Summary of Study ST001127

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 PR000754. The data can be accessed directly via it's Project DOI: 10.21228/M8709G This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST001127
Study Title	Lipid profiling of caecal samples from GF mice colonized with B. thetaiotaomicron WT or the ΔSPT mutants (part III)
Study Summary	To study Bacteroidetes sphingolipids in intestinal health, we colonized germ-free mice with wild type or a sphingolipid-deficient Bacteroides thetaiotaomicron strain. A lack of Bacteroides derived sphingolipids increased intestinal inflammation, dysregulated innate immunity and altered the host ceramide pool.
Institute	Broad Institute of MIT and Harvard
Last Name	Avila-Pacheco
First Name	Julian
Address	415 Main Street
Email	jravilap@broadinstitute.org
Phone	617-714-8264
Submit Date	2019-01-17
Num Groups	3
Total Subjects	14
Raw Data Available	Yes
Raw Data File Type(s)	raw(Thermo)
Analysis Type Detail	LC-MS
Release Date	2019-03-06
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR000754
Project DOI:	doi: 10.21228/M8709G
Project Title:	Bacteroides-derived sphingolipids are critical for maintaining intestinal homeostasis and symbiosis
Project Summary:	Sphingolipids are structural membrane components and important eukaryotic signaling molecules. We hypothesized that sphingolipids mediate intestinal health as they were identified as the most upregulated metabolite feature in stool of inflammatory bowel disease (IBD) patients. Commensal Bacteroidetes also produce sphingolipids, but the impact of these metabolites on host pathways is largely uncharacterized. To study Bacteroidetes sphingolipids in intestinal health, we colonized germ-free mice with a sphingolipid-deficient Bacteroides thetaiotaomicron strain. A lack of Bacteroides-derived sphingolipids increased intestinal inflammation, dysregulated innate immunity and altered the host ceramide pool. Using metabolomic analysis, we described the Bacteroides sphingolipid biosynthesis pathway and revealed a greater variety of Bacteroides-derived sphingolipids than previously recognized, including ceramide phosphoinositol and deoxy-sphingolipids. We annotated Bacteroides sphingolipids in an IBD metabolomic dataset, discovering lower abundances in IBD and negative correlations with gut inflammation and host sphingolipid production. These data highlight the role of sphingolipids in maintaining host-bacterial symbiosis and intestinal homeostasis.
Institute:	Broad Institute of MIT and Harvard
Last Name:	Avila-Pacheco
First Name:	Julian
Address:	415 Main Street, Cambridge MA
Email:	jravilap@broadinstitute.org
Phone:	617-714-8264
Funding Source:	National Institutes of Health (P30 DK043351 and R01 AT009708)
Contributors:	Eric M. Brown, Xiaobo Ke, Daniel Hitchcock, Timothy D. Arthur, Toru Nakata, Nadine Fornelos, Cortney Heim, Eric A. Franzosa1,4, Curtis Huttenhower1,4, Henry J. Haiser3, Glen 6 Dillow3, Daniel B. Graham1, B. Brett Finlay, Aleksandar D. Kostic, Jeffrey A. Porter, Hera Vlamakis, Sarah Jeanfavre, Julian Avila-Pacheco, Clary B. Clish, and Ramnik J. Xavier

Subject:

Subject ID:	SU001188
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090
Genotype Strain:	C57BL/6 germ-free mice
Animal Animal Supplier:	Taconic

Factors:

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

mb_sample_id	local_sample_id	Inoculum	Strain
SA078054	BTSPR Cecal_3	Bacteroides thetaiotaomicron	SPT K.O. (BT_0870)
SA078055	BTSPR Cecal_4	Bacteroides thetaiotaomicron	SPT K.O. (BT_0870)
SA078056	BTSPR Cecal_6	Bacteroides thetaiotaomicron	SPT K.O. (BT_0870)
SA078057	BTSPR Cecal_1	Bacteroides thetaiotaomicron	SPT K.O. (BT_0870)
SA078058	BTSPR Cecal_5	Bacteroides thetaiotaomicron	SPT K.O. (BT_0870)
SA078059	BTSPR Cecal_2	Bacteroides thetaiotaomicron	SPT K.O. (BT_0870)
SA078060	BTWT Cecal_3	Bacteroides thetaiotaomicron	WT (VPI-5482)
SA078061	BTWT Cecal_2	Bacteroides thetaiotaomicron	WT (VPI-5482)
SA078062	BTWT Cecal_1	Bacteroides thetaiotaomicron	WT (VPI-5482)
SA078063	BTWT Cecal_4	Bacteroides thetaiotaomicron	WT (VPI-5482)
SA078064	BTWT Cecal_5	Bacteroides thetaiotaomicron	WT (VPI-5482)
SA078065	BTWT Cecal_6	Bacteroides thetaiotaomicron	WT (VPI-5482)
SA078066	GF Cecal_1	None	None
SA078067	GF Cecal_2	None	None

Showing results 1 to 14 of 14

Showing results 1 to 14 of 14

Collection:

Collection ID:	CO001182
Collection Summary:	Caecal samples from GF mice colonized with B. thetaiotaomicron WT or the ΔSPT mutant were weighed, and 20 mg of each wet-weight sample was used for lipid profiling.
Sample Type:	Cecum
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR001203
Treatment Summary:	In an anaerobic chamber, bacterial cultures from frozen stocks were first plated on BHI supplemented with hemin and vitamin K, and subsequently pure cultures were selected and mixed together at a 1:1 ratio in sterile, reduced PBS. Bacterial mixtures in PBS were removed from the anaerobic chamber and immediately transported to the animal facility for gavage experiments. The volume of the mixture received per mouse was 100 μL, at a concentration of 10^9 cells/mL. The concentration of the mixture in cell/mL was determined using a UV spectrometer, and gavage doses were confirmed by back-titering the inocula.

Treatment ID:

TR001203

Treatment Summary:

In an anaerobic chamber, bacterial cultures from frozen stocks were first plated on BHI supplemented with hemin and vitamin K, and subsequently pure cultures were selected and mixed together at a 1:1 ratio in sterile, reduced PBS. Bacterial mixtures in PBS were removed from the anaerobic chamber and immediately transported to the animal facility for gavage experiments. The volume of the mixture received per mouse was 100 μL, at a concentration of 10^9 cells/mL. The concentration of the mixture in cell/mL was determined using a UV spectrometer, and gavage doses were confirmed by back-titering the inocula.

Sample Preparation:

Sampleprep ID:	SP001196
Sampleprep Summary:	20 mg of each wet-weight cecal sample was extracted using 200 μL isopropanol + 0.1 ng/μL C24:0 PC as internal standard, incubated at RT for 1 hr, and centrifuged for 10 min at 10,000g.

Combined analysis:

Analysis ID	AN001852
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Shimadzu Nexera X2
Column	Waters Acquity BEH C8 (100 x 2.1mm,1.7um)
MS Type	ESI
MS instrument type	Orbitrap
MS instrument name	Thermo Q Exactive Focus
Ion Mode	POSITIVE
Units	abundance

Chromatography:

Chromatography ID:	CH001340
Instrument Name:	Shimadzu Nexera X2
Column Name:	Waters Acquity BEH C8 (100 x 2.1mm,1.7um)
Chromatography Type:	Reversed phase

MS:

MS ID:	MS001713
Analysis ID:	AN001852
Instrument Name:	Thermo Q Exactive Focus
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Raw data were processed using TraceFinder 3.1 (Thermo Fisher) and Progenesis QI (Nonlinear dynamics). Retention time, m/z, MS/MS fragmentation were used to confirm the identity of metabolite using authentic reference standards when available (level 1 identifications), in addition to monitoring the product ions produced and the retention time and mass of species belonging to the each lipid class (level 2-3 annotation).
Ion Mode:	POSITIVE