Summary of Study ST002322
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 PR001488. The data can be accessed directly via it's Project DOI: 10.21228/M8CT5G This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST002322 |
| Study Title | Metabolomics study comparing SCAP KO and WT B cells |
| Study Type | Purified mouse B cells, stimulated ex vivo |
| Study Summary | Splenic B cells isolated from SCAP+/+ CD19-Cre (WT) and SCAPfl/fl CD19-Cre (KO) mice were stimulated with LPS or anti-CD40 for 24 and 48 hours. Cells were then analyzed by metabolomics. Metabolomics reveals global metabolic changes in SCAP deficient B cells. |
| Institute | Indiana University School of Medicine |
| Last Name | Luo |
| First Name | Wei |
| Address | 950 W Walnut Street - R2 E304 |
| wl47@iu.edu | |
| Phone | 3172748042 |
| Submit Date | 2022-10-19 |
| Analysis Type Detail | LC-MS |
| Release Date | 2022-11-18 |
| Release Version | 1 |
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Project:
| Project ID: | PR001488 |
| Project DOI: | doi: 10.21228/M8CT5G |
| Project Title: | Metabolomics reveals global metabolic changes between WT and SCAP deficient B cells |
| Project Type: | Purified mouse B cells, stimulated in vitro |
| Project Summary: | Cellular metabolism regulates almost all critical cellular activities including cell growth, proliferation, energy homeostasis and signaling transduction. SCAP controls SREBP signaling, an important pathway regulating cellular lipid biosynthesis. SCAP KO B cells have multiple defects and cannot form germinal centers. To test whether these defects of SCAP KO B cells are associated with an altered global metabolic state. We stimulated B cells isolated from SCAPfl/fl CD19Cre/+ mice (KO) and SCAP+/+ CD19Cre/+ mice (WT) with ⍺-CD40 or LPS for 24 and 48 hours, followed by targeted metabolomics analysis to evaluate global metabolic changes. We found that the metabolic profiles of unstimulated SCAP KO and WT B cells were largely overlapping, corroborating our findings that SCAP deficiency did not affect B cell development or their metabolic profile at steady state. In contrast, after stimulation with either LPS or ⍺-CD40, SCAP KO and WT B cells were clearly separated at 24 hours, with an even more pronounced separation at 48 hours. SREBP signaling can regulated the metabolism of many ceramides and sphingolipids. For example, lactosyl-N-palmitoyl-sphingosine (d18:1/16:0), one of the lactosylceramides (LacCer), is highly accumulated in SCAP KO B cells activated by either ⍺-CD40 or LPS. Strikingly, besides lipid metabolism, SCAP deficiency also alters many other metabolites that belong to different metabolic pathways such as energy, amino acids, peptide, cofactors and vitamins, nucleotide, carbohydrate, and xenobiotics. Taken together, the metabolomics analysis revealed previously unrecognized roles of SREBP signaling in regulating multiple cellular activities in activated B cells. |
| Institute: | Indiana University |
| Last Name: | Luo |
| First Name: | Wei |
| Address: | 950 W Walnut Street - R2 E304, Indianapolis, IN, 46202, USA |
| Email: | wl47@iu.edu |
| Phone: | 3172748042 |
Subject:
| Subject ID: | SU002408 |
| Subject Type: | Cultured cells |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | SCAP fl/fl CD19-Cre (KO) and SCAP+/+ CD19-Cre (WT) |
| Cell Strain Details: | Splenic B cell |
| Species Group: | Mammals |
Factors:
Subject type: Cultured cells; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Genotype | TREATMENT | TIME_POINT |
|---|---|---|---|---|
| SA227408 | KO_CD40_24hr_4 | SCAP KO | anti-CD40 | 24hr |
| SA227409 | KO_CD40_24hr_2 | SCAP KO | anti-CD40 | 24hr |
| SA227410 | KO_CD40_24hr_3 | SCAP KO | anti-CD40 | 24hr |
| SA227411 | KO_CD40_24hr_1 | SCAP KO | anti-CD40 | 24hr |
| SA227412 | KO_CD40_48hr_1 | SCAP KO | anti-CD40 | 48hr |
