Summary of Study ST001128
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
| Study ID | ST001128 |
| Study Title | WT and ΔSPT cultures of B. thetaiotaomicron grown in Minimal Media with or without d4-alanine (part IV) |
| Study Summary | Lipid profiling was applied on WT and ΔSPT cultures of B. thetaiotaomicron grown in minimal liquid media supplemented with or without deuterium (D4)-labelled alanine, in order to elucidate the production pathways of Bacteroides-derived sphingolipids. |
| Institute | Broad Institute of MIT and Harvard |
| Last Name | Avila-Pacheco |
| First Name | Julian |
| Address | 415 Main Street |
| jravilap@broadinstitute.org | |
| Phone | 617-714-8264 |
| Submit Date | 2019-01-17 |
| Num Groups | 3 |
| Total Subjects | 9 |
| 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: | SU001189 |
| Subject Type: | Bacteria |
| Subject Species: | Bacteroides thetaiotaomicron VPI-5482 |
| Taxonomy ID: | 226186 |
| Genotype Strain: | WT (VPI-5482), SPT K.O. (BT_0870) |
| Cell Biosource Or Supplier: | ATCC |
| Cell Strain Details: | Wild type Bacteroides tethaiotamicron strains and serine palmitoyltransferase (SPT) knockouts |
| Subject Comments: | Grown in Minimal Media with or without d4-alanine |
Factors:
Subject type: Bacteria; Subject species: Bacteroides thetaiotaomicron VPI-5482 (Factor headings shown in green)
| mb_sample_id | local_sample_id | Strain |
|---|---|---|
| SA078068 | SPT3 | SPT K.O. (BT_0870) + d4-Ala |
| SA078069 | SPT1 | SPT K.O. (BT_0870) + d4-Ala |
| SA078070 | SPT2 | SPT K.O. (BT_0870) + d4-Ala |
| SA078071 | MM2 | WT (VPI-5482) |
| SA078072 | MM1 | WT (VPI-5482) |
| SA078073 | MM3 | WT (VPI-5482) |
| SA078074 | WT3 | WT (VPI-5482) + d4-Ala |
| SA078075 | WT1 | WT (VPI-5482) + d4-Ala |
| SA078076 | WT2 | WT (VPI-5482) + d4-Ala |
| Showing results 1 to 9 of 9 |
Collection:
| Collection ID: | CO001183 |
| Collection Summary: | After 48 hrs of labeling, resulting cultures were pelleted by centrifugation at 8000 rpm for 10 min and cell density was normalized. Lipids were extracted using isopropanol and stored at -80C for lipidomic analysis. |
| Sample Type: | Bacterial cells |
| Storage Conditions: | -80℃ |
Treatment:
| Treatment ID: | TR001204 |
| Treatment Summary: | WT and ΔSPT cultures of B. thetaiotaomicron and B. ovatus were grown in 5 mL of minimal media (M9 salts (Teknova), 1% vitamin K1-hemin solution (Becton Dickinson), 1% trace mineral supplement (ATCC), 1% trace vitamin supplement (ATCC), 2% lactose), supplemented with 10μM of deuterium (D4)-labelled alanine (Sigma), or 10μM of -alanine (Sigma). |
| Cell Storage: | -80C |
| Cell Growth Container: | anaerobic chamber (Coy Laboratory Products) with an atmosphere of 20% CO2, 5% H2, and 75% N2 at 37°C. |
Sample Preparation:
| Sampleprep ID: | SP001197 |
| Sampleprep Summary: | After 48 hrs, resulting cultures were pelleted by centrifugation at 8000 rpm for 10 min and cell density was normalized. Lipids were extracted using isopropanol and stored at -80C until ready for lipidomic analysis. |
Combined analysis:
| Analysis ID | AN001853 |
|---|---|
| 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 | Abundances |
Chromatography:
| Chromatography ID: | CH001341 |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | Waters Acquity BEH C8 (100 x 2.1mm,1.7um) |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS001714 |
| Analysis ID: | AN001853 |
| 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 |
