Integrative Molecular Phenotyping
INTEGRATIVE MOLECULAR
PHENOTYPING
WHEELOCK LABORATORY
DEPARTMENT OF MEDICAL
BIOCHEMISTRY AND BIOPHYSICS
WHEELOCK LABORATORY
DEPARTMENT OF MEDICAL
BIOCHEMISTRY AND BIOPHYSICS
WHEELOCK LABORATORY
DEPARTMENT OF MEDICAL
BIOCHEMISTRY AND BIOPHYSICS
WHEELOCK LABORATORY
DEPARTMENT OF MEDICAL
BIOCHEMISTRY AND BIOPHYSICS
WHEELOCK LABORATORY
DEPARTMENT OF MEDICAL
BIOCHEMISTRY AND BIOPHYSICS
WHEELOCK LABORATORY

PubMed

Gypenosides suppress hepatocellular carcinoma cells by blocking cholesterol biosynthesis through inhibition of MVA pathway enzyme HMGCS1

Mon, 21/08/2023 - 12:00
Chem Biol Interact. 2023 Aug 19:110674. doi: 10.1016/j.cbi.2023.110674. Online ahead of print.ABSTRACTHepatocellular carcinoma (HCC) is one of the most common malignant tumors with high morbidity and mortality. Targeting abnormal cholesterol metabolism is a potential therapeutic direction. Therefore, more natural drugs targeting cholesterol in HCC need to be developed. Gypenosides (Gyp), the major constituent of Gynostemma pentaphyllum, has been demonstrated to have pharmacological properties on anti-cancer, anti-obesity, and hepatoprotective. We investigated whether Gyp, isolated and purified by our lab, could inhibit HCC progression by inhibiting cholesterol synthesis. The present research showed that Gyp inhibited proliferation and migration, and induced apoptosis in Huh-7 and Hep3B cells. Metabolomics, transcriptomics, and target prediction all suggested that lipid metabolism and cholesterol biosynthesis were the mechanisms of Gyp. Gyp could limit the production of cholesterol and target HMGCS1, the cholesterol synthesis-related protein. Downregulation of HMGCS1 could suppress the progression and abnormal cholesterol metabolism of HCC. In terms of mechanism, Gyp suppressed mevalonate (MVA) pathway mediated cholesterol synthesis by inhibiting HMGCS1 transcription factor SREBP2. And the high expression of HMGCS1 in HCC human specimens was correlated with poor clinical prognosis. The data suggested that Gyp could be a promising cholesterol-lowering drug for the prevention and treatment of HCC. And targeting SREBP2-HMGCS1 axis in MVA pathway might be an effective HCC therapeutic strategy.PMID:37604220 | DOI:10.1016/j.cbi.2023.110674

Comprehensive transcriptomic and metabolomic analysis of the effect of feed restriction on duck sternal development

Mon, 21/08/2023 - 12:00
Poult Sci. 2023 Jul 26;102(10):102961. doi: 10.1016/j.psj.2023.102961. Online ahead of print.ABSTRACTSkeletal characteristics are important to the growth and development of poultry. In feeding management, constant free feeding (FF) of poultry may lead to imbalance between bone development and weight gain. Feed restriction (FR), to a certain extent, is one way to solve this problem. However, the effect of feed restriction on poultry bone development needs further elucidation at the molecular level. Therefore, in the present study, we investigated the effects of different levels of feed restriction (60% FR, 70% FR, 80% FR, and FF) on the sternum development of ducks at 7 and 8 wk old. In the seventh wk, with increasing feed restriction, the values of traits including body weight, breast muscle weight, sternal weight, keel length, and calcified keel length decreased. However, in the eighth wk, the sternum weight and keel length of ducks treated with 60% FR were unexpectedly higher than those of FF individuals, indicative of catch-up growth. Then, we conducted RNA-seq and metabolomic analysis on sterna from 7- and 8-wk-old FF and 60% FR ducks. The results identified multiple differentially expressed genes (DEGs) associated with sternum development that were influenced by feed restriction. Among them, we found that the mRNA expression levels of the chondroitin sulfate synthase 3 (CHSY3) and annexin A2 (ANXA2) which are involved in glycosaminoglycan biosynthesis and bone mineralization, had smaller changes over time under FR treatment than under FF treatment, implying that the FR treatment to a certain extent prevented the premature calcification and prolonged the development time of duck sternum. In addition, the metabolomic and integrative analyses revealed that several antiaging-related metabolites and genes were associated with sternal catch-up growth. Pyrimidine metabolism was identified as the most significant pathway in which most differential metabolites (DMs) between FF and 60% FR were enriched. The results from integrative analysis revealed that the content and expression of 4-aminobutyric acid (GABA) and its related genes showed relatively higher activity in the 60% FR group than in the FF group. The present study identifies multiple biomarkers associated with duck sternum development that are influenced by feed restriction and suggests the potential mechanism of feed restriction-associated duck sternal catch-up growth.PMID:37604023 | DOI:10.1016/j.psj.2023.102961

