Showing papers by "Agilent Technologies published in 2019"
University of Bordeaux1, Wichita State University2, University of Manchester3, Mansfield University of Pennsylvania4, Florida State University5, Pacific Northwest National Laboratory6, University of Washington7, Amgen8, University of Natural Resources and Life Sciences, Vienna9, Washington State University10, Loughborough University11, University of Liège12, Florida International University13, Agilent Technologies14, Waters Corporation15, University of Bern16, University of Minnesota17, Vanderbilt University18, French Institute of Health and Medical Research19, Birkbeck, University of London20, West Virginia University21
TL;DR: A guide to ion mobility mass spectrometry experiments, which covers both linear and nonlinear methods: what is measured, how the measurements are done, and how to report the results, including the uncertainties of mobility and collision cross section values.
Abstract: Here we present a guide to ion mobility mass spectrometry experiments, which covers both linear and nonlinear methods: what is measured, how the measurements are done, and how to report the results, including the uncertainties of mobility and collision cross section values. The guide aims to clarify some possibly confusing concepts, and the reporting recommendations should help researchers, authors and reviewers to contribute comprehensive reports, so that the ion mobility data can be reused more confidently. Starting from the concept of the definition of the measurand, we emphasize that (i) mobility values (K0) depend intrinsically on ion structure, the nature of the bath gas, temperature, and E/N; (ii) ion mobility does not measure molecular surfaces directly, but collision cross section (CCS) values are derived from mobility values using a physical model; (iii) methods relying on calibration are empirical (and thus may provide method‐dependent results) only if the gas nature, temperature or E/N cannot match those of the primary method. Our analysis highlights the urgency of a community effort toward establishing primary standards and reference materials for ion mobility, and provides recommendations to do so.
284 citations
TL;DR: It is shown that CZ-48, a cell-permeant mimetic of NMN, activated SARM1 in vitro and in cellulo to cyclize NAD and produce a Ca2+ messenger, cADPR, with similar efficiency as NMN.
120 citations
TL;DR: In an all-comer cohort, tumor burden estimates and TP53 outperform any AR perturbation to infer prognosis and outperformed ARV expression and detection of genomic AR alterations.
Abstract: Purpose: To infer the prognostic value of simultaneous androgen receptor (AR) and TP53 profiling in liquid biopsies from patients with metastatic castration-resistant prostate cancer (mCRPC) starting a new line of AR signaling inhibitors (ARSi). Experimental Design: Between March 2014 and April 2017, we recruited patients with mCRPC (n = 168) prior to ARSi in a cohort study encompassing 10 European centers. Blood samples were collected for comprehensive profiling of CellSearch-enriched circulating tumor cells (CTC) and circulating tumor DNA (ctDNA). Targeted CTC RNA sequencing (RNA-seq) allowed the detection of eight AR splice variants (ARV). Low-pass whole-genome and targeted gene-body sequencing of AR and TP53 was applied to identify amplifications, loss of heterozygosity, mutations, and structural rearrangements in ctDNA. Clinical or radiologic progression-free survival (PFS) was estimated by Kaplan–Meier analysis, and independent associations were determined using multivariable Cox regression models. Results: Overall, no single AR perturbation remained associated with adverse prognosis after multivariable analysis. Instead, tumor burden estimates (CTC counts, ctDNA fraction, and visceral metastases) were significantly associated with PFS. TP53 inactivation harbored independent prognostic value [HR 1.88; 95% confidence interval (CI), 1.18–3.00; P = 0.008], and outperformed ARV expression and detection of genomic AR alterations. Using Cox coefficient analysis of clinical parameters and TP53 status, we identified three prognostic groups with differing PFS estimates (median, 14.7 vs. 7.51 vs. 2.62 months; P Conclusions: In an all-comer cohort, tumor burden estimates and TP53 outperform any AR perturbation to infer prognosis. See related commentary by Rebello et al., p. 1699
105 citations
TL;DR: The studies demonstrate that the switch from HIF‐1 to Hif‐2 constitutes a universal mechanism of cellular adaptation to hypoxic stress and that HIF1A and HIF2A mRNA stability differences contribute to HIF switch.
