Identification of proline, 1-pyrroline-5-carboxylate and glutamic acid as biomarkers of depression reflecting brain metabolism using carboxylomics, a new metabolomics method.

AIM: Depression is a psychiatric disease which is accompanied by metabolic disorder. Though depression has been widely studied, its metabolism is yet to be illustrated. We aimed to manifest the underlying mechanisms to diagnose depression. METHODS: One hundred thirty serum samples, including 65 patients and 65 healthy controls from different hospitals (training and validation cohorts), were recruited into the research. Sensitive Profiling for ChemoSelective Derivatization Carboxylomics (SPCSDCarboxyl) was applied to deeply hunt for the differential metabolites. Then, the serum, CSF, and hippocampus from depression rat models (CUMS group) were used to further confirm the results. Additionally, the co-occurrence between enzymes and biomarkers, as well as the combinatorial marker panel and the correlation of biomarkers among serum, CSF, or hippocampus were elucidated. RESULTS: Two hundred eight metabolites were identified from the sera of patients. Proline, 1-pyrroline-5-carboxylate (P5C), and glutamic acid could discriminate patients from healthy humans and were confirmed to be the potential biomarkers. After further validation through CUMS rats, proline, and P5C were enriched, while glutamic acid was depleted in the CUMS group. The co-occurrence analysis of enzymes and biomarkers indicated that they could be used for the diagnosis of depression. Moreover, the combinatorial marker panel and the correlation analysis of biomarkers between serum and CSF or between serum and hippocampus revealed that serum could be an alternative approach to directly reflect the potential physiological mechanisms and diagnose depression instead of brain samples. CONCLUSION: These integrated methods may facilitate the identification of biomarkers and help manifest the underlying mechanisms of depression.

[LC-MS fingerprint and multi-indicator components analysis of classical formula Gualou Xiebai Banxia Decoction].

This study developed an optimal pre-processing technique for the reference substance of the classic formula Gualou Xiebai Banxia Decoction(GXBD) and established a comprehensive quality control method for GXBD reference substance to provide a reference for its overall quality evaluation. The authors prepared 15 batches of GXBD samples and innovatively used the extracted ion chromatogram under the base peak chromatogram mode to establish a liquid chromatography-mass spectrometry(LC-MS) fingerprint, identify characteristic peaks, and perform quantitative analysis of indicator components. The yield of the 15 batches of GXBD samples ranged from 50.28% to 76.20%. In the positive ion mode, 12 common characteristic peaks were detected in the LC-MS fingerprint, and the structures of five common peaks were identified by comparison with reference standards. The similarity between the fingerprint profiles of different batches of samples and the reference fingerprint profile ranged from 0.920 to 0.984. Finally, liquid chromatography-triple quadrupole mass spectrometry(LC-QQQ/MS) in multiple reaction monitoring(MRM) mode was used to determine the content of eight indicator components in GXBD, including loliolide, chrysoeriol, rutin, cucurbitacin D, macrostemonoside Ⅰ, 25S-timosaponin B Ⅱ, 25R-timosaponin B Ⅱ, and peptide proline-tryptophan-valine-proline-glycine(PWVPG). The method established in this study can reduce matrix interference in the compound, and it has good accuracy, stability, and practical value. It effectively reflects the quality attributes of GXBD samples and can be used for the comprehensive quality control of GXBD.

Umbilical cord blood metabolomics: association with intrauterine hyperglycemia.

BACKGROUND: Intrauterine hyperglycemia can harm a fetus's growth and development, and this can be seen in the umbilical cord blood metabolism disorder. However, the metabolites and metabolic mechanisms involved in the condition remain unknown. METHODS: Targeted metabolomics using liquid chromatography and MetaboAnalyst were conducted in this study to explore differences in metabolites and metabolic pathways between individuals with hyperglycemia or well-controlled gestational diabetes mellitus (GDM) and healthy controls. RESULTS: Univariate analysis found that the hyperglycemic and healthy control groups differed in 30 metabolites, while the well-controlled GDM and the healthy control groups differed only in three metabolites-ursodeoxycholic acid, docosahexaenoic acid, and 8,11,14-eicosatrienoic acid. Most of these metabolic variations were negatively associated with neonatal weights. Further research showed that the variations in the metabolites were primarily associated with the metabolic pathways of linoleic acid (LA) and alpha-linolenic acid (ALA). CONCLUSION: Gestational hyperglycemia and well-controlled GDM, which may play a major role by inhibiting the LA and ALA metabolic pathways, have detrimental effects on cord blood metabolism. IMPACT: The main point of this paper is that intrauterine hyperglycemia has a negative effect on cord blood metabolism mainly through the linoleic acid and alpha-linolenic acid metabolic pathways. This is a study to report a new association between well-controlled GDM and cord blood metabolism. This study provides a possible explanation for the association between intrauterine hyperglycemia and neonatal adverse birth outcomes.