Changes in flavor and biological activities of Lentinula edodes hydrolysates after Maillard reaction

Mon, 21/08/2023 - 12:00
Food Chem. 2023 Aug 17;431:137138. doi: 10.1016/j.foodchem.2023.137138. Online ahead of print.ABSTRACTThis study aimed to elucidate how the Maillard reaction (MR) affects the flavor and bioactivities of Lentinula edodes hydrolysates (LEHs). Changes in flavor were investigated using non-targeted metabolomics techniques (GC-MS and LC-MS/MS) and sensory evaluation. Simultaneously, UV absorption, fluorescence, and FT-IR spectra were used to characterize the process of MR. We also evaluated the effects of MR on the antioxidant activity, hypoglycemic activity and antimicrobial activity of LEHs in vitro. The results revealed that MR produced many volatile aldehydes and ketones and decreased the content of most amino acids, sugars and flavonoids in the LEHs while increasing the content of l-theanine and succinic acid. MRPs had a strong caramel and like-meat flavor and an obvious improvement in umami, taste continuity, and total acceptability. Furthermore, MR improved the antioxidant and antimicrobial properties of LEHs. This research establishes a theoretical foundation for MR in the deep processing of edible mushrooms.PMID:37604001 | DOI:10.1016/j.foodchem.2023.137138

Phytochemical and functional characterization of cultivated varieties of Morus alba L. fruits grown in Italy

Mon, 21/08/2023 - 12:00
Food Chem. 2023 Aug 7;431:137113. doi: 10.1016/j.foodchem.2023.137113. Online ahead of print.ABSTRACTMorus alba L. fruits are considered functional foods with an important nutritional value for their high content of polyphenols. Therefore, the type and level of phytochemicals of the soroses from 13 M. alba cultivars grown in Italy were characterized due to the lack of data available about their nutraceutical properties. Mature M. alba fruits exhibited variable polyphenol, flavonoid, anthocyanin, proanthocyanins, and 1-deoxynojirimycin contents which resulted in different antioxidant and α-glucosidase inhibitory activities. Regression models built on UHPLC-HRMS results revealed a strong correlation between the expression of quercetin derivatives, cyanidin 3-O-glucoside, caffeoyl methyl quinates, and 5,5'-dehydrodivanillic acid, and the biological activity of each fruit. On another note, principal component analysis revealed that the quantity of caffeoyl/dicaffeoyl methyl quinate, caffeoylquinic acids, and quercetin derivatives decreased during ripening. The results on the compositional and functional characterization of mature M. alba fruits might improve their consumption and economic value in Italy.PMID:37604000 | DOI:10.1016/j.foodchem.2023.137113

Sugar composition and transcriptome analysis in developing 'Fengtang' plum (Prunus salicina Lindl.) reveal candidate genes regulating sugar accumulation

Mon, 21/08/2023 - 12:00
Plant Physiol Biochem. 2023 Aug 11;202:107955. doi: 10.1016/j.plaphy.2023.107955. Online ahead of print.ABSTRACTSweetness is an important attribute of fruit quality, which directly affects consumers' preference for fresh fruit and is mostly determined by carbohydrate composition. 'Fengtang' plum (Prunus salicina Lindl.) is recognized for its high soluble sugar content, but the sugar composition and the molecular mechanisms underlying sugar overproduction are not fully understood. In this work, the sugar components were analyzed using gas chromatography-mass spectrometry combined with transcription profiles from RNA-sequencing and Quantitative Real-time PCR during fruit development. The target metabolic group showed that sucrose was the dominant sugar component in mature fruit, followed by glucose, fructose, and sorbitol. Based on the transcriptome data and qRT-PCR validation, we identified 12 key structural genes that significantly responded to corresponding component accumulation: sucrose synthase (PsSUS4), sucrose phosphate synthase (PsSPS2), neutral invertase (PsNINV1/3/4), phosphoglucomutase (PsPGM1), UTP-glucose-1-phosphate uridylyl transferase (PsUGP1/2), hexose kinase (PsHXK1/3), sugar transport protein (PsSTP1), and Sugars Will Eventually be Exported Transporter (PsSWEET4). In which PsSUS4 and PsSPS2, whose encoding proteins immediately catalyze sucrose synthesis, were selected to be silenced using the virus-induced gene silencing technology. Silencing of PsSUS4 or PsSPS2 resulted in decreased sucrose content by 27.6% and 8%, respectively, compared with the control, verifying their important roles in sucrose accumulation. Subsequently, sugar metabolism networks in this high-sugar plum were constructed with 12 key structural genes, 72 putative transcription factors, and 4 major sugar components. These results might facilitate a better understanding of the molecular mechanisms of sugar accumulation in 'Fengtang' plum and provide a framework for future fruit quality improvement.PMID:37603969 | DOI:10.1016/j.plaphy.2023.107955