Abstract: During hypoxia, a cellular adaptive response activates hypoxia-inducible factors (HIFs; HIF-1 and HIF-2) that respond to low tissue-oxygen levels and induce the expression of a number of genes that promote angiogenesis, energy metabolism, and cell survival. HIF-1 and HIF-2 regulate endothelial cell (EC) adaptation by activating gene-signaling cascades that promote endothelial migration, growth, and differentiation. An HIF-1 to HIF-2 transition or switch governs this process from acute to prolonged hypoxia. In the present study, we evaluated the mechanisms governing the HIF switch in 10 different primary human ECs from different vascular beds during the early stages of hypoxia. The studies demonstrate that the switch from HIF-1 to HIF-2 constitutes a universal mechanism of cellular adaptation to hypoxic stress and that HIF1A and HIF2A mRNA stability differences contribute to HIF switch. Furthermore, using 4 genome-wide mRNA expression arrays of HUVECs during normoxia and after 2, 8, and 16 h of hypoxia, we show using bioinformatics analyses that, although a number of genes appeared to be regulated exclusively by HIF-1 or HIF-2, the largest number of genes appeared to be regulated by both.-Bartoszewski, R., Moszynska, A., Serocki, M., Cabaj, A., Polten, A., Ochocka, R., Dell'Italia, L., Bartoszewska, S., Kroliczewski, J., Dąbrowski, M., Collawn, J. F. Primary endothelial cell-specific regulation of hypoxia-inducible factor (HIF)-1 and HIF-2 and their target gene expression profiles during hypoxia.
104 citations
National Center for Toxicological Research1, University of Birmingham2, Food and Drug Administration3, Edith Cowan University4, Broad Institute5, Mayo Clinic6, National Institutes of Health7, Research Triangle Park8, Agilent Technologies9, Johns Hopkins University10, Wake Forest University11, University of Kentucky12, University of Arkansas for Medical Sciences13, National Institute of Standards and Technology14, University of Michigan15, United States Environmental Protection Agency16, Thermo Fisher Scientific17, European Bioinformatics Institute18, University of Washington19, University of Georgia20, Imperial College London21
TL;DR: The agreed upon first development steps and priority objectives of a community engagement effort to address current challenges in quality assurance (QA) and quality control (QC) in untargeted metabolomic studies are described.
Abstract: We describe here the agreed upon first development steps and priority objectives of a community engagement effort to address current challenges in quality assurance (QA) and quality control (QC) in untargeted metabolomic studies. This has included (1) a QA and QC questionnaire responded to by the metabolomics community in 2015 which recommended education of the metabolomics community, development of appropriate standard reference materials and providing incentives for laboratories to apply QA and QC; (2) a 2-day ‘Think Tank on Quality Assurance and Quality Control for Untargeted Metabolomic Studies’ held at the National Cancer Institute’s Shady Grove Campus and (3) establishment of the Metabolomics Quality Assurance and Quality Control Consortium (mQACC) to drive forward developments in a coordinated manner.
79 citations
TL;DR: The authors show that in lung cancer, genetic activation of distinct oncogenic receptor tyrosine kinases results in unique metabolic liabilities and, in particular, EGFR aberrant cancers rely on the serine biosynthetic pathway while FGFR aberrants cancers reliance on glycolysis.
Abstract: One of the biggest hurdles for the development of metabolism-targeted therapies is to identify the responsive tumor subsets. However, the metabolic vulnerabilities for most human cancers remain unclear. Establishing the link between metabolic signatures and the oncogenic alterations of receptor tyrosine kinases (RTK), the most well-defined cancer genotypes, may precisely direct metabolic intervention to a broad patient population. By integrating metabolomics and transcriptomics, we herein show that oncogenic RTK activation causes distinct metabolic preference. Specifically, EGFR activation branches glycolysis to the serine synthesis for nucleotide biosynthesis and redox homeostasis, whereas FGFR activation recycles lactate to fuel oxidative phosphorylation for energy generation. Genetic alterations of EGFR and FGFR stratify the responsive tumors to pharmacological inhibitors that target serine synthesis and lactate fluxes, respectively. Together, this study provides the molecular link between cancer genotypes and metabolic dependency, providing basis for patient stratification in metabolism-targeted therapies.
68 citations
TL;DR: The cellular uptake of galectin-3 inhibitors and their potency in the intracellular or extracellular space were compared and it was confirmed that the uptake was significantly higher for the inhibitor with the lowest PSA, as expected.
Abstract: Galectin-3 is a carbohydrate binding protein which has important roles in cancer and immunity. Potent galectin-3 inhibitors have been synthesized, for experimental purposes and potential clinical use. As galectin-3 is implicated in both intra- and extracellular activities, permeability of galectin-3 inhibitors is an important parameter determining biological effects. We compared the cellular uptake of galectin-3 inhibitors and their potency in the intracellular or extracellular space. The inhibitors differed in their polar surface area (PSA), but had similar affinities for galectin-3. Using a well-established permeability assay, we confirmed that the uptake was significantly higher for the inhibitor with the lowest PSA, as expected. To analyze intracellular activity of the inhibitors, we developed a novel assay based on galectin-3 accumulation around damaged intracellular vesicles. The results show striking differences between the inhibitors intracellular potency, correlating with their PSAs. To test extracellular activity of the inhibitors, we analyzed their potency to block binding of galectin-3 to cell surfaces. All inhibitors were equally able to block galectin-3 binding to cells and this was proportional to their affinity for galectin-3. These inhibitors may serve as useful tools in exploring biological roles of galectin-3 and may further our understanding of intracellular versus extracellular roles of galectin-3.