In Colorectal Cancer Cells With Mutant KRAS, SLC25A22-Mediated Glutaminolysis Reduces DNA Demethylation to Increase WNT Signaling, Stemness, and Drug Resistance.

BACKGROUND & AIMS: Mutant KRAS promotes glutaminolysis, a process that uses steps from the tricarboxylic cycle to convert glutamine to α-ketoglutarate and other molecules via glutaminase and SLC25A22. This results in inhibition of demethylases and epigenetic alterations in cells that increase proliferation and stem cell features. We investigated whether mutant KRAS-mediated glutaminolysis affects the epigenomes and activities of colorectal cancer (CRC) cells. METHODS: We created ApcminKrasG12D mice with intestine-specific knockout of SLC25A22 (ApcminKrasG12DSLC25A22fl/fl mice). Intestine tissues were collected and analyzed by histology, immunohistochemistry, and DNA methylation assays; organoids were derived and studied for stem cell features, along with organoids derived from 2 human colorectal tumor specimens. Colon epithelial cells (1CT) and CRC cells (DLD1, DKS8, HKE3, and HCT116) that expressed mutant KRAS, with or without knockdown of SLC25A22 or other proteins, were deprived of glutamine or glucose and assayed for proliferation, colony formation, glucose or glutamine consumption, and apoptosis; gene expression patterns were analyzed by RNA sequencing, proteins by immunoblots, and metabolites by liquid chromatography-mass spectrometry, with [U-13C5]-glutamine as a tracer. Cells and organoids with knocked down, knocked out, or overexpressed proteins were analyzed for DNA methylation at CpG sites using arrays. We performed immunohistochemical analyses of colorectal tumor samples from 130 patients in Hong Kong (57 with KRAS mutations) and Kaplan-Meier analyses of survival. We analyzed gene expression levels of colorectal tumor samples in The Cancer Genome Atlas. RESULTS: CRC cells that express activated KRAS required glutamine for survival, and rapidly incorporated it into the tricarboxylic cycle (glutaminolysis); this process required SLC25A22. Cells incubated with succinate and non-essential amino acids could proliferate under glutamine-free conditions. Mutant KRAS cells maintained a low ratio of α-ketoglutarate to succinate, resulting in reduced 5-hydroxymethylcytosine-a marker of DNA demethylation, and hypermethylation at CpG sites. Many of the hypermethylated genes were in the WNT signaling pathway and at the protocadherin gene cluster on chromosome 5q31. CRC cells without mutant KRAS, or with mutant KRAS and knockout of SLC25A22, expressed protocadherin genes (PCDHAC2, PCDHB7, PCDHB15, PCDHGA1, and PCDHGA6)-DNA was not methylated at these loci. Expression of the protocadherin genes reduced WNT signaling to ß-catenin and expression of the stem cell marker LGR5. ApcminKrasG12DSLC25A22fl/fl mice developed fewer colon tumors than ApcminKrasG12D mice (P < .01). Organoids from ApcminKrasG12DSLC25A22fl/fl mice had reduced expression of LGR5 and other markers of stemness compared with organoids derived from ApcminKrasG12D mice. Knockdown of SLC25A22 in human colorectal tumor organoids reduced clonogenicity. Knockdown of lysine demethylases, or succinate supplementation, restored expression of LGR5 to SLC25A22-knockout CRC cells. Knockout of SLC25A22 in CRC cells that express mutant KRAS increased their sensitivity to 5-fluorouacil. Level of SLC25A22 correlated with levels of LGR5, nuclear ß-catenin, and a stem cell-associated gene expression pattern in human colorectal tumors with mutations in KRAS and reduced survival times of patients. CONCLUSIONS: In CRC cells that express activated KRAS, SLC25A22 promotes accumulation of succinate, resulting in increased DNA methylation, activation of WNT signaling to ß-catenin, increased expression of LGR5, proliferation, stem cell features, and resistance to 5-fluorouacil. Strategies to disrupt this pathway might be developed for treatment of CRC.