Application of metabolomics in irritable bowel syndrome in recent 5 years

Mon, 21/08/2023 - 12:00
Int Immunopharmacol. 2023 Aug 19;124(Pt A):110776. doi: 10.1016/j.intimp.2023.110776. Online ahead of print.ABSTRACTIrritable bowel syndrome (IBS) is one of the most common functional gastrointestinal disorders worldwide, characterized by chronic abdominal pain or discomfort and altered bowel habits. To date, the exact pathogenesis of IBS remains elusive, but is clearly multifactorial, including environmental and host factors. However, the management of patients with IBS is challenging and the current diagnostic and therapeutic modalities have unsatisfactory outcomes. Therefore, it is important to develop more effective methods to diagnose IBS early. Metabolomics studies the metabolites most closely related to patient characteristics, which can provide useful clinical biomarkers that can be applied to IBS and may open up new diagnostic approaches. Traditional Chinese medicine (TCM) can play a role in improving symptoms and protecting target organs, but its mechanism needs to be studied in depth. In this review, based on PubMed/MEDLINE and other databases, we searched metabolomics studies related to IBS in the past 5 years, including those related to clinical studies and animal studies, as well as literatures on TCM interventions in IBS, to provide an updated overview of the application of metabolomics to the diagnosis and treatment of IBS and the improvement of IBS by TCM.PMID:37603947 | DOI:10.1016/j.intimp.2023.110776

Identification of canine mammary tumor-associated metabolites using untargeted metabolomics

Mon, 21/08/2023 - 12:00
Theriogenology. 2023 Aug 16;211:84-96. doi: 10.1016/j.theriogenology.2023.08.010. Online ahead of print.ABSTRACTThe canine mammary tumor is the most common tumor type in female dogs and seriously threatens their life. Currently, no effective treatments are available for this condition. Hence, it is essential to identify biomarkers that positively influence the early diagnosis and treatment and prognosis of this disease. To provide a basis for early diagnosis of canine breast tumors, in this study, 23 dogs with mammary tumors were identified via histopathological examination combined with ancillary diagnoses via blood examinations and diagnostic imaging. The canine mammary tumor and tumor-adjacent healthy tissues were collected, and their metabolites were identified utilizing a UHPLC-qTOF-MS-based untargeted metabolomics approach. The metabolic results revealed a total of 979 ion features in the positive polarity mode and 371 ion features in the negative polarity mode in the tissues of two groups; among them, 536 differential metabolites (385 in the positive and 151 in the negative polarity mode) were analyzed by PCA and PLS-DA. Subsequently, the enrichment pathways purine metabolism, alpha-linolenic acid metabolism, vitamin B6 metabolism, and fatty acid biosynthesis were analyzed using Metaboanalyst 4.0, which suggested that these pathways were valuable diagnostic and therapeutic targets. Moreover, the receiver operating characteristic curves further confirmed 13Z,16Z-docosadienoic acid, 23-nordeoxycholic acid, and (±)12(13)-DiHOME as expected candidate biomarkers of canine mammary tumors. In conclusion, the discovery of tumor biomarkers based on untargeted metabolomics is informative for pathological mechanism studies and facilitates the early diagnosis of canine mammary tumors.PMID:37603937 | DOI:10.1016/j.theriogenology.2023.08.010

Principles of metabolome conservation in animals

Mon, 21/08/2023 - 12:00
Proc Natl Acad Sci U S A. 2023 Aug 29;120(35):e2302147120. doi: 10.1073/pnas.2302147120. Epub 2023 Aug 21.ABSTRACTMetabolite levels shape cellular physiology and disease susceptibility, yet the general principles governing metabolome evolution are largely unknown. Here, we introduce a measure of conservation of individual metabolite levels among related species. By analyzing multispecies tissue metabolome datasets in phylogenetically diverse mammals and fruit flies, we show that conservation varies extensively across metabolites. Three major functional properties, metabolite abundance, essentiality, and association with human diseases predict conservation, highlighting a striking parallel between the evolutionary forces driving metabolome and protein sequence conservation. Metabolic network simulations recapitulated these general patterns and revealed that abundant metabolites are highly conserved due to their strong coupling to key metabolic fluxes in the network. Finally, we show that biomarkers of metabolic diseases can be distinguished from other metabolites simply based on evolutionary conservation, without requiring any prior clinical knowledge. Overall, this study uncovers simple rules that govern metabolic evolution in animals and implies that most tissue metabolome differences between species are permitted, rather than favored by natural selection. More broadly, our work paves the way toward using evolutionary information to identify biomarkers, as well as to detect pathogenic metabolome alterations in individual patients.PMID:37603743 | DOI:10.1073/pnas.2302147120