66 citations
TL;DR: The range of compound classes detected and different PFAS signatures between sample locations demonstrate the need for expanded quantitation lists when investigating PFAS, especially newer classes in aqueous environmental samples.
Abstract: A quantitative method for the determination of per- and polyfluoroalkyl substances (PFAS) using liquid chromatography (LC) tandem mass spectrometry (MS/MS) was developed and applied to aqueous wastewater, surface water, and drinking water samples. Fifty-three PFAS from 14 compound classes (including many contaminants of emerging concern) were measured using a single analytical method. After solid-phase extraction using weak anion exchange cartridges, method detection limits in water ranged from 0.28 to 18 ng/L and method quantitation limits ranged from 0.35 to 26 ng/L. Method accuracy ranged from 70 to 127% for 49 of the 53 extracted PFAS, with the remaining four between 66 and 138%. Method precision ranged from 2 to 28% RSD, with 49 out of the 53 PFAS being below 50 PFAS, many of which are currently unregulated in the environment and not included in typical analytical lists, this method has efficiency advantages over other similar methods as it utilizes a single chromatographic separation with a shorter runtime (14 min), while maintaining method accuracy and stability and the separation of branched and linear PFAS isomers. The method was applied to wastewater influent and effluent; surface water from a river, wetland, and lake; and drinking water samples to survey PFAS contamination in Australian aqueous matrices. The compound classes FTCAs, FOSAAs, PFPAs, and diPAPs were detected for the first time in Australian WWTPs and the method was used to quantify PFAS concentrations from 0.60 to 193 ng/L. The range of compound classes detected and different PFAS signatures between sample locations demonstrate the need for expanded quantitation lists when investigating PFAS, especially newer classes in aqueous environmental samples.
60 citations
TL;DR: In this article, the volatiles of four types of Japanese green tea (Sencha, Matcha, Gyokuro, and Hojicha) were extracted separately by headspace solid-phase microextraction (HS-SPME) and solvent-assisted flavour evaporation (SAFE), and then analyzed using GC-MS.
Abstract: Japanese green tea is becoming increasingly popular worldwide due to its pleasant aroma. The volatiles of four types of Japanese green tea (Sencha, Matcha, Gyokuro, and Hojicha) were extracted separately by headspace solid-phase microextraction (HS-SPME) and solvent-assisted flavour evaporation (SAFE), and then analysed using gas chromatography-mass spectrometry (GC-MS). The aroma-active compounds in each type of green tea were identified by aroma extract dilution analysis (AEDA) of its SAFE extract. The odourants exhibiting the highest flavour dilution (FD) factor of 27 in Sencha were indole and methional. For Matcha, nonanal had the highest FD factor of 243. The odourant in Gyokuro with the highest FD factor of 27 was indole. For Hojicha, the odour-active compounds exhibiting high FD factor of 729 and 243 were 2,3-diethylpyrazine and 2,3-diethyl-5-methylpyrazine, respectively. Sensory evaluation was then done on the SAFE extract to understand the overall aroma profile of each type of green tea. Based on sensory evaluation results, steamed green teas (Sencha, Matcha, and Gyokuro) had leafier and higher floral notes while roasted green tea (Hojicha) showed higher roasty and woody notes. Finally, principal component analysis (PCA) of data obtained from GC-MS analysis demonstrated marked separation of these four types of Japanese green tea.
55 citations
Okayama University1, Tokyo Medical and Dental University2, Kanazawa University3, University of Toyama4, Toyama Prefectural University5, Tokyo University of Marine Science and Technology6, Asia University (Japan)7, Asahi University8, Agilent Technologies9, Waseda University10, Kyushu University11, University of Tokyo12, Tohoku University13, Niigata University14, Tsurumi University15, Kanagawa Dental College16, American Physical Therapy Association17, Gunma University18, Japan Aerospace Exploration Agency19, Komatsu University20
TL;DR: This is the first study to report the inhibitory effect of melatonin on osteoclastic activation by microgravity and observed a novel action pathway ofmelatonin on fractures via an increase in CALCITONIN secretion, which could be the source of a potential novel drug to prevent bone loss during space flight.
