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.

Perform statistical analysis  |  Show all samples  |  Show named metabolites  |  Download named metabolite data  
Download mwTab file (text)   |  Download mwTab file(JSON)   |  Download data files (Contains raw data)
Study IDST002451
Study TitleAPOE modulates microglial immunometabolism in response to age, amyloid pathology, and inflammatory challenge (Part 2 of 3)
Study SummaryThe 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
DepartmentPhysiology
LaboratoryLance Johnson; Josh Morganti
Last NameDevanney
First NameNicholas
AddressPhysiology, 760 Press Ave, Healthy Kentucky Research Bldg, Rm152, Lexington, Kentucky, 40508, USA
EmailNicholas.Devanney@uky.edu
Phone8593238083
Submit Date2023-01-20
Study CommentsPart 2 of 3
Raw Data AvailableYes
Raw Data File Type(s)cdf
Analysis Type DetailGC-MS
Release Date2023-01-25
Release Version1
Nicholas Devanney Nicholas Devanney
https://dx.doi.org/10.21228/M8G711
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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
SA24505847_3_12E3_control E3/E3 Control
SA24505947_3_1E3_control E3/E3 Control
SA24506047_3_11E3_control E3/E3 Control
SA24506147_3_10E3_control E3/E3 Control
SA24506247_3_2E3_control E3/E3 Control
SA24506347_3_4E3_control E3/E3 Control
SA24506447_3_3E3_control E3/E3 Control
SA24506547_3_9E3_control E3/E3 Control
SA24506647_3_14E3_proinflammatory E3/E3 Proinflammatory
SA24506747_3_15E3_proinflammatory E3/E3 Proinflammatory
SA24506847_3_13E3_proinflammatory E3/E3 Proinflammatory
SA24506947_3_16E3_proinflammatory E3/E3 Proinflammatory
SA24507047_3_8E3_proinflammatory E3/E3 Proinflammatory
SA24507147_3_5E3_proinflammatory E3/E3 Proinflammatory
SA24507247_3_6E3_proinflammatory E3/E3 Proinflammatory
SA24507347_3_7E3_proinflammatory E3/E3 Proinflammatory
SA24507447_4_12E4_control E4/E4 Control
SA24507547_4_9E4_control E4/E4 Control
SA24507647_4_10E4_control E4/E4 Control
SA24507747_4_4E4_control E4/E4 Control
SA24507847_4_2E4_control E4/E4 Control
SA24507947_4_1E4_control E4/E4 Control
SA24508047_4_3E4_control E4/E4 Control
SA24508147_4_15E4_proinflammatory E4/E4 Proinflammatory
SA24508247_4_16E4_proinflammatory E4/E4 Proinflammatory
SA24508347_4_14E4_proinflammatory E4/E4 Proinflammatory
SA24508447_4_8E4_proinflammatory E4/E4 Proinflammatory
SA24508547_4_6E4_proinflammatory E4/E4 Proinflammatory
SA24508647_4_7E4_proinflammatory E4/E4 Proinflammatory
SA24508747_4_13E4_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
  logo