<em>De Novo</em> Cleaning of Chimeric MS/MS Spectra for LC-MS/MS-Based Metabolomics

Mon, 21/08/2023 - 12:00
Anal Chem. 2023 Aug 21. doi: 10.1021/acs.analchem.3c00736. Online ahead of print.ABSTRACTThe purity of tandem mass spectrometry (MS/MS) is essential to MS/MS-based metabolite annotation and unknown exploration. This work presents a de novo approach to cleaning chimeric MS/MS spectra generated in liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomics. The assumption is that true fragments and their precursors are well correlated across the samples in a study, while false or contamination fragments are rather independent. Using data simulation, this work starts with an investigation of the negative effects of chimeric MS/MS spectra on spectral similarity analysis and molecular networking. Next, the characteristics of true and false fragments in chimeric MS/MS spectra were investigated using MS/MS of chemical standards. We recognized three fragment peak attributes indicative of whether a peak is a false fragment, including (1) intensity ratio fluctuation, (2) appearance rate, and (3) relative intensity. Using these attributes, we tested three machine learning models and identified XGBoost as the best model to achieve an area under the precision-recall curve of 0.98 for a clear separation between true and false fragments. Based on the trained model, we constructed an automated bioinformatic platform, DNMS2Purifier (short for de novo MS2Purifier), for metabolic features from metabolomics studies. DNMS2Purifier recognizes and processes chimeric MS/MS spectra without additional sample analysis or library confirmation. DNMS2Purifer was evaluated on a metabolomics data set generated with different MS/MS precursor isolation windows. It successfully captured the increase in the number of false fragments from the increased isolation window. DNMS2Purifier was also compared to MS2Purifier, an existing MS/MS spectral cleaning tool based on the addition of data-independent acquisition (DIA) analysis. Results indicated that DNMS2Purifier uniquely recognizes false fragments, which complements the previous DIA-based approach. Finally, DNMS2Purifier was demonstrated using a real experimental metabolomics study, showing improved MS/MS spectral quality and leading to an improved spectral match ratio and molecular networking outcome.PMID:37603462 | DOI:10.1021/acs.analchem.3c00736

Metformin, cognitive function, and changes in the gut microbiome

Mon, 21/08/2023 - 12:00
Endocr Rev. 2023 Aug 21:bnad029. doi: 10.1210/endrev/bnad029. Online ahead of print.ABSTRACTThe decline in cognitive function and the prevalence of neurodegenerative disorders are among the most serious threats to health in old age. The prevalence of dementia has reached 50 million people worldwide and has become one of the major public health problems. The causes of age-related cognitive impairment are multiple, complex, and difficult to determine. However, type 2 diabetes (T2D) is linked to an enhanced risk of cognitive impairment and dementia. Human studies have shown that patients with T2D exhibit dysbiosis of the gut microbiota. This dysbiosis may contribute to the development of insulin resistance and increased plasma lipopolysaccharide concentrations. Metformin medication mimics some of the benefits of calorie restriction and physical activity, such as greater insulin sensitivity and decreased cholesterol levels, and hence may also have a positive impact on aging in humans. According to recent human investigations, metformin might partially restore gut dysbiosis related to T2D. Likewise, some studies showed that metformin reduced the risk of dementia and improved cognition, although not all studies are concordant. Therefore, this review focused on those human studies describing the effects of metformin on the gut microbiome (specifically the changes in taxonomy, function, and circulating metabolomics), the changes in cognitive function and their possible bidirectional implications.PMID:37603460 | DOI:10.1210/endrev/bnad029

Use of N-(4-aminophenyl)piperidine derivatization to improve organic acid detection with supercritical fluid chromatography-mass spectrometry