Abstract: Astronauts experience osteoporosis-like loss of bone mass because of microgravity conditions during space flight. To prevent bone loss, they need a riskless and antiresorptive drug. Melatonin is reported to suppress osteoclast function. However, no studies have examined the effects of melatonin on bone metabolism under microgravity conditions. We used goldfish scales as a bone model of coexisting osteoclasts and osteoblasts and demonstrated that mRNA expression level of acetylserotonin O-methyltransferase, an enzyme essential for melatonin synthesis, decreased significantly under microgravity. During space flight, microgravity stimulated osteoclastic activity and significantly increased gene expression for osteoclast differentiation and activation. Melatonin treatment significantly stimulated Calcitonin (an osteoclast-inhibiting hormone) mRNA expression and decreased the mRNA expression of receptor activator of nuclear factor κB ligand (a promoter of osteoclastogenesis), which coincided with suppressed gene expression levels for osteoclast functions. This is the first study to report the inhibitory effect of melatonin on osteoclastic activation by microgravity. We also observed a novel action pathway of melatonin on osteoclasts via an increase in CALCITONIN secretion. Melatonin could be the source of a potential novel drug to prevent bone loss during space flight.
54 citations
TL;DR: It is concluded that a single NTA method, even when optimized, can likely characterize only a subset of ToxCast substances (and, by extension, other CECs) and Rigorous quality control and self-evaluation practices should be required of labs generating NTA data to support exposure and health studies.
Abstract: Non-targeted analysis (NTA) methods are increasingly used to discover contaminants of emerging concern (CECs), but the extent to which these methods can support exposure and health studies remains to be determined. EPA's Non-Targeted Analysis Collaborative Trial (ENTACT) was launched in 2016 to address this need. As part of ENTACT, 1269 unique substances from EPA's ToxCast library were combined to make ten synthetic mixtures, with each mixture containing between 95 and 365 substances. As a participant in the trial, we first performed blinded NTA on each mixture using liquid chromatography (LC) coupled with high-resolution mass spectrometry (HRMS). We then performed an unblinded evaluation to identify limitations of our NTA method. Overall, at least 60% of spiked substances could be observed using selected methods. Discounting spiked isomers, true positive rates from the blinded and unblinded analyses reached a maximum of 46% and 65%, respectively. An overall reproducibility rate of 75% was observed for substances spiked into more than one mixture and observed at least once. Considerable discordance in substance identification was observed when comparing a subset of our results derived from two separate reversed-phase chromatography methods. We conclude that a single NTA method, even when optimized, can likely characterize only a subset of ToxCast substances (and, by extension, other CECs). Rigorous quality control and self-evaluation practices should be required of labs generating NTA data to support exposure and health studies. Accurate and transparent communication of performance results will best enable meaningful interpretations and defensible use of NTA data. Graphical abstract ᅟ.
TL;DR: It is demonstrated that the reported LC-MS/MS method provides a quantitative analysis of antioxidants in plant tissues, for example to investigate interactions between plants and nanomaterials.
TL;DR: The GENESIS study allowed us to specify BC relative risks associated with deleterious‐predicted variants in PALB2, ATM and CHEK2 and to add MAST1, POLH, RTEL1 and FANCI to the list of DNA repair genes possibly involved in BC susceptibility.
Abstract: Pathogenic variants in BRCA1 and BRCA2 only explain the underlying genetic cause of about 10% of hereditary breast and ovarian cancer families. Because of cost-effectiveness, multigene panel testing is often performed even if the clinical utility of testing most of the genes remains questionable. The purpose of our study was to assess the contribution of rare, deleterious-predicted variants in DNA repair genes in familial breast cancer (BC) in a well-characterized and homogeneous population. We analyzed 113 DNA repair genes selected from either an exome sequencing or a candidate gene approach in the GENESIS study, which includes familial BC cases with no BRCA1 or BRCA2 mutation and having a sister with BC (N = 1,207), and general population controls (N = 1,199). Sequencing data were filtered for rare loss-of-function variants (LoF) and likely deleterious missense variants (MV). We confirmed associations between LoF and MV in PALB2, ATM and CHEK2 and BC occurrence. We also identified for the first time associations between FANCI, MAST1, POLH and RTEL1 and BC susceptibility. Unlike other associated genes, carriers of an ATM LoF had a significantly higher risk of developing BC than carriers of an ATM MV (ORLoF = 17.4 vs. ORMV = 1.6; p Het = 0.002). Hence, our approach allowed us to specify BC relative risks associated with deleterious-predicted variants in PALB2, ATM and CHEK2 and to add MAST1, POLH, RTEL1 and FANCI to the list of DNA repair genes possibly involved in BC susceptibility. We also highlight that different types of variants within the same gene can lead to different risk estimates.