Aspirin Reduces Colorectal Tumor Development in Mice and Gut Microbes Reduce its Bioavailability and Chemopreventive Effects.

BACKGROUND & AIMS: Alterations in the intestinal microbiota affect development of colorectal cancer and drug metabolism. We studied whether the intestinal microbiota affect the ability of aspirin to reduce colon tumor development in mice. METHODS: We performed studies with APCmin/+ mice and mice given azoxymethane and dextran sulfate sodium to induce colorectal carcinogenesis. Some mice were given antibiotics to deplete intestinal microbes, with or without aspirin, throughout the entire experiment. Germ-free mice were studied in validation experiments. Colon tissues were collected and analyzed by histopathology, quantitative reverse-transcription polymerase chain reaction, and immunoblots. Blood samples and gut luminal contents were analyzed by liquid chromatography/mass spectrometry and an arylesterase activity assay. Fecal samples were analyzed by 16S ribosomal RNA gene and shotgun metagenome sequencing. RESULTS: Administration of aspirin to mice reduced colorectal tumor number and load in APCmin/+ mice and mice given azoxymethane and dextran sulfate sodium that had been given antibiotics (depleted gut microbiota), but not in mice with intact microbiota. Germ-free mice given aspirin developed fewer colorectal tumors than conventionalized germ-free mice given aspirin. Plasma levels of aspirin were higher in mice given antibiotics than in mice with intact gut microbiota. Analyses of luminal contents revealed that aerobic gut microbes, including Lysinibacillus sphaericus, degrade aspirin. Germ-free mice fed L sphaericus had lower plasma levels of aspirin than germ-free mice that were not fed this bacterium. There was an inverse correlation between aspirin dose and colorectal tumor development in conventional mice, but this correlation was lost with increased abundance of L sphaericus. Fecal samples from mice fed aspirin were enriched in Bifidobacterium and Lactobacillus genera, which are considered beneficial, and had reductions in Alistipes finegoldii and Bacteroides fragili, which are considered pathogenic. CONCLUSIONS: Aspirin reduces development of colorectal tumors in APCmin/+ mice and mice given azoxymethane and dextran sulfate sodium, depending on the presence of intestinal microbes. L sphaericus in the gut degrades aspirin and reduced its chemopreventive effects in mice. Fecal samples from mice fed aspirin were enriched in beneficial bacteria, with reductions in pathogenic bacteria.

Metabolomics reveals a correlation between hydroxyeicosatetraenoic acids and allergic asthma: Evidence from three years' immunotherapy.

BACKGROUND: Subcutaneous immunotherapy (SCIT) is an effective, safe, preventative treatment for allergic asthma; however, potential biomarkers for monitoring SCIT have rarely been reported. OBJECTIVE: Metabolomics was utilized for the discovery of new biomarkers and analyzing disease pathophysiology of allergic asthma, and it was also applied to determine the metabolomic profiles of serum samples from children with asthma undergoing SCIT and identify potential biomarkers for allergic asthma and its therapeutic monitoring. METHODS: Untargeted metabolomics using ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry was performed on 15 asthmatic and 15 healthy pediatric sera to profile carboxylic acids. Statistical analysis combined with pathway enrichment analysis was applied to identify potential biomarkers. Then, targeted metabolomics was performed to study longitudinal changes of eicosanoid profiles on sera from 20 participants with asthma who received SCIT at baseline, 6 months, one, two, and three years (ChiCTR-DDT-13003728). RESULTS: Metabolomic analysis revealed that levels of eicosanoids, particularly 12(S)-hydroxyeicosatetraenoic acid (HETE; AUC = 0.94, p < .0001) and 15(S)-HETE (AUC = 0.89, p = .0028), metabolized from arachidonic acid by lipoxygenase and glutathione peroxidase enzymes, were significantly higher in asthma group than in healthy individuals. Furthermore, levels of these important metabolites increased in the first year of SCIT treatment and then decreased from years one to three, being significantly lower after three years of treatment than baseline levels. CONCLUSION: 12(S)- and 15(S)-HETEs are potential biomarkers to participate in the pathogenesis and treatment of allergic asthma. Moreover, these metabolites may be a new target for biological indicators to monitor the therapeutic effect of SCIT, particularly in the setting of allergic asthma.