Mon, 21/08/2023 - 12:00
J Sep Sci. 2023 Aug 21:e2300343. doi: 10.1002/jssc.202300343. Online ahead of print.ABSTRACTThe analysis of organic acids in complex mixtures by LC-MS can often prove challenging, especially due to the poor sensitivity of negative ionization mode required for detection of these compounds in their native (i.e., underivatized or untagged) form. These compounds have also been difficult to measure using supercritical fluid chromatography (SFC)-MS, a technique of growing importance for metabolomic analysis, with similar limitations based on negative ionization. In this report, the use of a high proton affinity N-(4-aminophenyl)piperidine derivatization tag is explored for the improvement of organic acid detection by SFC-MS. Four organic acids (lactic, succinic, malic, and citric acids) with varying numbers of carboxylate groups were derivatized with N-(4-aminophenyl)piperidine to achieve detection limits down to 0.5 ppb, with overall improvements in detection limit ranging from 25-to-2100-fold. The effect of the derivatization group on sensitivity, which increased by at least 200-fold for compounds that were detectable in their native form, and mass spectrometric detection are also described. Preliminary investigations into the separation of these derivatized compounds identified multiple stationary phases that could be used for complete separation of all four compounds by SFC. This derivatization technique provides an improved approach for the analysis of organic acids by SFC-MS, especially for those that are undetectable in their native form.PMID:37603367 | DOI:10.1002/jssc.202300343

Molecular mechanisms of physiological change under acute total dissolved gas supersaturation stress in yellow catfish (Pelteobagrus fulvidraco)

Mon, 21/08/2023 - 12:00
Environ Sci Pollut Res Int. 2023 Aug 21. doi: 10.1007/s11356-023-29157-6. Online ahead of print.ABSTRACTDuring the dam discharging period, the strong aeration of high-speed water leads to the supersaturation of total dissolved gas (TDG) in the downstream water, which causes gas bubble disease (GBD) in fish and threatens their survival. TDG supersaturation has now become an ecological and environmental issue of global concern; however, the molecular mechanism underlying the physiological effect of TDG supersaturation on fish is poorly known. Here, we comprehensively investigated the effect of TDG supersaturation on Pelteobagrus fulvidraco at the histopathological, biochemical, transcriptomic, and metabolomic levels. After exposure to 116% TDG, P. fulvidraco exhibited classic GBD symptoms and pathological changes in gills. The level of superoxide dismutase was highly significantly decreased. Transcriptomic results revealed that heat shock proteins (HSPs) and a large number of genes involved in immunity were increased by TDG stress. A key environmental sensor PI3K/Akt/mTOR pathway was significantly stimulated for defence against stress. Integrated transcriptomic and metabolomic analyses revealed that key metabolites and genes were upregulated in the triacylglycerol synthesis pathway and that amino acid levels decreased, which might be associated with TDG supersaturation stress. The present study demonstrated that TDG supersaturation could cause severe physiological damage in fish. HSP genes, immune functions, and energy metabolic pathways were enhanced to counteract the adverse effects.PMID:37603244 | DOI:10.1007/s11356-023-29157-6

Distinctive metabolic remodeling in TYMP deficiency beyond mitochondrial dysfunction

Mon, 21/08/2023 - 12:00
J Mol Med (Berl). 2023 Aug 21. doi: 10.1007/s00109-023-02358-9. Online ahead of print.ABSTRACTMitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is caused by mutations in the TYMP gene, which encodes thymidine phosphorylase (TP). As a cytosolic metabolic enzyme, TP defects affect biological processes that are thought to not be limited to the abnormal replication of mitochondrial DNA. This study aimed to elucidate the characteristic metabolic alterations and associated homeostatic regulation caused by TYMP deficiency. The pathogenicity of novel TYMP variants was evaluated in terms of clinical features, genetic analysis, and structural instability. We analyzed plasma samples from three patients with MNGIE; three patients with m.3243A > G mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS); and four healthy controls (HC) using both targeted and untargeted metabolomics techniques. Transcriptomics analysis and bioenergetic studies were performed on skin fibroblasts from participants in these three groups. A TYMP overexpression experiment was conducted to rescue the observed changes. Compared with controls, specific alterations in nucleosides, bile acids, and steroid metabolites were identified in the plasma of MNGIE patients. Comparable mitochondrial dysfunction was present in fibroblasts from patients with TYMP deficiency and in those from patients with the m.3243A > G mutation. Distinctively decreased sterol regulatory element binding protein (SREBP) regulated cholesterol metabolism and fatty acid (FA) biosynthesis as well as reduced FA degradation were revealed in fibroblasts with TYMP deficiency. The restoration of thymidine phosphorylase activity rescued the observed changes in MNGIE fibroblasts. Our findings indicated that more widespread metabolic disturbance may be caused by TYMP deficiency in addition to mitochondrial dysfunction, which expands our knowledge of the biochemical outcome of TYMP deficiency. KEY MESSAGES: Distinct metabolic profiles in patients with TYMP deficiency compared to those with m.3243A > G mutation. TYMP deficiency leads to a global disruption of nucleoside metabolism. Cholesterol and fatty acid metabolism are inhibited in individuals with MNGIE. TYMP is functionally related to SREBP-regulated pathways. Potential metabolite biomarkers that could be valuable clinical tools to improve the diagnosis of MNGIE.PMID:37603049 | DOI:10.1007/s00109-023-02358-9

