Summary of Study ST000890

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000619. The data can be accessed directly via it's Project DOI: 10.21228/M8NT1F This work is supported by NIH grant, U2C- DK119886.

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Study IDST000890
Study TitleIdentification of RXR Ligands
Study TypeIdenfication of Ligands by HPLC-MS
Study SummaryFree fatty acids in mouse plasma were identified and quantified by LC-MS. Through differential feeding and PHZ (phnylhydrazine) dosing, coupled with mass spectrometry, we identified the long chain fatty acid C24:5 as a natural RXRA ligand, which was dynamically increased in concentration in response to hematopoietic stress. Collectively, these data demonstrate that natural RXRA ligands are present and are dynamically regulated in vivo in mouse hematopoietic cells.
Institute
Washington University in St. Louis
DepartmentDiabetic Cardiovascular Disease Center, School of Medicine
LaboratoryMetabolomics Core
Last NameFujiwara
First NameHideji
Address660 South Euclid Ave, St. Louis MO 63110
Emailhfujiwar@wustl.edu
Phone314-747-0494
Submit Date2017-09-22
Study CommentsUnits of measurement:peak area ratio: analyte peak area/peak area of internal standard
Analysis Type DetailLC-MS
Release Date2017-10-22
Release Version1
Hideji Fujiwara Hideji Fujiwara
https://dx.doi.org/10.21228/M8NT1F
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000619
Project DOI:doi: 10.21228/M8NT1F
Project Title:Endogenous Retinoid X Receptor Ligands in Mouse Hemotopoietic Cells
Project Type:MS Identification and Quantification of Ligands
Project Summary:The retinoid X receptor α (RXRA) has been implicated in diverse hematological processes. However, it is unknown whether natural ligands of RXRA are present or regulated in hematopoietic cells. We quantified lipids in the serum of mice treated with a vitamin A deficient diet, a fatty acid deficient diet, and following phenylhydrazine treatment. In parallel, these serum samples were applied to RXRA reporter cells to identify conditions which contained increased concentrations of natural RXRA ligands. Mass spectrometry quantification of serum lipids was correlated with data from the RXRA reporter cells to identify serum lipids that increased in conditions associated with augmented RXRA ligand concentrations in the RXRA reporter assay.
Institute:Washington University in St. Louis
Department:Diabetic Cardiovascular Disease Center
Laboratory:Metabolomics Core
Last Name:Fujiwara
First Name:Hideji
Address:660 South Euclide Avenue, St. Lois, MO 63110
Email:hfujiwar@wustl.edu
Phone:314-747-0494
Funding Source:NIH R01 HL128447, NIH P30 DK020579
Publications:Science Signaling 2018, being accepted for publication
Contributors:Haixia Niu, Hideji Fujiwara, Orsola di Martino, Gayla Hadwiger, Thomas E. Frederick, María P. Menéndez-Gutiérrez, Mercedes Ricote, Gregory R. Bowman, John S. Welch

Subject:

Subject ID:SU000927
Subject Type:Animals
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57/B6
Age Or Age Range:8-16 weeks
Species Group:Mammal

Factors:

Subject type: Animals; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Diet of subject mice Feeding period (wk) PHZ dose (mg)
SA0524494630 BM2-PHZcontrol chow 12 1
SA052450VADC-PHZ-4control chow 12 1
SA0524514629 BM2-PHZcontrol chow 12 1
SA0524524629 BM1-PHZcontrol chow 12 1
SA052453VADC-PHZ-3control chow 12 1
SA0524544630 PB1-PHZcontrol chow 12 1
SA0524554630 BM1-PHZcontrol chow 12 1
SA0524564629 PB1-PHZcontrol chow 12 1
SA0524574630 PB2-PHZcontrol chow 12 1
SA0524584629 PB2-PHZcontrol chow 12 1
SA052459VADC-PHZ-1control chow 12 1
SA052460VADC-PHZ-2control chow 12 1
SA0524614633 PB1control chow 12 mock
SA052462VDAC41control chow 12 mock
SA052463VADC-23control chow 12 mock
SA052464VADC-13control chow 12 mock
SA052465VADC-33control chow 12 mock
SA052466VADC-43control chow 12 mock
SA052467VADC-53control chow 12 mock
SA052468VDAC52control chow 12 mock
SA052469VDAC42control chow 12 mock
SA052470VDAC21control chow 12 mock
SA052471VDAC51control chow 12 mock
SA052472VDAC22control chow 12 mock
SA052473VDAC32control chow 12 mock
SA0524744632 PB1control chow 12 mock
SA052475VDAC31control chow 12 mock
SA052476VADC-2control chow 12 mock
SA052477VADC-3control chow 12 mock
SA052478VADC-1control chow 12 mock
SA0524794632 PB2control chow 12 mock
SA0524804632 BM2control chow 12 mock
SA0524814633 PB2control chow 12 mock
SA052482VADC-4control chow 12 mock
SA0524834633 BM2control chow 12 mock
SA0524844632 BM1control chow 12 mock
SA0524854633 BM1control chow 12 mock
SA052486VADC-PHZ-5control chow 12 mock
SA052487NFC-PHZ-22control chow 4 1
SA052488NFC-PHZ-23control chow 4 1
SA052489NFC-PHZ-12control chow 4 1
SA052490NFC-PHZ-21control chow 4 1
SA052491NFC-PHZ-13control chow 4 1
SA052492NFC-PHZ-11control chow 4 1
SA052493FC-31control chow 4 mock
SA052494FC-21control chow 4 mock
SA052495FC-41control chow 4 mock
SA052496FC-12control chow 4 mock
SA052497FC-11control chow 4 mock
SA052498FC-42control chow 4 mock
SA052499FC-22control chow 4 mock
SA052500FC-32control chow 4 mock
SA052418NF-PHZ-41Non-fat-diet 4 1
SA052419NF-PHZ-31Non-fat-diet 4 1
SA052420NF-PHZ-32Non-fat-diet 4 1
SA052421NF-PHZ-23Non-fat-diet 4 1
SA052422NF-PHZ-43Non-fat-diet 4 1
SA052423NF-PHZ-21Non-fat-diet 4 1
SA052424NF-PHZ-42Non-fat-diet 4 1
SA052425NF-PHZ-33Non-fat-diet 4 1
SA052426F-41Non-fat-diet 4 mock
SA052427F-31Non-fat-diet 4 mock
SA052428F-51Non-fat-diet 4 mock
SA052429NF-11Non-fat-diet 4 mock
SA052430NF-23Non-fat-diet 4 mock
SA052431NF-33Non-fat-diet 4 mock
SA052432F-21Non-fat-diet 4 mock
SA052433F-11Non-fat-diet 4 mock
SA052434NF-2Non-fat-diet 4 mock
SA052435NF-3Non-fat-diet 4 mock
SA052436NF-1Non-fat-diet 4 mock
SA052437NF-31Non-fat-diet 4 mock
SA052438NF-13Non-fat-diet 4 mock
SA052439NF-12Non-fat-diet 4 mock
SA052440NF-21Non-fat-diet 4 mock
SA052441NF-22Non-fat-diet 4 mock
SA052442NF-PHZ-22Non-fat-diet 4 mock
SA052443F-12Non-fat-diet 4 mock
SA052444F-52Non-fat-diet 4 mock
SA052445NF-32Non-fat-diet 4 mock
SA052446F-42Non-fat-diet 4 mock
SA052447F-32Non-fat-diet 4 mock
SA052448F-22Non-fat-diet 4 mock
SA052501VAD-PHZ2-32vitamin A deficient diet 12 1
SA052502VAD-PHZ2-12vitamin A deficient diet 12 1
SA052503VAD-PHZ2-42vitamin A deficient diet 12 1
SA052504VAD-PHZ2-22vitamin A deficient diet 12 1
SA052505VAD-PHZ1-11vitamin A deficient diet 12 1
SA052506VAD-PHZ1-31vitamin A deficient diet 12 1
SA052507VAD-PHZ1-21vitamin A deficient diet 12 1
SA052508VAD-PHZ2-52vitamin A deficient diet 12 1
SA052509VAD-PHZ1-41vitamin A deficient diet 12 1
SA052510VAD-PHZ2-51vitamin A deficient diet 12 1
SA052511VAD-PHZ1-42vitamin A deficient diet 12 1
SA052512VAD-PHZ1-22vitamin A deficient diet 12 1
SA052513VAD-PHZ2-41vitamin A deficient diet 12 1
SA052514VAD-PHZ2-31vitamin A deficient diet 12 1
SA052515VAD-PHZ2-11vitamin A deficient diet 12 1
SA052516VAD-PHZ2-21vitamin A deficient diet 12 1
SA052517VAD-PHZ1-12vitamin A deficient diet 12 1
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Collection:

Collection ID:CO000921
Collection Summary:Mouse plasma was collected by obtaining peripheral blood via peri-orbital bleeding. Approximately 100 ul blood is placed in a microvette vial with pre-loaded KEDTA and then centrifuged at 14,000 rpm for 10 minutes at 4 degrees. Plasma is removed and stored at -80 C until analysis.
Sample Type:Blood

Treatment:

Treatment ID:TR000941
Treatment Summary:Phenylhydrazine was administered at 1mg once on day 1 and plasma was collected on day 4. Vitamin A deficient diet: Teklad, TD.86143. Control diet: Teklad, TD.91280. Fatty acid deficient diet: Teklad, TD.03314. Control diet: Teklad, TD.130321.

Sample Preparation:

Sampleprep ID:SP000934
Sampleprep Summary:For free fatty acids analysis, 100 μL of plasma extraction sample or 400 μL pull-down samples was used for DMAPA (dimethylaminopropylamine) derivatization for improving MS sensitivity of free fatty acids. Prior to the derivatization, 50 ng of AA-d8 (deuterated arachidonic acid –d8) as the internal standard was added to each sample. The solvents in the sample were dried under a stream of nitrogen. To derivatize the sample, 50 μL of 50 mM EDC (N-ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride) and 50 mM DMAPA and 50 mM DMAP (4-dimethylaminopyridine) were added to the dried fatty acid samples and heated at 50 ◦C for 30 minutes. The samples were dried under nitrogen and then dissolved in 1 mL of ethanol : water for MS.

Combined analysis:

Analysis ID AN001452
Analysis type MS
Chromatography type Reversed phase
Chromatography system Shimadzu 20AD
Column Agilent Eclipse XDB-C18 (100 x 3.0mm,3.5um)
MS Type ESI
MS instrument type Triple quadrupole
MS instrument name ABI Sciex API 4000 QTrap
Ion Mode POSITIVE
Units peak area ratio

Chromatography:

Chromatography ID:CH001021
Chromatography Summary:For DMAPA derivatized Fatty acid analysis. Shimadzu 20AD two pump system was used. Solvent A: 0.1% formic acid in water. Solvent B: 0.1% in Acetonitrile. The initial LC condition was 30% B. The solvent gradient was programmed from 30% B to 99% B in 4 min and was hold for 1 min at the flow rate of 1 mL/min. Then, it was immediately brought back to 30 % B and was hold for 2 min. An Agilent Eclipse XDB-C18 column (3.0 x100 mm x 3.5 µm) was used for this analysis.
Instrument Name:Shimadzu 20AD
Column Name:Agilent Eclipse XDB-C18 (100 x 3.0mm,3.5um)
Column Temperature:60 C
Flow Gradient:30% B to 99% B in 4 min
Flow Rate:1 mL/min
Solvent A:0.1% formic acid in water
Solvent B:0.1% formic acid in ACN
Analytical Time:7 min
Chromatography Type:Reversed phase

MS:

MS ID:MS001342
Analysis ID:AN001452
Instrument Name:ABI Sciex API 4000 QTrap
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:Units of measurement:peak area ratio: analyte peak area/peak area of internal standard
Ion Mode:POSITIVE
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