach as a result of its high cost, lack of acceptance, and poor availability. Thus, novel markers that can be used in cost-effective diagnosis and noninvasive screening for GC are needed. METHODS: A total of 154 urine samples from GC patients and healthy individuals and 30 pairs of matched tumor and normal stomach tissues were collected. Multivariate analysis was performed on urinary and tissue metabolic profiles acquired using (1)H nuclear magnetic resonance and (1)H high-resolution magic angle spinning spectroscopy, respectively. In addition, metabolic profiling of urine from GC patients after curative surgery was performed. RESULTS: Multivariate statistical analysis showed significant separation in the urinary and tissue data of GC patients and healthy individuals. The metabolites altered in the urine of GC patients were related to amino acid and lipid metabolism, consistent with changes in GC tissue. In the external validation, the presence of GC (early or advanced) from the urine model was predicted with high accuracy, which showed much higher sensitivity than carbohydrate antigen 19-9 and carcinoembryonic antigen. Furthermore, 4-hydroxyphenylacetate, alanine, phenylacetylglycine, mannitol, glycolate, and arginine levels were significantly correlated with cancer T stage and, together with hypoxanthine level, showed a recovery tendency toward healthy controls in the postoperative samples compared to the preoperative samples. CONCLUSIONS: An urinary metabolomics approach may be useful for the effective diagnosis of GC.

Potential metabolomic biomarkers for evaluation of adriamycin efficacy using a urinary 1H-NMR spectroscopy.

A metabolomics approach using proton nuclear magnetic resonance (NMR) was applied to investigate metabolic alterations following adriamycin (ADR) treatment for gastric adenocarcinoma. After BALB/c-nu/nu mice were implanted with human gastric adenocarcinoma, ADR (1 or 3 mg kg(-1) per day) was intraperitoneally administered for 5 days. Urine was collected on days 2 and 5 and analyzed by NMR. The levels of trimethylamine oxide (TMAO, ×0.3), hippurate (×0.3) and taurine (×0.6) decreased significantly (P < 0.05), whereas the levels of 3-indoxylsulfate (×12.6), trigonelline (×1.5), citrate (×2.5), trimethylamine (TMA, ×2.0) and 2-oxoglutarate (×2.3) increased significantly (P < 0.05) in the tumor model. After ADR treatment, TMAO, hippuarte and taurine were increased significantly on day 5 compared with those of the tumor model. The levels of 2-oxoglutarate, 3-indoxylsulfate, trigonelline, TMA and citrate, which increased in the tumor model, significantly decreased to those of normal control by ADR treatment. Furthermore, the ratio between TMA and TMAO was dramatically altered in both tumor and ADR-treated groups. Overall, metabolites such as TMAO, TMA, 3-indoxylsulfate, hippurate, trigonelline, citrate and 2-oxoglutarate related to the tricarboxylic acid (TCA) cycle might be considered as therapeutic targets to potentiate the efficacy of ADR. Thus, these results suggest that the metabolomics analysis of tumor response to ADR treatment may be applicable for demonstrating the efficacy of anticancer agent, ADR and treatment adaptation.

Characterization of dandelion species using 1H NMR- and GC-MS-based metabolite profiling.

Taraxacum, known as dandelion, is a large genus of plants in the family Asteraceae. Pharmacological studies have shown that these plants display a wide variety of medicinal properties because Taraxacum extracts contain many pharmacologically active metabolites that display anti-inflammatory, antinociceptive, antioxidant, and anticancer activity. Each plant species displays several different natural constituents, the majority of which have not been studied as no global metabolite screen of the diverse Taraxacum species has been performed. In this study, we investigated the metabolite difference in three species of Taraxacum (T. coreanum, T. officinale, and T. platycarpum) by (1)H NMR spectroscopy and gas chromatography-mass spectrometry (GC-MS) coupled with multivariate statistical analyses. The aim of this study was to identify the different chemical compositions of the polar and nonpolar extracts in these species. A partial least-squares discriminant analysis showed a significantly higher separation among nonpolar extracts (mainly fatty acids and sterols) compared to polar extracts (mainly amino acids, organic acids, and sugars) between these species. A one-way ANOVA was performed to statistically certify the metabolite differences of these nonpolar extracts. Taken together, these data suggest that a metabolomic approach using combined (1)H NMR and GC-MS analysis is an effective analytical method to differentiate biochemical compositions among different species in plants.

1H NMR-based metabolomic study on resistance to diet-induced obesity in AHNAK knock-out mice.