Hierarchical Contribution Of Argonaute Proteins To Antiviral Protection

Mon, 21/08/2023 - 12:00
J Exp Bot. 2023 Aug 21:erad327. doi: 10.1093/jxb/erad327. Online ahead of print.ABSTRACTAntiviral RNA interference (RNAi) is the main protective measure employed by plants in the fight against viruses. The main steps of this process have been clarified in recent years, primarily relying on the extensive genetic resources of Arabidopsis thaliana. Our knowledge of viral diseases of crops however is still limited, mainly due to the fact that A. thaliana is a non-host for many agriculturally important viruses. Contrary, Nicotiana benthamiana has an unparalleled susceptibility to viruses, and since it belongs to the Solanaceae family, it is considered an adequate system for modeling infectious diseases of crops such as tomatoes. We used a series of N. benthamiana mutants created by genome editing to analyze the RNAi response elicited by the emerging tomato pathogen, Pepino mosaic virus (PepMV). We uncovered hierarchical roles of several Argonaute proteins (AGOs) in anti-PepMV defense, with AGO2's predominant contribution. Interestingly, the anti-PepMV activities of AGO1A, AGO5 and AGO10 only become apparent when AGO2 is mutated. Taken together, our results prove that hierarchical actions of several AGOs are needed for the plant to build effective anti-PepMV resistance. The genetic resources created here will be valuable assets for analyzing RNAi responses triggered by other agriculturally important pathogenic viruses.PMID:37603044 | DOI:10.1093/jxb/erad327

Tips And Tricks for Proteome Sample Preparation

Mon, 21/08/2023 - 12:00
J Vis Exp. 2023 Feb 3;(192). doi: 10.3791/64830.ABSTRACTKovalchuk, S. I., Ziganshin, R., Shelukhina, I. Simple in-house ultra-high performance capillary column manufacturing with the FlashPack approach. Journal of Visualized Experiments. (178), e62522 (2021). Sirois, I., Isabelle, M., Duquette, J. D., Saab, F., Caron, E. Immunopeptidomics: Isolation of mouse and human MHC Class I- and II-associated peptides for mass spectrometry analysis. Journal of Visualized Experiments. (176), e63052 (2021). Han, Y., Thomas, C. T., Wennersten, S. A., Lau, E., Lam, M. P. Y. Shotgun proteomics sample processing automated by an open-source lab robot. Journal of Visualized Experiments. (176), e63092 (2021). Nickerson, J. L. et al. Organic solvent-based protein precipitation for robust proteome purification ahead of mass spectrometry. Journal of Visualized Experiments. (180), e63503 (2022). Li, D., Liang, J., Zhang, Y., Zhang, G. An integrated workflow of identification and quantification on FDR control-based untargeted metabolome. Journal of Visualized Experiments. doi: 10.3791/63625-v (2022). Petelski, A. A., Nikolai Slavov, N., Specht, N. Single-cell proteomics preparation for mass spectrometry analysis using freeze-heat lysis and an isobaric carrier. Journal of Visualized Experiments. doi: 10.3791/63802 (2022).PMID:37602878 | DOI:10.3791/64830

Serum metabolomics identifies uric acid as a possible novel biomarker for predicting recurrence of chronic rhinosinusitis with nasal polyps

Mon, 21/08/2023 - 12:00
Rhinology. 2023 Aug 21. doi: 10.4193/Rhin23.236. Online ahead of print.ABSTRACTBACKGROUND: Metabolomics has proven to be a valuable tool in gaining new insights into disease progression and prognosis, the specific metabolic alterations in the serum of recurrent chronic rhinosinusitis with nasal polyps (CRSwNP) patients remain unknown. This study aims to explore the serum metabolomic profiles of recurrent CRSwNP and identify potential predictive biomarkers.METHODS: A prospective, single-center study was conducted on CRSwNP patients prior to endoscopic sinus surgery. Serum samples were subjected to untargeted metabolomic profiling. Patients were followed up for over 2 years and categorized into recurrence and non-recurrence groups. Metabolite differences between the two groups were compared, and the identified differentially regulated metabolites were subsequently validated in a large clinical cohort.RESULTS: 67 CRSwNP patients completed the follow-up schedule, with 47 classified into the non-recurrent group and 20 into the recurrent group. Significant differences were found in the metabolomic profiles between both groups, and serum uric acid (SUA) showed promising predictive potential for postoperative recurrence in both positive and negative ion models. A validation cohort comprising 398 non-recurrent and 142 recurrent CRSwNP patients was recruited, and a significant elevation in SUA levels was observed in recurrent cases. Patients were stratified into tertiles based on the distribution of baseline SUA levels. Multivariate Cox regression analysis showed that higher tertiles of SUA were associated with an increased risk of CRSwNP recurrence compared to lower tertiles, even after adjusting for potential confounding factors. The receiver operating characteristic curve and Kaplan-Meier survival analysis highlighted that elevated SUA levels exhibited potential predictive values for postoperative recurrence.CONCLUSION: Serum metabolic signatures might predict postoperative recurrence in CRSwNP patients. Increased SUA concentrations were found to be associated with a higher risk of future postoperative recurrence in CRSwNP, independent of traditional risk factors.PMID:37602858 | DOI:10.4193/Rhin23.236

