Summary of Study ST002451
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 PR001580. The data can be accessed directly via it's Project DOI: 10.21228/M8G711 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 | ST002451 |
| Study Title | APOE modulates microglial immunometabolism in response to age, amyloid pathology, and inflammatory challenge (Part 2 of 3) |
| Study Summary | The E4 allele of Apolipoprotein E (APOE) is associated with both metabolic dysfunction and a heightened pro-inflammatory response – two findings that may be intrinsically linked through the concept of immunometabolism. Here, we combined bulk, single-cell, and spatial transcriptomics with cell-specific and spatially resolved metabolic analyses to systematically address the role of APOE across age, neuroinflammation, and AD pathology. RNAseq highlighted immunometabolic changes across the APOE4 glial transcriptome, specifically in subsets of metabolically distinct microglia enriched in the E4 brain during aging or following an inflammatory challenge. E4 microglia display increased Hif1α expression, a disrupted TCA cycle, and are inherently pro-glycolytic, while spatial transcriptomics and MALDI mass spectrometry imaging highlight an E4-specific response to amyloid that is characterized by widespread alterations in lipid metabolism. Taken together, our findings emphasize a central role for APOE in regulating microglial immunometabolism. |
| Institute | University of Kentucky |
| Department | Physiology |
| Laboratory | Lance Johnson; Josh Morganti |
| Last Name | Devanney |
| First Name | Nicholas |
| Address | Physiology, 760 Press Ave, Healthy Kentucky Research Bldg, Rm152, Lexington, Kentucky, 40508, USA |
| Nicholas.Devanney@uky.edu | |
| Phone | 8593238083 |
| Submit Date | 2023-01-20 |
| Study Comments | Part 2 of 3 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | cdf |
| Analysis Type Detail | GC-MS |
| Release Date | 2023-01-25 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001580 |
| Project DOI: | doi: 10.21228/M8G711 |
| Project Title: | APOE modulates microglial immunometabolism in response to age, amyloid pathology, and inflammatory challenge |
| Project Summary: | The E4 allele of Apolipoprotein E (APOE) is associated with both metabolic dysfunction and a heightened pro-inflammatory response – two findings that may be intrinsically linked through the concept of immunometabolism. Here, we combined bulk, single-cell, and spatial transcriptomics with cell-specific and spatially resolved metabolic analyses to systematically address the role of APOE across age, neuroinflammation, and AD pathology. RNAseq highlighted immunometabolic changes across the APOE4 glial transcriptome, specifically in subsets of metabolically distinct microglia enriched in the E4 brain during aging or following an inflammatory challenge. E4 microglia display increased Hif1α expression, a disrupted TCA cycle, and are inherently pro-glycolytic, while spatial transcriptomics and MALDI mass spectrometry imaging highlight an E4-specific response to amyloid that is characterized by widespread alterations in lipid metabolism. Taken together, our findings emphasize a central role for APOE in regulating microglial immunometabolism. |
| Institute: | University of Kentucky, Department of Physiology |
| Last Name: | Devanney |
| First Name: | Nicholas |
| Address: | Physiology, 760 Press Ave, Healthy Kentucky Research Bldg, Rm152, Lexington, Kentucky, 40508, USA |
| Email: | Nicholas.Devanney@uky.edu |
| Phone: | 8593238083 |
| Project Comments: | Part 2 of 3 |
Subject:
| Subject ID: | SU002540 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | APOE-targeted replacement mice homozygous for human E3 (B6.129P2-Apoe^tm2(APOE*3)Mae N8, Taconic #1548-F) or human E4 (B6.129P2- Apoe^tm3(APOE*4)Mae N8, Taconic #1549-F) alleles |
| Age Or Age Range: | P0-P3 |
| Gender: | Pooled |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Group | APOE genotype | Treatment |
|---|---|---|---|---|
| SA245058 | 47_3_12 | E3_control | E3/E3 | Control |
| SA245059 | 47_3_1 | E3_control | E3/E3 | Control |
| SA245060 | 47_3_11 | E3_control | E3/E3 | Control |
| SA245061 | 47_3_10 | E3_control | E3/E3 | Control |
| SA245062 | 47_3_2 | E3_control | E3/E3 | Control |
| SA245063 | 47_3_4 | E3_control | E3/E3 | Control |
| SA245064 | 47_3_3 | E3_control | E3/E3 | Control |
| SA245065 | 47_3_9 | E3_control | E3/E3 | Control |
| SA245066 | 47_3_14 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245067 | 47_3_15 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245068 | 47_3_13 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245069 | 47_3_16 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245070 | 47_3_8 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245071 | 47_3_5 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245072 | 47_3_6 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245073 | 47_3_7 | E3_proinflammatory | E3/E3 | Proinflammatory |