| SA227413 | KO_CD40_48hr_2 | SCAP KO | anti-CD40 | 48hr |
| SA227414 | KO_CD40_48hr_3 | SCAP KO | anti-CD40 | 48hr |
| SA227415 | KO_CD40_48hr_4 | SCAP KO | anti-CD40 | 48hr |
| SA227396 | KO_0hr_2 | SCAP KO | Freshly isolated (No treatment) | 0hr |
| SA227397 | KO_0hr_3 | SCAP KO | Freshly isolated (No treatment) | 0hr |
| SA227398 | KO_0hr_1 | SCAP KO | Freshly isolated (No treatment) | 0hr |
| SA227399 | KO_0hr_4 | SCAP KO | Freshly isolated (No treatment) | 0hr |
| SA227400 | KO_LPS_24hr_3 | SCAP KO | LPS | 24hr |
| SA227401 | KO_LPS_24hr_2 | SCAP KO | LPS | 24hr |
| SA227402 | KO_LPS_24hr_4 | SCAP KO | LPS | 24hr |
| SA227403 | KO_LPS_24hr_1 | SCAP KO | LPS | 24hr |
| SA227404 | KO_LPS_48hr_3 | SCAP KO | LPS | 48hr |
| SA227405 | KO_LPS_48hr_2 | SCAP KO | LPS | 48hr |
| SA227406 | KO_LPS_48hr_4 | SCAP KO | LPS | 48hr |
| SA227407 | KO_LPS_48hr_1 | SCAP KO | LPS | 48hr |
| SA227428 | WT_CD40_24hr_2 | WT | anti-CD40 | 24hr |
| SA227429 | WT_CD40_24hr_1 | WT | anti-CD40 | 24hr |
| SA227430 | WT_CD40_24hr_4 | WT | anti-CD40 | 24hr |
| SA227431 | WT_CD40_24hr_3 | WT | anti-CD40 | 24hr |
| SA227432 | WT_CD40_48hr_2 | WT | anti-CD40 | 48hr |
| SA227433 | WT_CD40_48hr_1 | WT | anti-CD40 | 48hr |
| SA227434 | WT_CD40_48hr_3 | WT | anti-CD40 | 48hr |
| SA227435 | WT_CD40_48hr_4 | WT | anti-CD40 | 48hr |
| SA227416 | WT_0hr_4 | WT | Freshly isolated (No treatment) | 0hr |
| SA227417 | WT_0hr_3 | WT | Freshly isolated (No treatment) | 0hr |
| SA227418 | WT_0hr_2 | WT | Freshly isolated (No treatment) | 0hr |
| SA227419 | WT_0hr_1 | WT | Freshly isolated (No treatment) | 0hr |
| SA227420 | WT_LPS_24hr_1 | WT | LPS | 24hr |
| SA227421 | WT_LPS_24hr_2 | WT | LPS | 24hr |
| SA227422 | WT_LPS_24hr_4 | WT | LPS | 24hr |
| SA227423 | WT_LPS_24hr_3 | WT | LPS | 24hr |
| SA227424 | WT_LPS_48hr_4 | WT | LPS | 48hr |
| SA227425 | WT_LPS_48hr_1 | WT | LPS | 48hr |
| SA227426 | WT_LPS_48hr_2 | WT | LPS | 48hr |
| SA227427 | WT_LPS_48hr_3 | WT | LPS | 48hr |
| Showing results 1 to 40 of 40 |
Collection:
| Collection ID: | CO002401 |
| Collection Summary: | Mouse splenic B cells were isolated using STEM CELL B cell isolation kit (Cat:19854) |
| Sample Type: | B cells |
Treatment:
| Treatment ID: | TR002420 |
| Treatment Summary: | Splenic B cells isolated from SCAP+/+ CD19-Cre (WT) and SCAPfl/fl CD19-Cre (KO) mice were stimulated with 10 micro g/ml LPS or 5 micro g/ml anti-CD40 for 24 and 48 hours. Freshly isolated untreated WT and KO B cells were used as untreated control. |
Sample Preparation:
| Sampleprep ID: | SP002414 |
| Sampleprep Summary: | Sample preparation: Proteins were precipitated with methanol under vigorous shaking for 2 min (Glen Mills GenoGrinder 2000) followed by centrifugation. The resulting extract was divided into five fractions: two for analysis by two separate reverse phases (RP)/UPLC-MS/MS methods with positive ion mode electrospray ionization (ESI), one for analysis by RP/UPLC-MS/MS with negative ion mode ESI, one for analysis by HILIC/UPLC-MS/MS with negative ion mode ESI, and one sample was reserved for backup. Samples were placed briefly on a TurboVap® (Zymark) to remove the organic solvent. The sample extracts were stored overnight under nitrogen before preparation for analysis. |
Chromatography:
| Chromatography ID: | CH002802 |
| Methods Filename: | B_cell_MS_protocol.docx |
| Instrument Name: | Waters ACQUITY ultra-performance liquid chromatography (UPLC) |
| Column Name: | Waters UPLC BEH C18 (100 x 2.1mm,1.7um)/Waters Acquity BEH Amide (150 x 2.1mm,1.7um) |
| Chromatography Type: | Reversed phase/HILIC |
Analysis:
| Analysis ID: | AN003789 |
| Analysis Type: | MS |
| Analysis Protocol File: | B_cell_MS_protocol.docx |
| Chromatography ID: | CH002802 |
| Num Factors: | 10 |
| Num Metabolites: | 404 |
| Units: | Normalized AUC |