Analysis of serum polyunsaturated fatty acid metabolites in allergic bronchopulmonary aspergillosis.

BACKGROUND: The importance of lipid mediators in allergic diseases has been long recognized, whereas little is known about their role in allergic bronchopulmonary aspergillosis (ABPA). We investigated whether lipid mediators are associated with ABPA. METHODS: We recruited 12 ABPA patients, 23 asthma patients and 12 healthy control in our study. Serum of 11 ABPA patients were collected before and following treatment. 36 polyunsaturated fatty acid metabolites were measured in serum samples by using liquid chromatography-mass spectrometry. This study was approved by the Ethics Committee of the First Affiliated Hospital of Guangzhou Medical University, with ethics number GYFYY-2016-73. RESULTS: Levels of arachidonic acid (AA), 15(S)-hydroxyeicosatetraenoic acid (HETE), 12(S)-HETE, 8(S)-HETE, 5(S)-HETE, LTB4, PGB2, 12(S)-hydroxyeicosapentaenoic acid (HEPE), 12-hydro-xyheptadecatrienoic acid (HHTrE) were significantly higher in ABPA patients than that in HC groups. Compared with asthma group, ABPA group expressed lower levels of 15(S)-hy-droperoxyeicosatetraenoic acid (HPETE), 5(S)-HPETE, 13(S)-hydroperoxyoctadecadienoic acid (HPODE) and 9(S)-HPODE. In APBA patients, AA level was positively correlated with serumtotal IgE (tIgE). The levels of 12(S)-HPETE, 15(S)-HEPE and 12(S)-HEPE correlated with Asp-ergillus fumigatus specific IgE(A. fumigatus sIgE) positively. Peripheral blood eosinophilia correlated with high levels of 12(S)-HETE and 15(S)-HETE. In addition, the serum levels of15(S)-HETE and 12(S)-HETE in ABPA subjects both declined with the decrease of tIgE, A. fumigatus sIgE and sIgG concentrations after treatment. CONCLUSIONS: We present data regarding the role of polyunsaturated fatty acid metabolites in APBA for the first time. Most of the tested metabolites increased in ABPA when co-mpared with healthy controls and 15(S)-HETE and 12(S)-HETE may play a role in the pat-hogenesis of ABPA. These findings can provide new ideas for diagnosis, therapy and mon-itor of ABPA.

Identifying potential serum biomarkers of breast cancer through targeted free fatty acid profiles screening based on a GC-MS platform.

Recent advances suggest that abnormal fatty acid metabolism highly correlates with breast cancer, which provide clues to discover potential biomarkers of breast cancer. This study aims to identify serum free fatty acid (FFA) metabolic profiles and screen potential biomarkers for breast cancer diagnosis. Gas chromatography-mass spectrometry and our in-house fatty acid methyl ester standard substances library were combined to accurately identify FFA profiles in serum samples of breast cancer patients and breast adenosis patients (as controls). Potential biomarkers were screened by applying statistical analysis. A total of 18 FFAs were accurately identified in serum sample. Two groups of patients were correctly discriminated by the orthogonal partial least squares-discriminant analysis model based on FFA profiles. Seven FFA levels were significantly higher in serum from breast cancer patients than that in controls, and exhibited positive correlation with malignant degrees of disease. Furthermore, five candidates (palmitic acid, oleic acid, cis-8,11,14-eicosatrienoic acid, docosanoic acid and the ratio of oleic acid to stearic acid) were selected as potential serum biomarkers for differential diagnosis of breast cancer. Our study will help to reveal the metabolic signature of FFAs in breast cancer patients, and provides valuable information for facilitating clinical noninvasive diagnosis.