TL;DR: The influence of a longer peak width on the accuracy of SP-ICP-MS measurement data (NP size, particle number density and mass concentration) must be taken into account when using CRC technology as a means to overcome spectral overlap.
TL;DR: This comprehensive study on the glycosides changes revealed the molecular bases for GBVs increments during the pan firing and showed that enzymatic synthesis contributed to the increase in the content of GBVs during thepan firing.
TL;DR: HLA loss provides genetic evidence that allogeneic immune recognition may be mediated by minor histocompatibility antigens and suggests opportunities for novel immunologic approaches for relapse prevention.
TL;DR: The results demonstrated that the method effectively removes unwanted matrix co-extractives better than conventional PSA/C18 dispersive solid phase extraction (dSPE) cleanup, while still delivering acceptable recovery results (recoveries of 60-120%) for the majority of pesticides tested.
TL;DR: This is the first report of a pH-gradient LC–MS/MS method for glyphosate and AMPA analysis at ultratrace levels, with minimal sample processing, avoiding costly, time-consuming derivatization and preconcentration steps.
Abstract: Glyphosate is currently the most widely used herbicide in the world, yet screening of environmental waters for this chemical is limited by the need for specialized derivatization and measurement methods that can be tedious and time-consuming. In this work, we present a novel method for the detection and quantification at trace levels of glyphosate and aminomethylphosphonic acid (AMPA) in environmental water samples. The detection and quantification of the analytes was performed by liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS). Chromatographic separation was achieved with an ion-exchange column and a pH-gradient elution of a solution of ammonium hydroxide and ammonium acetate. The limit of detection for glyphosate and AMPA was 0.25 μg L-1 and the limit of quantification was 0.5 μg L-1with a 20-μL injection. The method was used to investigate the levels of glyphosate and AMPA in surface water samples from the Yarra River catchment area and urban constructed stormwater wetlands. The results indicate that at the time of sampling, no glyphosate or AMPA was present in the samples from the Yarra River catchment area (n = 10). However, glyphosate was detected above the limit of quantification in 33% of the wetland samples (n = 12), with concentrations ranging from 1.95 to 2.96 μg L-1. Similarly, AMPA was quantified in 83% of the wetland samples, with concentrations ranging from 0.55 to 2.42 μg L-1. To our knowledge, this is the first report of a pH-gradient LC-MS/MS method for glyphosate and AMPA analysis at ultratrace levels, with minimal sample processing, avoiding costly, time-consuming derivatization and preconcentration steps. Graphical abstract ᅟ.
TL;DR: Can reduced representation genome sequencing offer an alternative to single nucleotide polymorphism (SNP) arrays as a generic and genome-wide approach for comprehensive preimplantation genetic testing for monogenic disorders, aneuploidy and structural rearrangements in human embryo biopsy samples?
Abstract: Study question Can reduced representation genome sequencing offer an alternative to single nucleotide polymorphism (SNP) arrays as a generic and genome-wide approach for comprehensive preimplantation genetic testing for monogenic disorders (PGT-M), aneuploidy (PGT-A) and structural rearrangements (PGT-SR) in human embryo biopsy samples? Summary answer Reduced representation genome sequencing, with OnePGT, offers a generic, next-generation sequencing-based approach for automated haplotyping and copy-number assessment, both combined or independently, in human single blastomere and trophectoderm samples. What is known already Genome-wide haplotyping strategies, such as karyomapping and haplarithmisis, have paved the way for comprehensive PGT, i.e. leveraging PGT-M, PGT-A and PGT-SR in a single workflow. These methods are based upon SNP array technology. Study design, size, duration This multi-centre verification study evaluated the concordance of PGT results for a total of 225 embryos, including 189 originally tested for a monogenic disorder and 36 tested for a translocation. Concordance for whole chromosome aneuploidies was also evaluated where whole genome copy-number reference data were available. Data analysts were kept blind to the results from the reference PGT method. Participants/materials, setting, methods Leftover blastomere/trophectoderm whole genome amplified (WGA) material was used, or secondary trophectoderm biopsies were WGA. A reduced representation library from WGA DNA together with bulk DNA from phasing references was processed across two study sites with the Agilent OnePGT solution. Libraries were sequenced on an Illumina NextSeq500 system, and data were analysed with Agilent Alissa OnePGT software. The embedded PGT-M pipeline utilises the principles of haplarithmisis to deduce haplotype inheritance whereas both the PGT-A and PGT-SR pipelines are based upon read-count analysis in order to evaluate embryonic ploidy. Concordance analysis was performed for