Multiomics reveals the mechanism of <em>B. longum</em> in promoting the formation of mixed-species biofilms

Mon, 21/08/2023 - 12:00
Food Funct. 2023 Aug 21. doi: 10.1039/d3fo01751f. Online ahead of print.ABSTRACTIt has been found previously that Bifidobacterium longum, Bacteroides ovatus, Enterococcus faecalis, and Lactobacillus gasseri can form a biofilm better when co-cultured in vitro and B. longum is the core biofilm-formation-promoting strain in this community. B. longum is part of the core microbiota in the gut and is widely recognized as a probiotic. Therefore, it is necessary to explore its role in mixed-species biofilms through transcriptomics and metabolomics. Metabolomics showed that the increase in amino acid and purine content could promote biofilm formation. In transcriptomic analysis, many genes related to carbohydrate metabolism, amino acid metabolism, and environmental tolerance of B. longum were up-regulated. Combined with the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and Gene Ontology (GO) analysis, the differentially expressed genes (DEGs) of B. longum in mixed-species biofilms were mainly correlated to "quorum sensing (QS)", "ABC transporters", "biosynthesis of amino acids", "microbial metabolism in different environments", "carbohydrate metabolism" and "two-component system". In addition, the rpl and rps gene families, which function in the metabolism of organic substances and the biosynthesis of amino acids, were the core DEGs according to the analysis of the protein-protein interaction (PPI) network. Finally, by combining metabolomics and quorum sensing mechanisms, it was found that the metabolism of autoinducer peptides (proliylglycine and glycylleucine), N-acyl homoserine lactone (N-(3-oxo hydroxy) homoserine lactone), and AI-2 can promote the formation of biofilms, both mono- and mixed-species biofilms composed of B. longum. Our research enabled us to understand the critical role of B. longum in mixed-species biofilms and the interactions between biofilm metabolism and gut health. In addition, the generated knowledge will be of great significance for us to develop biofilm products with beneficial functions in future.PMID:37602484 | DOI:10.1039/d3fo01751f

Mechanistic exploration of Yiqi Liangxue Shengji prescription on restenosis after balloon injury by integrating metabolomics with network pharmacology

Mon, 21/08/2023 - 12:00
Pharm Biol. 2023 Dec;61(1):1260-1273. doi: 10.1080/13880209.2023.2244533.ABSTRACTCONTEXT: Yiqi Liangxue Shengji prescription (YQLXSJ) is a traditional Chinese medicine (TCM) formula that has long been used for treatment after percutaneous coronary intervention (PCI).OBJECTIVE: To investigate the putative pharmacological mechanism of YQLXSJ on restenosis through an integrated approach utilizing metabolomics and network pharmacology.MATERIALS AND METHODS: Forty male Sprague-Dawley rats were divided into sham, model, YQLXSJ, and positive groups. YQLXSJ group received the treatment of YQLXSJ (6 g/kg/d, i.g.) and the positive group was treated with atorvastatin (2 mg/kg/d, i.g.). After 4 weeks, the improvement in intimal hyperplasia was evaluated by ultrasound, H&E staining, and immunofluorescence. UPLC-MS/MS technology was utilized to screen the differential metabolites. Network pharmacology was conducted using TCMSP, GeneCards, and Metascape, etc., in combination with metabolomics. Eventually, the core targets were acquired and validated.RESULTS: Compared to models, YQLXSJ exhibited decreased intima-media thickness on ultrasound (0.23 ± 0.02 mm vs. 0.20 ± 0.01 mm, p < 0.01) and reduced intima thickness by H&E (30.12 ± 6.05 μm vs. 14.32 ± 1.37 μm, p < 0.01). We identified 18 differential metabolites and 5 core targets such as inducible nitric oxide synthase (NOS2), endothelial nitric oxide synthase (NOS3), vascular endothelial growth factor-A (VEGFA), ornithine decarboxylase-1 (ODC1) and group IIA secretory phospholipase A2 (PLA2G2A). These targets were further confirmed by molecular docking and ELISA.DISCUSSION AND CONCLUSIONS: This study confirms the effects of YQLXSJ on restenosis and reveals some biomarkers. TCM has great potential in the prevention and treatment of restenosis by improving metabolic disorders.PMID:37602438 | DOI:10.1080/13880209.2023.2244533