Integrated Proteomics, Biological Functional Assessments, and Metabolomics Reveal Toosendanin-Induced Hepatic Energy Metabolic Disorders.

Toosendanin (TSN), a compound from Melia toosendan, exhibits severe hepatotoxicity, which restricts its clinical application. However, the mechanism is not clear. Our previous research found that covalent modification of TSN for proteins might be a possible reason using human liver microsomes, and the glycolytic enzymes, triosephosphate isomerase 1 (TPIS) and α-enolase (ENOA), were responsible for the hepatotoxicity. In this study, we tried to prove these findings in cell and animal models by integration of proteomics, metabolomics, and biological methods. Proteomics analysis in rats showed that TPIS and ENOA were covalently modified by TSN reactive metabolites. The biological functional assessments revealed that the modifications inhibited the activity of TPIS and induced the activity of ENOA, in vitro and in vivo, followed by an increase in the level of cellular methylglyoxal, advanced glycation end products, and reactive oxygen species/superoxide, and the induction of mitochondrial dysfunction, which further inhibited oxidative phosphorylation and stimulated glycolysis. Furthermore, metabolomics demonstrated the decrease in the level of metabolites in the tricarboxylic acid cycle, fatty acid ß-oxidation, and amino acid metabolism; i.e., TSN induced hepatocyte energy metabolism disorder. In conclusion, these data suggest novel mechanistic insights into TSN-induced liver injury on the upstream level and provide valuable proteins and energy metabolic targets for diagnosis and therapy in the clinic.

Mushroom polysaccharides and jiaogulan saponins exert cancer preventive effects by shaping the gut microbiota and microenvironment in ApcMin/+ mice.

The incidence of colorectal cancer (CRC) is alarming among younger peoples. While no effective chemopreventive drug available in the market, researchers have been searching for alternative strategies against CRC that are in demand. Therefore, we tested the cancer-preventive properties of Ganoderma lucidum (Lingzhi) polysaccharides (GLP), along with the saponins extracted from Gynostemma pentaphyllum (GpS), an herbal tea with prebiotic-like effects. Here, we report that saponins from Gynostemma pentaphyllum (GpS) and polysaccharides from Ganoderma lucidum (GLP together with GpS) profoundly improved the inflamed gut barrier of ApcMin/+ mice by reducing polyps, shifting colonic M1 to M2 macrophages, positively reverting E-cadherin/N-cadherin ratio, and downregulating oncogenic signaling molecules. The treatments also markedly promoted short-chain fatty acids (SCFAs)-producing bacteria and abridged sulfate-reducing bacteria in a time-dependent manner. G-protein coupled-receptors were significantly stimulated in the treated mice, accompanied by the modulated expressions of histone deacetylases, anti-cancer gut hormone PYY, and PPAPγ. These findings suggest that some of the herbal medicinal foods could modulate the relationship between the host and the gut microbiota (GM) to exert their beneficial properties to the host. Our study also implicates that these dietary mushroom polysaccharides and the Gp saponins have the potential to be developed as new preventive medicines against CRC.

Polarity-Tuning Derivatization-LC-MS Approach for Probing Global Carboxyl-Containing Metabolites in Colorectal Cancer.

Carboxyl-containing metabolites (CCMs) widely exist in living systems and are the essential components for life. Global characteristics of CCMs in biological samples are critical for the understanding of physiological processes and the discovery for the onset of relevant diseases. However, their determination represents a challenge due to enormous polarity differences, structural diversity, high structural similarity, and poor ionization efficiency in mass spectrometry. Herein, 5-(diisopropylamino)amylamine (DIAAA) derivatization coupled with liquid chromatography-mass spectrometry (LC-MS) was developed for mapping the CCMs. With this methodology, the sensitivity was significantly enhanced. More importantly, the hydrophobicity of polar CCMs, amino acids, TCA cycle intermediates, and short-chain fatty acids and the hydrophilicity of low-polar CCMs, long-chain fatty acids, and bile acids were significantly increased, resulting in a remarkable separation efficiency for which 68 CCMs can be simultaneously determined. Furthermore, the polarity-tuning effect was confirmed to be induced by the different impacts of aliphatic chains and nitrogen atom in DIAAA, the latter existing as a cation in the acidic mobile phase, using different derivatization reagents. Finally, this derivatization method was utilized to hunt for the potential biomarkers in colorectal cancer (CRC) patients and 52 CCMs, related with several key metabolic pathways, including amino acids metabolism, TCA cycle, fatty acid metabolism, pyruvate metabolism, and gut flora metabolism were identified. This innovative polarity-tuning derivatization-LC-MS approach was proved to be a valuable tool for probing global metabolome with high separation efficiency and sensitivity in various biological samples.