both analysis strategies against the reference PGT method. Main results and the role of chance PGT-M analysis was performed on 189 samples. For nine samples, the data quality was too poor to analyse further, and for 20 samples, no result could be obtained mainly due to biological limitations of the haplotyping approach, such as co-localisation of meiotic crossover events and nullisomy for the chromosome of interest. For the remaining 160 samples, 100% concordance was obtained between OnePGT and the reference PGT-M method. Equally for PGT-SR, 100% concordance for all 36 embryos tested was demonstrated. Moreover, with embryos originally analysed for PGT-M or PGT-SR for which genome-wide copy-number reference data were available, 100% concordance was shown for whole chromosome copy-number calls (PGT-A). Limitations, reasons for caution Inherent to haplotyping methodologies, processing of additional family members is still required. Biological limitations caused inconclusive results in 10% of cases. Wider implications of the findings Employment of OnePGT for PGT-M, PGT-SR, PGT-A or combined as comprehensive PGT offers a scalable platform, which is inherently generic and thereby, eliminates the need for family-specific design and optimisation. It can be considered as both an improvement and complement to the current methodologies for PGT. Study funding/competing interest(s) Agilent Technologies, the KU Leuven (C1/018 to J.R.V. and T.V.) and the Horizon 2020 WIDENLIFE (692065 to J.R.V. and T.V). H.M. is supported by the Research Foundation Flanders (FWO, 11A7119N). M.Z.E, J.R.V. and T.V. are co-inventors on patent applications: ZL910050-PCT/EP2011/060211- WO/2011/157846 'Methods for haplotyping single cells' and ZL913096-PCT/EP2014/068315 'Haplotyping and copy-number typing using polymorphic variant allelic frequencies'. T.V. and J.R.V. are co-inventors on patent application: ZL912076-PCT/EP2013/070858 'High-throughput genotyping by sequencing'. Haplarithmisis ('Haplotyping and copy-number typing using polymorphic variant allelic frequencies') has been licensed to Agilent Technologies. The following patents are pending for OnePGT: US2016275239, AU2014345516, CA2928013, CN105874081, EP3066213 and WO2015067796. OnePGT is a registered trademark. D.L., J.T. and R.L.R. report personal fees during the conduct of the study and outside the submitted work from Agilent Technologies. S.H. and K.O.F. report personal fees and other during the conduct of the study and outside the submitted work from Agilent Technologies. J.A. reports personal fees and other during the conduct of the study from Agilent Technologies and personal fees from Agilent Technologies and UZ Leuven outside the submitted work. B.D. reports grants from IWT/VLAIO, personal fees during the conduct of the study from Agilent Technologies and personal fees and other outside the submitted work from Agilent Technologies. In addition, B.D. has a patent 20160275239 - Genetic Analysis Method pending. The remaining authors have no conflicts of interest.
TL;DR: The metabolically engineered yeast cell factories provide an alternative approach to glycyrrhetinic acid production, replacing the traditional method of plant extraction.
Abstract: Glycyrrhetinic acid (GA) is the most important ingredient in licorice due to its outstanding anti-inflammatory activity and wide application in the medicine and cosmetics industries. Contemporary industrial production of GA by acid hydrolysis of glycyrrhizin which was extracted from Glycyrrhiza plants, is not environment-friendly and devastates farmland since the Glycyrrhiza rhizomes grow up to 10 m underground. In this study, GA was produced through metabolically engineering Saccharomyces cerevisiae by introducing the entire heterogeneous biosynthetic pathway of GA. Codon optimized CYP88D6 and CYP72A154, combined with β-AS (β-amyrin synthase encoding gene) and the NADPH-cytochrome P450 reductase gene of Arabidopsis thaliana were introduced into S. cerevisiae. The resulting strain (Y1) produced 2.5 mg/L of β-amyrin and 14 μg/L of GA. The cytochrome b5 from G. uralensis (GuCYB5) was identified and the introduction of this novel GuCYB5 increased the efficiency of GA production by eightfold. The joint utilization of the GuCYB5 gene along with 10 known MVA pathway genes from S. cerevisiae were overexpressed in a stable chromosome integration to achieve higher GA production. Using the combined strategy, GA concentration improved by 40-fold during batch fermentation. The production was further improved to 8.78 mg/L in fed-batch fermentation, which was increased by a factor of nearly 630. This study first investigated the influence of carbon flux in the upstream module and the introduction of a newly identified GuCYB5 on GA production. The newly identified GuCYB5 was highly effective in improving GA production. An integrated strategy including enzyme discovery, pathway optimization, and fusion protein construction was provided in improving GA production, achieving a 630 fold increase in GA production. The metabolically engineered yeast cell factories provide an alternative approach to glycyrrhetinic acid production, replacing the traditional method of plant extraction.