AHNAK is a giant protein of approximately 700 kDa identified in human neuroblastomas and skin epithelial cells. Recently, we found that AHNAK knock-out (AHNAK(-/-)) mice have a strong resistance to high-fat diet-induced obesity. In this study, we applied (1)H NMR-based metabolomics with multivariate statistical analysis to compare the altered metabolic patterns detected in urine from high-fat diet (HFD) fed wild-type and AHNAK(-/-) mice and investigate the mechanisms underlying the resistance to high-fat diet-induced obesity in AHNAK(-/-) mice. In global profiling, principal components analysis showed a clear separation between the chow diet and HFD groups; wild-type and AHNAK(-/-) mice were more distinctly separated in the HFD group compared to the chow diet group. Based on target profiling, the urinary metabolites of HFD-fed AHNAK(-/-) mice gave higher levels of methionine, putrescine, tartrate, urocanate, sucrose, glucose, threonine, and 3-hydroxyisovalerate. Furthermore, two-way ANOVAs indicated that diet type, genetic type, and their interaction (gene × diet) affect the metabolite changes differently. Most metabolites were affected by diet type, and putrescine, threonine, urocanate, and tartrate were also affected by genetic type. In addition, cis-aconitate, succinate, glycine, histidine, methylamine (MA), phenylacetylglycine (PAG), methionine, putrescine, uroconate, and tartrate showed interaction effects. Through the pattern changes in urinary metabolites of HFD-fed AHNAK(-/-) mice, our data suggest that the strong resistance to HFD-induced obesity in AHNAK(-/-) mice comes from perturbations of amino acids, such as methionine, putrescine, threonine, and histidine, which are related to fat metabolism. The changes in metabolites affected by microflora such as PAG and MA were also observed. In addition, resistance to obesity in HFD-fed AHNAK(-/-) mice was not related to an activated tricarboxylic acid cycle. These findings demonstrate that (1)H NMR-based metabolic profiling of urine is suitable for elucidating possible biological pathways perturbed by functional loss of AHNAK on HFD feeding and could elucidate the mechanism underlying the resistance to high-fat diet-induced obesity in AHNAK(-/-) mice.

Toxicometabolomics of urinary biomarkers for human gastric cancer in a mouse model.

Toxicometabolomics of urinary biomarkers for human gastric cancer in a mouse model was investigated using (1)H-nuclear magnetic resonance (NMR) spectroscopy. A human gastric adenocarcinoma cell line (1 × 10(7) cells/ml) was grafted onto the skin of the back of intact male BALB/c-nu/nu mice. After the xenografted tumors developed, urine was collected and analyzed for endogenous metabolites. Global profiling combined with principal components analysis (PCA), partial least squares-discriminant analysis (PLS-DA), and orthogonal projections to latent squares-discriminant analysis (OPLS-DA) showed distinct separation of clusters between control and tumor-bearing mice. Targeted profiling revealed significant changes in trimethylamine oxide (TMAO), 3-indoxylsulfate, hippurate, and citrate levels in mice carrying human gastric cancer cells compared to normal mice. The levels of TMAO (0.41-fold) and hippurate (0.26-fold) in tumor-bearing mice were significantly decreased, whereas the levels of 3-indoxylsulfate (3.39-fold), 2-oxoglutarate (2.32-fold), and citrate (1.9-fold) were significantly increased in urine samples of tumor-bearing mice. Data suggest that TMAO, hippurate, 3-indoxylsulfate, 2-oxoglutarate, and citrate may serve as useful urinary biomarkers for gastric tumorigenesis in a mouse model.

Enantioselective synthesis of beta-iodo Morita-Baylis-Hillman esters by a catalytic asymmetric three-component coupling reaction.

A catalytic route toward chiral Morita-Baylis-Hillman esters by asymmetric coupling between alpha,beta-acetylenic esters, aldehydes, and trimethylsilyl iodide has been developed (see scheme). The reaction proceeds with high to excellent enantioselectivities, and the products can be transformed into beta-branched derivatives in a single step and with excellent retention of configuration. TMS = trimethylsilyl.

A highly E-stereoselective approach to beta-iodo Morita-Baylis-Hillman esters: synthesis of secokotomolide A.

A geometric-selective synthesis of (E)-beta-iodo Morita-Baylis-Hillman esters has been developed through a three-component aldol-type reaction using BF3.Et2O and TMS-I. The synthetic utility of the (E)-beta-iodo Morita-Baylis-Hillman esters was demonstrated in the first reported short synthesis of secokotomolide A.