Metabolomic characterization benefits the identification of acute lung injury in patients with type A acute aortic dissection

Mon, 21/08/2023 - 12:00
Front Mol Biosci. 2023 Aug 3;10:1222133. doi: 10.3389/fmolb.2023.1222133. eCollection 2023.ABSTRACTIntroduction: Acute aortic dissection (AAD) often leads to the development of acute lung injury (ALI). However, the early detection and diagnosis of AAD in patients with ALI pose significant challenges. The objective of this study is to investigate distinct metabolic alterations in the plasma samples of AAD patients with ALI, AAD patients without ALI, and healthy individuals. Method: Between September 2019 and September 2022, we retrospectively collected data from 228 AAD patients who were diagnosed with ALI through post-surgery chest X-ray and PaO2/FiO2 assessments. Univariate analysis was employed to identify pre-surgery risk factors for ALI. Additionally, we conducted high-throughput target metabolic analysis on 90 plasma samples, comprising 30 samples from AAD patients with ALI, 30 from patients with AAD only, and 30 from healthy controls. After LC-MS spectral processing and metabolite quantification, the recursive feature elimination with cross-validation (RFECV) analysis based on the random forest was used to select the optimal metabolites as a diagnostic panel for the detection of AAD patients with ALI. The support vector machines (SVM) machine learning model was further applied to validate the diagnostic accuracy of the established biomarker panel. Results: In the univariate analysis, preoperative β-HB and TNF-α exhibited a significant association with lung injury (OR = 0.906, 95% CI 0.852-0.965, p = 0.002; OR = 1.007, 95% CI 1.003-1.011, p < 0.0001). The multiple-reaction monitoring analysis of 417 common metabolites identified significant changes in 145 metabolites (fold change >1.2 or <0.833, p < 0.05) across the three groups. Multivariate statistical analysis revealed notable differences between AAD patients and healthy controls. When compared with the non-ALI group, AAD patients with ALI displayed remarkable upregulation in 19 metabolites and downregulation in 4 metabolites. Particularly, combining citric acid and glucuronic acid as a biomarker panel improved the classification performance for distinguishing between the ALI and non-ALI groups. Discussion: Differentially expressed metabolites in the ALI group were primarily involved in amino acids biosynthesis, carbohydrate metabolism (TCA cycle), arginine and proline metabolism, and glucagon signaling pathway. These findings demonstrate a great potential of the targeted metabolomic approach for screening, routine surveillance, and diagnosis of pulmonary injury in patients with AAD.PMID:37602331 | PMC:PMC10434778 | DOI:10.3389/fmolb.2023.1222133

Right in two: capabilities of ion mobility spectrometry for untargeted metabolomics

Mon, 21/08/2023 - 12:00
Front Mol Biosci. 2023 Aug 4;10:1230282. doi: 10.3389/fmolb.2023.1230282. eCollection 2023.ABSTRACTThis mini review focuses on the opportunities provided by current and emerging separation techniques for mass spectrometry metabolomics. The purpose of separation technologies in metabolomics is primarily to reduce complexity of the heterogeneous systems studied, and to provide concentration enrichment by increasing sensitivity towards the quantification of low abundance metabolites. For this reason, a wide variety of separation systems, from column chemistries to solvent compositions and multidimensional separations, have been applied in the field. Multidimensional separations are a common method in both proteomics applications and gas chromatography mass spectrometry, allowing orthogonal separations to further reduce analytical complexity and expand peak capacity. These applications contribute to exponential increases in run times concomitant with first dimension fractionation followed by second dimension separations. Multidimensional liquid chromatography to increase peak capacity in metabolomics, when compared to the potential of running additional samples or replicates and increasing statistical confidence, mean that uptake of these methods has been minimal. In contrast, in the last 15 years there have been significant advances in the resolution and sensitivity of ion mobility spectrometry, to the point where high-resolution separation of analytes based on their collision cross section approaches chromatographic separation, with minimal loss in sensitivity. Additionally, ion mobility separations can be performed on a chromatographic timescale with little reduction in instrument duty cycle. In this review, we compare ion mobility separation to liquid chromatographic separation, highlight the history of the use of ion mobility separations in metabolomics, outline the current state-of-the-art in the field, and discuss the future outlook of the technology. "Where there is one, you're bound to divide it. Right in two", James Maynard Keenan.PMID:37602325 | PMC:PMC10436490 | DOI:10.3389/fmolb.2023.1230282

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