TL;DR: In this article, the concentrations of two cyclic phosphonates (PMMMPs) were quantitatively measured for the first time in the North American environment and were much higher than those of other flame retardants.
Abstract: Brominated and phosphorus-based flame retardants (PFRs) were measured in foam and fabric samples from 18 newly marketed children’s car seats. The concentrations of two cyclic phosphonates {PMMMPs, 5-ethyl-2-methyl-2-oxido-1,3,2-dioxaphosphinan-5-yl)methyl methyl methylphosphonate and bis[(5-ethyl-2-methyl-1,3,2-dioxaphosphorinan-5-yl)methyl] methyl phosphonate p,p′-dioxide} were quantitatively measured for the first time in the North American environment and were much higher than those of other flame retardants. Median PMMMP concentrations were 73.6 μg/g, accounting on average for 52% of the total FR concentrations, indicating an intentional addition of PMMMPs during the manufacturing process of these car seats. Two other emerging PFRs [tris(2,4-di-Hi Katie,t-butylphenyl) phosphate (TDTBPP) and resorcinol bis(diphenyl phosphate) (RDP)] were detected for the first time in baby products at median levels of 1.11 and 6.15 μg/g, respectively. Other frequently detected PFRs included triethyl phosphate (TEP), tr...
TL;DR: This work demonstrates the genetic origin of tubulopathies in one out of three adult patients, half of the rate observed in children, and enhances the understanding of complex phenotypes and enriches the database with the causal variants described.
TL;DR: By understanding monoterpene glycoconjugation in wine grapes, scientists and winemakers can better understand grape and wine aromas and align with underlying genetic relationships among cultivars.
TL;DR: The optimized method was successfully applied to investigate the removal efficiency of emerging contaminants in a drinking water treatment plant and to analyze the environmental occurrence of target analytes, and has the potential to advance large-scale environmental monitoring programs.
TL;DR: In this article, the trace element concentration pattern in muscle and liver of different age groups differed, may be according to the different diet types of the groups, and no differentiation among the age groups based on the trace elements in the gills was observed.
TL;DR: This workflow uses a reversed-phase LC/DTIM-MS workflow able to profile and quantify the oxylipin and fatty acid content of biological samples while simultaneously acquiring full scan and product ion spectra, confirming its ability in providing further insights into lipid analysis of Biological samples.
Abstract: Analysis of oxylipins by liquid chromatography mass spectrometry (LC/MS) is challenging because of the small mass range occupied by this diverse lipid class, the presence of numerous structural isomers, and their low abundance in biological samples. Although highly sensitive LC/MS/MS methods are commonly used, further separation is achievable by using drift tube ion mobility coupled with high-resolution mass spectrometry (DTIM-MS). Herein, we present a combined analytical and computational method for the identification of oxylipins and fatty acids. We use a reversed-phase LC/DTIM-MS workflow able to profile and quantify (based on chromatographic peak area) the oxylipin and fatty acid content of biological samples while simultaneously acquiring full scan and product ion spectra. The information regarding accurate mass, collision-cross-section values in nitrogen (DTCCSN2), and retention times of the species found are compared to an internal library of lipid standards as well as the LIPID MAPS Structure Database by using specifically developed processing tools. Features detected within the DTCCSN2 and m/ z ranges of the analyzed standards are flagged as oxylipin-like species, which can be further characterized using drift-time alignment of product and precursor ions distinctive of DTIM-MS. This not only helps identification by reducing the number of annotations from LIPID MAPS but also guides discovery studies of potentially novel species. Testing the methodology on Salmonella enterica serovar Typhimurium-infected murine bone-marrow-derived macrophages and thrombin activated human platelets yields results in agreement with literature. This workflow has also annotated features as potentially novel oxylipins, confirming its ability in providing further insights into lipid analysis of biological samples.
TL;DR: Most metabolites, including catechins, flavonol/flavone glycosides, amino acids, nucleosides, organic acids, aroma precursors, lipids, and carbohydrates, decreased with increasing storage duration, while 8-C N-ethyl-2-pyrrolidinone-substituted flavan-3-ols (EPSFs) and pyroglutamic acid increased.
TL;DR: The objective of this work is to determine if hypothesis testing using Adverse Outcome Pathways (AOPs) can provide quantitative chemical hazard predictions and to conclude that quantitative AOPs provide a flexible hypothesis framework for predicting chemical hazards.