| SA245074 | 47_4_12 | E4_control | E4/E4 | Control |
| SA245075 | 47_4_9 | E4_control | E4/E4 | Control |
| SA245076 | 47_4_10 | E4_control | E4/E4 | Control |
| SA245077 | 47_4_4 | E4_control | E4/E4 | Control |
| SA245078 | 47_4_2 | E4_control | E4/E4 | Control |
| SA245079 | 47_4_1 | E4_control | E4/E4 | Control |
| SA245080 | 47_4_3 | E4_control | E4/E4 | Control |
| SA245081 | 47_4_15 | E4_proinflammatory | E4/E4 | Proinflammatory |
| SA245082 | 47_4_16 | E4_proinflammatory | E4/E4 | Proinflammatory |
| SA245083 | 47_4_14 | E4_proinflammatory | E4/E4 | Proinflammatory |
| SA245084 | 47_4_8 | E4_proinflammatory | E4/E4 | Proinflammatory |
| SA245085 | 47_4_6 | E4_proinflammatory | E4/E4 | Proinflammatory |
| SA245086 | 47_4_7 | E4_proinflammatory | E4/E4 | Proinflammatory |
| SA245087 | 47_4_13 | E4_proinflammatory | E4/E4 | Proinflammatory |
| Showing results 1 to 30 of 30 |
Collection:
| Collection ID: | CO002533 |
| Collection Summary: | Primary microglia were plated at 7x10^6 cells/well in 6-well plates (VWR #10062-894) and incubated at 5% CO2 37°C. Upon reaching confluence, cells were washed with warm, sterile phosphate-buffered saline (PBS; Thomas #QZY-11666789001-4L) to remove traces of non-13C media and then incubated in glucose- and sodium pyruvate-free DMEM (Thermo #11966-025) containing 2mM GlutaMAX (Thermo #35050-061), 1% penicillin/streptomycin, and 10mM universally labelled 13C-glucose (Cambridge Isotope Laboratories # CLM-1396-PK) for two hours with either a pro-inflammatory cocktail of 20ng/ml interferon-γ (IFNγ, R&D Systems #485-MI-100) and 50ng/ml tumor necrosis factor α (TNFα, R&D Systems #410-MT-025) or vehicle control. Cells were then removed from the incubator and washed with warm 0.9% NaCl solution. Culture plates were placed on a bed of crushed dry ice and 1mL of ice cold 50% methanol (HPLC-grade, Sigma #A456-4) was added to quench cellular metabolic activity followed by a 10 minute incubation at -80°C to ensure cell lysis. After removing from the freezer, cells were detached with a cell scraper (VWR #10062-906) and the entire contents collected into a microcentrifuge tube, vortexed briefly, and placed on ice until all samples were collected. The tubes were then placed on a Disruptor Genie Cell Disruptor Homogenizer (Scientific Industries) for 5 min at 3,000 rpm. Tubes were then centrifuged at 20,000 x g for 10 min at 4 °C. The supernatant containing polar metabolites was isolated to a new tube and stored at -80C until use. |
| Sample Type: | Brain |
Treatment:
| Treatment ID: | TR002552 |
| Treatment Summary: | APOE3/3 and APOE4/4 primary microglia were treated for 2 hours with a pro-inflammatory cocktail of 20ng/ml interferon-γ (IFNγ, R&D Systems #485-MI-100) and 50ng/ml tumor necrosis factor α (TNFα, R&D Systems #410-MT-025) or vehicle control. |
Sample Preparation:
| Sampleprep ID: | SP002546 |
| Sampleprep Summary: | The supernatant fraction containing polar metabolites was thawed gently on ice and dried at 10-3 mbar using a CentriVap vacuum concentrator (LabConco) to evaporate methanol. The dried polar metabolite pellet was derivatized by a two-step methoxyamine protocol first by addition of 70µL methoxyamine HCl (Sigma-Aldrich #226904-5G) in pyridine (20 mg/mL; Sigma-Aldrich #TS25730) to each pellet followed by 90 min dry heat incubation at 30°C. Samples were then centrifuged at 20,000 x g for 10 minutes after which 50µL of each sample was transferred to an amber V-shaped glass chromatography vial (Agilent #5184-3554) containing 80µL N-methyl-trimethylsilyl-trifluoroacetamide (MSTFA; ThermoFisher #TS48915) and gently vortexed followed by 30 min dry heat incubation at 37°C. |
| Extraction Method: | 50% ice-cold methanol |
| Extract Storage: | -80℃ |
| Sample Derivatization: | MSTFA |
Combined analysis:
| Analysis ID | AN003999 |
|---|---|
| Analysis type | MS |
| Chromatography type | GC |
| Chromatography system | Agilent 8890 |
| Column | Agilent HP5-MS (30m x 0.25mm, 0.25 um) |
| MS Type | EI |
| MS instrument type | Single quadrupole |
| MS instrument name | Agilent 5977B |
| Ion Mode | POSITIVE |
| Units | Fractional Enrichment (adjusted for natural abundance) |
Chromatography:
| Chromatography ID: | CH002953 |
| Instrument Name: | Agilent 8890 |
| Column Name: | Agilent HP5-MS (30m x 0.25mm, 0.25 um) |
| Column Temperature: | 60°C, rising at 10°C/min to 325°C, and held for 10 min |
| Flow Gradient: | N/A |
| Flow Rate: | 0.581 mL/min |
| Solvent A: | N/A |
| Solvent B: | N/A |
| Chromatography Type: | GC |
MS:
| MS ID: | MS003747 |
| Analysis ID: | AN003999 |
| Instrument Name: | Agilent 5977B |
| Instrument Type: | Single quadrupole |
| MS Type: | EI |
| MS Comments: | Metabolites were identified and quantified using DExSI v1.11 (https://github.com/DExSI/DExSI) - Dagley, M. J. and McConville, M. J. (2018) "DExSI: A new tool for the rapid quantitation of 13C-labelled metabolites detected by GC-MS", Bioinformatics, bty025, https://doi.org/10.1093/bioinformatics/bty025. |
| Ion Mode: | POSITIVE |
