Summary of Study ST002846
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 PR001781. The data can be accessed directly via it's Project DOI: 10.21228/M8H432 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 | ST002846 |
| Study Title | Apolipoprotein E suppresses hyperlipidemia-driven hematopoiesis & inflammation by controlling mitochondrial metabolism |
| Study Summary | Apolipoprotein E (ApoE) is recognized for its pleiotropic properties that suppress inflammation. We report that ApoE serves as a metabolic rheostat that regulates microRNA-control of glycolytic and mitochondrial activity in myeloid cells and hematopoietic stem & progenitor cells (HSPCs). ApoE expression in myeloid cells increases microRNA-146a, which reduces NF-κB-driven GLUT1 expression and glycolytic activity. In contrast, ApoE expression reduces microRNA-142a, which increases CPT1A expression, fatty acid oxidation, and oxidative phosphorylation. Improved mitochondrial metabolism by ApoE expression causes an enrichment of TCA cycle metabolites and NAD+ in macrophages. The study of mice with conditional ApoE expression supports the capacity for ApoE to foster microRNA-controlled immunometabolism. Modulation of microRNA-146a & -142a in the hematopoietic system of hyperlipidemic mice using RNA mimics & antagonists, respectively, improves mitochondrial metabolism, which suppresses inflammation and hematopoiesis. Our findings unveil an RNA regulatory network, controlled by ApoE, which exerts metabolic control over hematopoiesis and inflammation in hyperlipidemia. |
| Institute | Northwestern University |
| Last Name | Stoolman |
| First Name | Joshua |
| Address | 303 E Superior Street, Chicago, IL 60611 |
| joshua.stoolman@northwestern.edu | |
| Phone | 7343559440 |
| Submit Date | 2023-07-20 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2024-05-01 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001781 |
| Project DOI: | doi: 10.21228/M8H432 |
| Project Title: | Apolipoprotein E suppresses hyperlipidemia-driven hematopoiesis & inflammation by controlling mitochondrial metabolism |
| Project Summary: | Apolipoprotein E (ApoE) is recognized for its pleiotropic properties that suppress inflammation. We report that ApoE serves as a metabolic rheostat that regulates microRNA-control of glycolytic and mitochondrial activity in myeloid cells and hematopoietic stem & progenitor cells (HSPCs). ApoE expression in myeloid cells increases microRNA-146a, which reduces NF-?B-driven GLUT1 expression and glycolytic activity. In contrast, ApoE expression reduces microRNA-142a, which increases CPT1A expression, fatty acid oxidation, and oxidative phosphorylation. Improved mitochondrial metabolism by ApoE expression causes an enrichment of TCA cycle metabolites and NAD+ in macrophages. The study of mice with conditional ApoE expression supports the capacity for ApoE to foster microRNA-controlled immunometabolism. Modulation of microRNA-146a & -142a in the hematopoietic system of hyperlipidemic mice using RNA mimics & antagonists, respectively, improves mitochondrial metabolism, which suppresses inflammation and hematopoiesis. Our findings unveil an RNA regulatory network, controlled by ApoE, which exerts metabolic control over hematopoiesis and inflammation in hyperlipidemia. |
| Institute: | Northwestern University |
| Department: | Pulmonary and Critical Care |
| Laboratory: | Chandel Laboratory |
| Last Name: | Stoolman |
| First Name: | Joshua |
| Address: | 303 E Superior Street, Chicago, IL, 60611 |
| Email: | joshua.stoolman@northwestern.edu |
| Phone: | 7343559440 |
Subject:
| Subject ID: | SU002958 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | genotype | treatment |
|---|---|---|---|
| SA308535 | 17 | ApoE-KO | no treatment |
| SA308536 | 18 | ApoE-KO | no treatment |
| SA308537 | 20 | ApoE-KO | no treatment |
| SA308538 | 19 | ApoE-KO | no treatment |
| SA308539 | 8 | wild-type | IL-4 |
| SA308540 | 7 | wild-type | IL-4 |
| SA308541 | 6 | wild-type | IL-4 |
| SA308542 | 5 | wild-type | IL-4 |
| SA308543 | 2 | wild-type | no treatment |
| SA308544 | 3 | wild-type | no treatment |
| SA308545 | 4 | wild-type | no treatment |
| SA308546 | 1 | wild-type | no treatment |
| Showing results 1 to 12 of 12 |
Collection:
| Collection ID: | CO002951 |
| Collection Summary: | BMDMs were seeded in a 6-well cell culture plate at a density of 50,000/well, and incubated in complete media at 37C & 5% CO2 overnight. Cells were gently washed 4x with DPBS before freezing at -80C. For metabolite extraction, cells were extracted with 1 mL HPLC-grade methanol/water 80:20. Cell lysates were centrifuged at 18,000 x g for 10 min. Supernatants were collected for downstream metabolomic analysis. The samples were evaporated using a stream of nitrogen. Subsequently metabolites were reconstituted in 50% acetonitrile in HPLC-grade water,vortexed and centrifuged to remove any debris. Samples were analyzed by Ultra-High-Performance Liquid Chromatography and High-Resolution Mass Spectrometry and Tandem Mass Spectrometry (UHPLC-MS/MS). Specifically, the system consisted of a Thermo Q-Exactive in line with