Abstract: Current efforts in chemical safety are focused on utilizing human in vitro or alternatives to animal data in a biological pathway context. However, it remains unclear how biological pathways, and toxicology data developed in that context, can be used to quantitatively facilitate decision-making. The objective of this work is to determine if hypothesis testing using adverse outcome pathways (AOPs) can provide quantitative chemical hazard predictions. Current methods for predicting hazards of chemicals in a biological pathway context were extensively reviewed, specific case studies examined, and computational modeling used to demonstrate quantitative hazard prediction based on an AOP. Since AOPs are chemically agnostic, we propose that AOPs function as hypotheses for how specific chemicals may cause adverse effects via specific pathways. Three broad approaches were identified for testing the hypothesis with AOPs, semi-quantitative weight of evidence, probabilistic, and mechanistic modeling. We then demonstrate how these approaches could be used to test hypotheses using high throughput in vitro data and data from alternatives to animal testing. Finally, we discuss standards in development and documentation that would facilitate use in a regulatory context. We conclude that quantitative AOPs provide a flexible hypothesis framework for predicting chemical hazards, which accommodates a wide range of approaches that are useful at many stages and build upon one another to become increasingly quantitative.
TL;DR: It is suggested that exposure to DDTs may contribute to the risk of metabolic disease among Asian Indians by affecting hepatic fat levels independent of obesity.
Abstract: Persistent organic pollutants (POPs), such as dichlorodiphenyltrichloroethane (DDT) and other organochlorine compounds, are abundant in the environment and in foodstuffs from the Indian subcontinent. These environmental contaminants have been associated with a higher risk of diabetes in numerous studies. Asian Indians are well known to have a high risk of diabetes compared with other populations, and this risk is also found in migrant populations of Asian Indians in the United States, Europe, and elsewhere. We hypothesized that high plasma concentrations of POPs in Asian Indian migrants are linked to a variety of diabetes-related pathologies and explored the mechanism for the induction of these effects. We measured 30 environmental pollutants in plasma samples obtained from 147 participants in the Metabolic syndrome and Atherosclerosis in South Asians Living in America pilot study using a gas chromatography-tandem mass spectrometry analytical method that uses less than 0.5 mL of plasma. We found that plasma levels of o,p'-DDT and p,p'-DDT were independently associated with both body mass index (BMI) and waist circumference. Doubling the levels of the sums of these DDTs was associated with insulin insensitivity (-0.38 Matsuda index, p = 0.001), increased adiposity (1.26 kg/m2 BMI and 3.58 cm waist circumference increase, p < 0.0001), circulating insulin (12.9 mIU/L, p = 0.002), hepatic fat (-0.051 HU, p = 0.001), as well as increased odds of obesity (OR = 2.17, p < 0.001, BMI-based; OR = 2.37, p = 0.001, waist-based), prediabetes (OR = 1.55, p = 0.02), diabetes (OR = 1.72, p = 0.01), and fatty liver (OR = 1.66, p = 0.01) in multivariable models accounting for confounding by age, sex, years in the US, education, and fish protein. Furthermore, levels of DDTs were associated with increased hepatic fat and circulating insulin, independent of obesity and confounders. These findings suggest that exposure to DDTs may contribute to the risk of metabolic disease among Asian Indians by affecting hepatic fat levels independent of obesity.
TL;DR: ociation between metabolites and COPD phenotypes are more robustly represented in BAL compared to plasma, and three BAL co-expression modules associated with FEV1/FVC and emphysema are identified.
Abstract: Smoking causes chronic obstructive pulmonary disease (COPD). Though recent studies identified a COPD metabolomic signature in blood, no large studies examine the metabolome in bronchoalveolar lavage (BAL) fluid, a more direct representation of lung cell metabolism. We performed untargeted liquid chromatography-mass spectrometry (LC-MS) on BAL and matched plasma from 115 subjects from the SPIROMICS cohort. Regression was performed with COPD phenotypes as the outcome and metabolites as the predictor, adjusted for clinical covariates and false discovery rate. Weighted gene co-expression network analysis (WGCNA) grouped metabolites into modules which were then associated with phenotypes. K-means clustering grouped similar subjects. We detected 7939 and 10,561 compounds in BAL and paired plasma samples, respectively. FEV1/FVC (Forced Expiratory Volume in One Second/Forced Vital Capacity) ratio, emphysema, FEV1 % predicted, and COPD exacerbations associated with 1230, 792, eight, and one BAL compounds, respectively. Only two plasma compounds associated with a COPD phenotype (emphysema). Three BAL co-expression modules associated with FEV1/FVC and emphysema. K-means BAL metabolomic signature clustering identified two groups, one with more airway obstruction (34% of subjects, median FEV1/FVC 0.67), one with less (66% of subjects, median FEV1/FVC 0.77; p < 2 × 10-4). Associations between metabolites and COPD phenotypes are more robustly represented in BAL compared to plasma.