Structure determination of two new indole-diterpenoids from Penicillium sp. CM-7 by NMR spectroscopy.

Two new indole-diterpenoids 4b-deoxy-1'-O-acetylpaxilline (1) and 4b-deoxypenijanthine A (2) were isolated from the fermentation broth and the mycelia of the soil fungus Penicillium sp. CM-7, along with three known structurally related compounds, 1'-O-acetylpaxilline (3), paspaline (4) and 3-deoxo-4b-deoxypaxilline (5). The structures of compounds 1 and 2 were elucidated by extensive spectroscopic methods, especially 2D NMR, and their absolute configurations were suggested on the basis of the circular dichroism spectral analysis and the NOESY data.

LPCAT2 inhibits colorectal cancer progression via the PRMT1/SLC7A11 axis.

Colorectal cancer (CRC) has a high degree of heterogeneity and identifying the genetic information of individual tumor cells could help enhance our understanding of tumor biology and uncover potential therapeutic targets for CRC. In this study, we identified LPCAT2+ tumor cell populations with less malignancy than LPCAT2- tumor cells in human and mouse CRC tissues using scRNA-seq. Combining in vitro and in vivo experiments, we found that LPCAT2 could inhibit the proliferation of CRC cells by inducing ferroptosis. Mechanistically, LPCAT2 arrested PRMT1 in cytoplasm of CRC cells via regulating acetylation of PRMT1 at the K145 site. In turn, PRMT1 enhanced SLC7A11 promoter activity. Thus, LPCAT2 attenuated the positive regulatory effect of PRMT1 on SLC7A11 promoter. Notably, SLC7A11 acts as a ferroptosis regulator. Furthermore, in LPCAT2 knockout mice (LPCAT2-/-) colon cancer model, we found that LPCAT2-/- mice exhibited more severe lesions, while PRMT1 or SLC7A11 inhibitors delayed the progression. Altogether, we elucidated that LPCAT2 suppresses SLC7A11 expression by inhibiting PRMT1 nuclear translocation, thereby inducing ferroptosis in CRC cells. Moreover, inhibitors of the PRMT1/SLC7A11 axis could delay tumor progression in CRC with low LPCAT2 expression, making it a potentially effective treatment for CRC.

Structural characterization, molecular dynamic simulation, and conformational visualization of a water-soluble glucan with high molecular weight from Gastrodia elata Blume.

Polysaccharides have been widely used in the development of natural drugs and health food. However, polysaccharide characterization lags due to inherently complicated features and the limitations of existing detection approaches. We aimed to provide new insight into the fine structure and conformational visualization of polysaccharides from Gastrodia elata Blume, a medicinal and edible plant. A water-soluble polysaccharide (GEP2-6) with the high molecular weight of 2.7 × 106 Da was first obtained, and its purity reached 99.2 %. Chemical and spectroscopic analyses jointly revealed that GEP2-6 was a glucan linked by α-(1 â†’ 4) and α-(1 â†’ 6) glycosidic bonds. After enzymolysis, the local structure of GEP2-6 included α-1,4-Glcp, α-1,6-Glcp, α-1,4,6-Glcp, and α-1-Glcp at a molar ratio of 31.27∶1.32∶1.08∶0.93. The glycosidic linkage pattern of repeating units was further simulated by a glycan database and spatial examination software. The good dissolution performance was interpreted by dynamics simulation and practical molecular characteristics. Spherical flexible chains and the porous stable conformation were corroborated using atomic force microscopy. In addition, GEP2-6 could effectively scavenge DPPH and hydroxyl radicals as a promising natural antioxidant. These efforts will contribute to the expansion of clinical applications of this G. elata polysaccharide and the structural elucidation for macromolecular polysaccharides combined with traditional and modern analysis techniques.

