Summary of Study ST001143

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 PR000764. The data can be accessed directly via it's Project DOI: 10.21228/M8XH5B This work is supported by NIH grant, U2C- DK119886.

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Study IDST001143
Study TitleMicrobial depletion and ozone exposure - Lung tissue (part I)
Study SummaryGlobal biochemical profiles were determined in lung tissue collected from untreated control mice and mice treated for two weeks with untreated drinking water or water containing an antibiotic cocktail (ampicillin, neomycin, metronidazole, and vancomycin), followed by a three hour exposure to ambient air or ozone (2ppm). Sample collection occurred 24 hours post-ozone exposure.
Institute
Harvard School of Public Health
Last NameShore
First NameStephanie
Address677 Huntington Ave
Emailsshore@hsph.harvard.edu
Phone6174320989
Submit Date2019-02-26
Raw Data AvailableYes
Analysis Type DetailGC-MS/LC-MS
Release Date2019-05-15
Release Version1
Stephanie Shore Stephanie Shore
https://dx.doi.org/10.21228/M8XH5B
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000764
Project DOI:doi: 10.21228/M8XH5B
Project Title:Microbial depletion and ozone exposure
Project Summary:Ozone is an asthma trigger. In mice, the gut microbiome contributes to ozone-induced airway hyperresponsiveness, a defining feature of asthma. The purpose of this study was to identify metabolites that could be mediating this role. Gut bacterial enzymes modify ingested substances producing metabolites that enter the blood and circulate to host tissues where they may exert a variety of effects. Therefore, we performed global metabolomic profiling on serum of mice after acute ozone exposure. To identify the role of the microbiome in mediating ozone-induced metabolomic changes, mice were treated for two weeks with a cocktail of antibiotics in the drinking water or with control water and then exposed to air or ozone (2 ppm for 3 hours). Twenty four hours later, blood was harvested and serum analyzed via liquid-chromatography or gas-chromatography coupled to mass spectrometry. We observed marked effects of both ozone exposure and antibiotics on the serum metabolome. Known bacterially-derived metabolites were reduced in antibiotic-treated mice. Importantly, our data also indicated that ozone-induced changes in serum lipids, including long chain fatty acids and bile acids, as well as ozone-induced changes in polyamines were different in control and antibiotic-treated mice. Each of these metabolites has the capacity to alter airway responsiveness and may account for the role of the microbiome in pulmonary responses to ozone.
Institute:Harvard School of Public Health
Department:Molecular and Integrative Physiological Sciences
Last Name:Shore
First Name:Stephanie
Address:677 Huntington Ave
Email:sshore@hsph.harvard.edu
Phone:6174320989

Subject:

Subject ID:SU001208
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57BL/6J
Age Or Age Range:8 weeks
Gender:Male
Animal Animal Supplier:Taconic Farms (New York)

Factors:

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

mb_sample_id local_sample_id Treament Exposure
SA079500Mouse 11Antibiotics Air
SA079501Mouse 10Antibiotics Air
SA079502Mouse 12Antibiotics Air
SA079503Mouse 14Antibiotics Air
SA079504Mouse 15Antibiotics Air
SA079505Mouse 9Antibiotics Air
SA079506Mouse 13Antibiotics Air
SA079507Mouse 30Antibiotics Ozone
SA079508Mouse 16Antibiotics Ozone
SA079509Mouse 31Antibiotics Ozone
SA079510Mouse 29Antibiotics Ozone
SA079511Mouse 26Antibiotics Ozone
SA079512Mouse 25Antibiotics Ozone
SA079513Mouse 27Antibiotics Ozone
SA079514Mouse 28Antibiotics Ozone
SA079515Mouse 32Antibiotics Ozone
SA079516Mouse 1Water Air
SA079517Mouse 6Water Air
SA079518Mouse 7Water Air
SA079519Mouse 5Water Air
SA079520Mouse 4Water Air
SA079521Mouse 8Water Air
SA079522Mouse 3Water Air
SA079523Mouse 2Water Air
SA079524Mouse 17Water Ozone
SA079525Mouse 19Water Ozone
SA079526Mouse 18Water Ozone
SA079527Mouse 20Water Ozone
SA079528Mouse 21Water Ozone
SA079529Mouse 23Water Ozone
SA079530Mouse 22Water Ozone
SA079531Mouse 24Water Ozone
Showing results 1 to 32 of 32

Collection:

Collection ID:CO001202
Collection Summary:Global biochemical profiles were determined in lung tissue and serum collected from untreated control mice and mice treated for two weeks with untreated drinking water or water containing an antibiotic cocktail (ampicillin, neomycin, metronidazole, and vancomycin), followed by a three hour exposure to ambient air or ozone (2ppm). Sample collection occurred 24 hours post-ozone exposure.
Collection Protocol Filename:PROTOCOL_AND_REPORT_150324.docx
Sample Type:Lung

Treatment:

Treatment ID:TR001223
Treatment Summary:Global biochemical profiles were determined in lung tissue and serum collected from untreated control mice and mice treated for two weeks with untreated drinking water or water containing an antibiotic cocktail (ampicillin, neomycin, metronidazole, and vancomycin), followed by a three hour exposure to ambient air or ozone (2ppm). Sample collection occurred 24 hours post-ozone exposure.
Treatment Protocol Filename:PROTOCOL_AND_REPORT_150324.docx

Sample Preparation:

Sampleprep ID:SP001216
Sampleprep Summary:Sample preparation: Samples were shipped to Metabolon for processing and prepared for metabolomics as previously described. Briefly, an equivalent amount serum (100µl) on a per sample basis was prepared using the automated MicroLab STAR® system from Hamilton Company. For QC purposes, a recovery standard was added prior to the first step in the extraction process. To remove protein, dissociate small molecules bound to protein or trapped in the precipitated protein matrix, and to recover chemically diverse metabolites, proteins were first precipitated with methanol under vigorous shaking for 2 min (Glen Mills GenoGrinder 2000) followed by centrifugation. The resulting extract was divided into five fractions as follows. One fraction was used for analysis by UPLC-MS/MS with positive ion mode electrospray ionization. Another fraction was used for analysis by UPLC-MS/MS with negative ion mode electrospray ionization. The third and fourth fractions were used for LC polar platform, and for analysis by GC-MS. The final fraction was reserved as a backup. Samples were placed briefly on a TurboVap® (Zymark) to remove the organic solvent. For LC, the samples were stored overnight under nitrogen before preparation for analysis. For GC, each sample was dried under vacuum overnight before preparation for analysis.

Combined analysis:

Analysis ID AN001882 AN001883 AN001884 AN001885
Analysis type MS MS MS MS
Chromatography type Reversed phase/HILIC Reversed phase/HILIC Reversed phase/HILIC GC
Chromatography system Waters Acquity Waters Acquity Waters Acquity Thermo Trace DSQ
Column Waters Acquity BEH C18 (100 x 2mm,1.7um)/Waters UPLC BEH Amide (2.1x150 mm,1.7um) Waters Acquity BEH C18 (100 x 2mm,1.7um)/Waters UPLC BEH Amide (2.1x150 mm,1.7um) Waters Acquity BEH C18 (100 x 2mm,1.7um)/Waters UPLC BEH Amide (2.1x150 mm,1.7um) 5% diphenyl/95% dimethyl polysiloxane GC column
MS Type ESI ESI ESI EI
MS instrument type Orbitrap Orbitrap Orbitrap Single quadrupole
MS instrument name Thermo Q Exactive Plus Orbitrap Thermo Q Exactive Plus Orbitrap Thermo Q Exactive Plus Orbitrap Thermo Trace DSQ
Ion Mode POSITIVE NEGATIVE NEGATIVE POSITIVE
Units area under the curve area under the curve area under the curve area under the curve

Chromatography:

Chromatography ID:CH001363
Chromatography Summary:The LC/MS portion of the platform was based on a Waters ACQUITY ultra-performance liquid chromatography (UPLC) and a Thermo Scientific Q-Exactive high resolution/accurate mass spectrometer interfaced with a heated electrospray ionization (HESI-II) source and Orbitrap mass analyzer operated at 35,000 mass resolution. Each sample extract was first dried, then reconstituted in acidic or basic LC-compatible solvents. To ensure injection and chromatographic consistency, these solvents each contained 8 or more injection standards at fixed concentrations. One aliquot was analyzed using acidic positive ion optimized conditions and the other using basic negative ion optimized conditions in two independent injections using separate dedicated columns (Waters UPLC BEH C18-2.1x100 mm, 1.7 µm). Extracts reconstituted in acidic conditions were gradient eluted from a C18 column using water and methanol containing 0.1% formic acid. The basic extracts were similarly eluted from C18 using methanol and water, however with 6.5mM Ammonium Bicarbonate. The third aliquot was analyzed via negative ionization following elution from a HILIC column (Waters UPLC BEH Amide 2.1x150 mm, 1.7 µm) using a gradient consisting of water and acetonitrile with 10mM Ammonium Formate. The MS analysis alternated between MS and data-dependent MS2 scans using dynamic exclusion, and the scan range was from 80-1000 m/z.
Methods Filename:PROTOCOL_AND_REPORT_150324.docx
Instrument Name:Waters Acquity
Column Name:Waters Acquity BEH C18 (100 x 2mm,1.7um)/Waters UPLC BEH Amide (2.1x150 mm,1.7um)
Solvent A:100% water; 0.1% formic acid
Solvent B: 100% methanol; 0.1% formic acid
Chromatography Type:Reversed phase/HILIC
  
Chromatography ID:CH001364
Chromatography Summary:For GC-MS analysis, samples were dried under vacuum for at least 18 h prior to derivatization under dried nitrogen using bistrimethyl-silyltrifluoroacetamide. Derivatized samples were separated on a 5% diphenyl / 95% dimethyl polysiloxane fused silica column (20 m x 0.18 mm ID; 0.18 um film thickness) with helium as carrier gas and a temperature ramp from 60° to 340°C over a 17.5 min period. Samples were analyzed on a Thermo-Finnigan Trace DSQ fast-scanning single-quadrupole mass spectrometer using electron impact ionization (EI) and operated at unit mass resolving power. The scan range was from 50–750 m/z.
Methods Filename:PROTOCOL_AND_REPORT_150324.docx
Instrument Name:Thermo Trace DSQ
Column Name:5% diphenyl/95% dimethyl polysiloxane GC column
Chromatography Type:GC

MS:

MS ID:MS001738
Analysis ID:AN001882
Instrument Name:Thermo Q Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Reversed phase LC-MS Positive
Ion Mode:POSITIVE
  
MS ID:MS001739
Analysis ID:AN001883
Instrument Name:Thermo Q Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Reversed phase LC-MS Negative
Ion Mode:NEGATIVE
  
MS ID:MS001740
Analysis ID:AN001884
Instrument Name:Thermo Q Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:HILIC LC-MS Negative
Ion Mode:NEGATIVE
  
MS ID:MS001741
Analysis ID:AN001885
Instrument Name:Thermo Trace DSQ
Instrument Type:Single quadrupole
MS Type:EI
MS Comments:GC-MS
Ion Mode:POSITIVE
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