an electrospray source and an Ultimate3000 (Thermo Fisher, USA) series HPLC consisting of a binary pump, degasser, and auto-sampler outfitted with an Xbridge Amide column (Waters; dimensions of 4.6mm × 100mm and a 3.5μm particle size). Mobile phase A contained 95% (vol/vol) water, 5% (vol/vol) acetonitrile, 10mM ammonium hydroxide, 10mM ammonium acetate, pH = 9.0; and mobile phase B was 100% Acetonitrile. The gradient was as follows: 0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1-18 min, 75% A; 18-25 min, 15% A with a flow rate of 400μL/min. The capillary of the ESI source was set to 275°C, with sheath gas at 45 arbitrary units, auxiliary gas at 5 arbitrary units, and the spray voltage at 4.0kV. In positive/negative polarity switching mode, an m/z scan automatic gain control (AGC) target was set at 1x106 and the maximum injection time was 200 ms. The top 5 precursor ions were subsequently fragmented, in a data-dependent manner, using the higher energy collisional dissociation (HCD) cell set to 30% normalized collision energy in MS2 at a resolution power of 17,500. |
| Sample Type: | Macrophages |
| Storage Conditions: | -80℃ |
Treatment:
| Treatment ID: | TR002967 |
| Treatment Summary: | Briefly, bone marrow cells were flushed from the tibia and femurs of 6- to 12-week-old male Apoe-/-, Apoe+/-, or wildtype mice on C57BL/6J background. Cells were cultured in complete media containing DMEM (Corning, USA) supplemented with 10% fetal bovine serum (GIBCO, USA), 1% GlutaMax (GIBCO, USA), and 1% penicillin-streptomycin (GIBCO, USA) and differentiated with 25 ng/ml mouse M-CSF (Peprotech, USA) for 6 days in 37C and 5% CO2. BMDM were then seeded into 12-well culture plates (Corning, USA) at a concentration of 3 x 10^5. Stimulation was done with 20 ng/mL IL-4 for 16 hours prior to collection. |
Sample Preparation:
| Sampleprep ID: | SP002964 |
| Sampleprep Summary: | Following incubation/ stimulation cells were washed with 2mL ice-cold 0/9% NaCl. Following plate wash, plates were placed on dry ice 100uL 80:20 methanol/water solution in dry ice. Plates were scraped on dry ice and lysates were transferred to eppendorf tubes and centrifuged at 21,000xg for 10 min. Supernatants were transferred to fresh tubes and kept at -80dC until analysis. |
Combined analysis:
| Analysis ID | AN004664 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Thermo Fisher UltiMate 3000 |
| Column | Waters XBridge Amide (100 x 4.6mm, 3.5um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive Plus Orbitrap |
| Ion Mode | UNSPECIFIED |
| Units | Peak area |
Chromatography:
| Chromatography ID: | CH003510 |
| Chromatography Summary: | Samples were analyzed by high-performance LC (HPLC) and high-resolution MS and MS/MS (HPLC-MS/MS). The system consists of Thermo Q Exactive with an electrospray source and an UltiMate3000 (Thermo Fisher Scientific) series HPLC consisting of a binary pump, degasser, and autosampler outfitted with an XBridge Amide column (Waters; dimensions of 4.6 mm by 100 mm and a 3.5-μm particle size). Mobile phase A contained water and acetonitrile (95/5, v/v), 10 mM ammonium hydroxide and 10 mM ammonium acetate (pH 9.0). Mobile phase B was 100% acetonitrile. The gradient was set to 0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1–18 min, 75% A; 18–25 min, 15% A, with a flow rate of 400 μl min–1. |
| Instrument Name: | Thermo Fisher UltiMate 3000 |
| Column Name: | Waters XBridge Amide (100 x 4.6mm, 3.5um) |
| Column Temperature: | 275 |
| Flow Gradient: | 0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1 to 18 min, 75% A; and 18 to 25 min, 15% A |
| Flow Rate: | 400 μl/min |
| Solvent A: | 95% water/5% acetonitrile; 10 mM ammonium hydroxide; 10 mM ammonium acetate (pH 9.0) |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS004411 |
| Analysis ID: | AN004664 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | A 15μl aliquot of the sample was used for high-resolution HPLC-tandem mass spectrometry. High-resolution HPLC-tandem mass spectrometry was performed on a Q-Exactive (ThermoFisher Scientific) in line with an electrospray source and an UltiMate 3000 (ThermoFisher Scientific) series HPLC consisting of a binary pump, degasser and autosampler outfitted with a XBridge Amide column (Waters; 4.6 mm × 100 mm dimension and a 3.5 μm particle size). The capillary of the electrospray ionization source was set to 275 °C, with sheath gas at 45 arbitrary units, auxiliary gas at 5 arbitrary units and the spray voltage at 4.0 kV. A mass/charge ratio scan ranging from 70 to 850 was used in positive/negative polarity switching mode. MS1 data were collected at a resolution The automatic gain control (AGC) target was set at 1 × 106, with a maximum injection time of 200 ms. The top five precursor ions were fragmented using the higher-energy collisional dissociation cell with normalized collision energy of 30% in MS2 at a resolution of 17,500. Data were acquired with Xcalibur software (v.4.1; ThermoFisher Scientific). Analysis was performed using MeatboAnalyst (v5.0) with peak area normalized to total ion count for each sample. |
| Ion Mode: | UNSPECIFIED |