Ganoderma lucidum Polysaccharide Supplementation Significantly Activates T-Cell-Mediated Antitumor Immunity and Enhances Anti-PD-1 Immunotherapy Efficacy in Colorectal Cancer.

Ganoderma lucidum polysaccharide (GLP) is a prebiotic with immunomodulatory effects. However, the therapeutic potential of GLP in tumor immunotherapy has not been fully explored, especially in T cell-mediated antitumor immunity. In this study, we found that GLP significantly inhibited tumor growth and activated antitumor immunity in colorectal cancer (CRC). In the spleens and tumor tissues, the proportion of cytotoxic CD8+T cells and Th1 helper cells increased, while immunosuppressive Tregs decreased. Additionally, microbiota dysbiosis was alleviated by GLP, and short-chain fatty acid production was increased. Meanwhile, GLP decreased the ratio of kynurenine and tryptophan (Kyn/Trp) in the serum, which contributed to antitumor immunity of T cells. More importantly, the combination of GLP and the immune checkpoint inhibitor anti-PD-1 monoclonal antibody further enhanced the efficacy of anti-PD-1 immunotherapy. Thus, GLP as a prebiotic has the potential to be used in tumor immunotherapy.

Spatiotemporal changes in snow depth and the influence factors in China from 1979 to 2019.

Snow depth is an important parameter to characterize the characteristics of snow cover, and it is also one of the most sensitive response factors to regional climate change. However, the extent of snow depth variability and its driving mechanisms are still unknown in China. Therefore, in this study, we used the regression analysis, root-mean-square error analysis, anomalous year analysis, and correlation analysis methods to explore the spatiotemporal variation characteristics of snow depth in China from 1979 to 2019 based on the reanalysis snow depth dataset. The results show that (1) the snow distribution in China is obviously spatially heterogeneous, and the southeastern, western, and southern regions of the Qinghai-Tibet Plateau, northern Xinjiang, and northeastern China have high values of snow depth; (2) the high-value regions are also the sensitive regions for anomalous variations in snow depth in China; (3) in the past 41 years, the interannual variability of snow depth in China has shown a significantly decreasing trend, and the linear tendency of snow depth is - 0.093 cm/10 a (p < 0.01) and the snow depth in four seasons showed a decreasing trend (p < 0.05); and (4) the driving factors of snow heterogeneity are dissimilar in different regions and seasons. In temperate zones, average air temperature is the main factor affecting snow depth in cold temperature, mid temperature, and warm temperature zones; the maximum air temperature is the main factor affecting snow depth in mid temperate and warm temperate zones. Both the minimum air temperature and the average land-surface temperature are important factors affecting the snow depth in the cold temperate, mid temperate and warm temperate zones, and all passed the significance test of 0.01.

Polarity-regulated derivatization-assisted LC-MS method for amino-containing metabolites profiling in gastric cancer.

Amino-containing compounds, including amino acids, aliphatic amines, aromatic amines, small peptides and catecholamines, are involved in various biological processes and play vital roles in multiple metabolic pathways. Previous studies indicated that some amino-containing metabolites are significant diagnostic and prognostic biomarkers of gastric cancer. However, the discovery of precise biomarkers for the preoperative diagnosis of gastric cancer is still in an urgent need. Herein, we established a polarity-regulated derivatization method coupled with liquid chromatography-mass spectrometry (LC-MS) for amino-containing metabolites profiling in the serum samples of patients with gastric cancer and healthy controls, based on our newly designed and synthesized derivatization reagent (S)-3-(1-(diisopropoxyphosphoryl) pyrrolidine-2-carboxamido)-N-hydroxysuccinimidyl ester (3-DP-NHS). Enhanced separation efficiency and detection sensitivity for amino-containing metabolites were achieved after derivatization. This method exhibited good linearity, recovery, intra- and inter-day precision and accuracy. Only 5 µL serum is needed for untargeted analysis, enabling 202 amino-containing metabolites to be detected. Statistical analysis revealed altered amino acid metabolisms in patients with gastric cancer. Furthermore, ultra high performance liquid chromatography coupled with mass spectrometry (UHPLC-MS/MS) analysis quantification revealed increased serum levels of tryptamine and decreased concentrations of arginine and tryptophan in patients with gastric cancer. Receiver operating characteristic (ROC) curves indicated that an increased tryptamine/tryptophan ratio could serve as a potential biomarker for gastric cancer diagnosis. This study demostrated the possibility of using serum amino acid biomarkers for gastric cancer diagnosis, providing new avenues for the treatment of gastric cancer.

Polarity-extended composition profiling via LC-MS-based metabolomics approaches: A key to functional investigation of Citrus aurantium L.

Citrus is a genus containing diverse edible species, among them Citrus aurantium L. is widely utilized while short of composition research. Herein, utilizing multiple liquid chromatography-mass spectrometry (LC-MS)-based metabolomics approaches, we comprehensively characterized its components. We first systematized both LC and MS characteristics of polymethoxyflavones (PMFs), by which 13 PMFs were identified in C. aurantium, and their biosynthesis pathway was further established. Using derivatization-LC-MS targeted metabolomics approaches, 28 carbohydrates and 18 carboxylic acids were firstly found in C. aurantium. Combined with untargeted metabolomics method, total 147 compositions were characterized, among which 92 were firstly reported in C. aurantium. We further obtained their geographical features and sought out principal discriminative compounds. Moreover, typical biofunctions of C. aurantium from diverse regions were speculated using pharmacological platform and partly verified by experiments. The present study provided systematic component information for C. aurantium, which laid the foundation for its further exploitation as functional food.

Identification of polyunsaturated fatty acids as potential biomarkers of osteoarthritis after sodium hyaluronate and mesenchymal stem cell treatment through metabolomics.

Introduction: Osteoarthritis (OA) is a prevalent joint disorder worldwide. Sodium hyaluronate (SH) and mesenchymal stem cells (MSCs) are promising therapeutic strategies for OA. Previous studies showed they could improve knee function and clinical symptoms of OA. However, the mechanism of the therapeutic effects on the improvement of OA has not been clearly explained. Methods: In our study, we used a technique called 5-(diisopropylamino)amylamine derivatization liquid chromatography coupled with mass spectrometry to find the metabolites in OA synovial fluid under different treatments. Results and Discussion: After looking into the metabolomics, we discovered that SH and MSC treatment led to the downregulation of ω-6 polyunsaturated fatty acids (PUFAs) and the upregulation of ω-3 PUFAs. Significantly, the contents of 5(S)-HETE, PGA2, PGB2, and PGJ2 were lower in the MSC group than in the SH group after quantification using 5-(diisopropylamino)amylamine derivatization-UHPLC-QQQ-MS. This is the first report on the relationship of 11(S)-HETE, PGA2, PGB2, PGF2ß, 11ß-PGF2α, and DK-PGE2 with OA. Moreover, the correlation analysis of metabolites and inflammation factors showed the positive association of ω-6 PUFAs with pro-inflammation cytokines, and of ω-3 PUFAs with anti-inflammation cytokines. Our results indicated the therapeutic effect of SH and MSCs in patients with OA. In addition, this reliable metabolic approach could uncover novel biomarkers to treat OA.

Discovery of novel ascorbic acid derivatives and other metabolites in fruit of Rosa roxburghii Tratt through untargeted metabolomics and feature-based molecular networking.

Fruit of Rosa roxburghii Tratt (FRR) is widely used in functional food industry while short of metabolites research. Herein, we firstly identified 251 metabolites in FRR based on untargeted liquid chromatography-mass spectrometry (LC-MS) approach. Then, 42 differential compounds were sought out to avoid the confusing use of FRR and fruit of R. sterilis S. D. Shi (FRS), and FRR was evaluated exhibiting higher biofunction potential. Moreover, a quantitative LC-MS approach was established to determine the contents of 3 ascorbyl hexosides, and FRR with higher contents should be better source than FRS. Additionally, 17 ascorbic acid (AA) derivatives formed by conjugation of ascorbyl unit with organic acids, flavonoids, or glucuronic acid were also discovered in FRR through characteristic ions of AA and feature-based molecular networking (FBMN), enlightening that AA derivatives were not limited to ascorbyl glycosides. This study provided abundant metabolites information of FRR, laying the basis for exploitation of FRR.