Summary of Study ST002379

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 PR001524. The data can be accessed directly via it's Project DOI: 10.21228/M8Q13W This work is supported by NIH grant, U2C- DK119886.

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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.

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Study IDST002379
Study TitleGlucose flux analysis of NLRP3 inflammasome activated macrophages
Study TypeBasic research
Study SummaryActivating macrophage NLRP3 inflammasome can promote excessive inflammation, with severe cell and tissue damage and organ dysfunction. Here, we show that pharmacological or genetic inhibition of pyruvate dehydrogenase kinase (PDHK) significantly attenuates NLRP3 inflammasome activation in murine and human macrophages and septic mice by lowering caspase-1 cleavage and IL-1beta secretion. Inhibiting PDHK reverses NLRP3 inflammasome-induced metabolic reprogramming, enhances autophagy, promotes mitochondrial fusion over fission, preserves cristae ultrastructure, and attenuates mitochondrial ROS production. The suppressive effect of PDHK inhibition on the NLRP3 inflammasome is independent of its canonical role as a pyruvate dehydrogenase regulator. We suggest that PDHK inhibition improves mitochondrial fitness by reversing NLRP3 inflammasome activation in acutely inflamed macrophages.
Institute
Wake Forest School of Medicine
Last NameZhu
First NameXuewei
Address575 Patterson Ave, Winston-Salem, NC 27101
Emailxwzhu@wakehealth.edu
Phone3367131445
Submit Date2022-11-15
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2022-12-15
Release Version1
Xuewei Zhu Xuewei Zhu
https://dx.doi.org/10.21228/M8Q13W
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001524
Project DOI:doi: 10.21228/M8Q13W
Project Title:Pyruvate dehydrogenase kinase supports macrophage NLRP3 inflammasome activation during acute inflammation
Project Type:Basic research
Project Summary:Activating macrophage NLRP3 inflammasome can promote excessive inflammation, with severe cell and tissue damage and organ dysfunction. Here, we show that pharmacological or genetic inhibition of pyruvate dehydrogenase kinase (PDHK) significantly attenuates NLRP3 inflammasome activation in murine and human macrophages and septic mice by lowering caspase-1 cleavage and IL-1beta secretion. Inhibiting PDHK reverses NLRP3 inflammasome-induced metabolic reprogramming, enhances autophagy, promotes mitochondrial fusion over fission, preserves cristae ultrastructure, and attenuates mitochondrial ROS production. The suppressive effect of PDHK inhibition on the NLRP3 inflammasome is independent of its canonical role as a pyruvate dehydrogenase regulator. We suggest that PDHK inhibition improves mitochondrial fitness by reversing NLRP3 inflammasome activation in acutely inflamed macrophages.
Institute:Wake Forest School of Medicine
Last Name:Zhu
First Name:Xuewei
Address:575 Patterson Ave
Email:xwzhu@wakehealth.edu
Phone:3367131445

Subject:

Subject ID:SU002468
Subject Type:Cultured cells
Subject Species:Mus musculus
Taxonomy ID:10090
Species Group:Mammals

Factors:

Subject type: Cultured cells; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Treatment
SA237431LPS_JX06_ATP_2JPS+JX06+ATP
SA237432LPS_JX06_ATP_1JPS+JX06+ATP
SA237433LPS_JX06_ATP_3JPS+JX06+ATP
SA237434LPS_3LPS
SA237435LPS_1LPS
SA237436LPS_2LPS
SA237437LPS_ATP_3LPS+ATP
SA237438LPS_ATP_1LPS+ATP
SA237439LPS_ATP_2LPS+ATP
SA237440Ctrl_C_2No treatment
SA237441Ctrl_C_1No treatment
SA237442Ctrl_C_3No treatment
Showing results 1 to 12 of 12

Collection:

Collection ID:CO002461
Collection Summary:Mouse bone marrow was cultured in low glucose DMEM supplemented with 30% L929 cell-conditioned medium, 20% FBS, 2 mM L-glutamine, 1 mM sodium pyruvate, 100 U/ml penicillin, and 100 µg/ml streptomycin for 6-7 days until the cells reached confluence. BMDMs were then reseeded in culture dishes overnight in RPMI-1640 medium containing 1% Nutridoma-SP medium (Sigma-Aldrich). LPS-primed BMDMs were stimulated with 5 mM ATP in the presence or absence of 10 uM JX06 for 30 min. Unstimulated macrophages were used as control.13C tracing started by replacing medium with glucose-free medium supplemented with 2.06 g/L [U-13C]-glucose (Cayman Chemical) for 90 min. Cells were washed with ice-cold saline and intracellular metabolites were extracted using cold methanol and H2O (80:20; HPLC Grade; Sigma-Aldrich). Here, we show that pharmacological or genetic inhibition of pyruvate dehydrogenase kinase (PDHK) significantly attenuates NLRP3 inflammasome activation in murine and human macrophages and septic mice by lowering caspase-1 cleavage and IL-1beta secretion. Inhibiting PDHK reverses NLRP3 inflammasome-induced metabolic reprogramming, enhances autophagy, promotes mitochondrial fusion over fission, preserves cristae ultrastructure, and attenuates mitochondrial ROS production. The suppressive effect of PDHK inhibition on the NLRP3 inflammasome is independent of its canonical role as a pyruvate dehydrogenase regulator. We suggest that PDHK inhibition improves mitochondrial fitness by reversing NLRP3 inflammasome activation in acutely inflamed macrophages.
Sample Type:Macrophages
Collection Method:Cells were washed with ice-cold saline and intracellular metabolites were extracted using cold methanol and H2O (80:20; HPLC Grade; Sigma-Aldrich). The samples were pre-dissolved into 40 µL solution (LC-water, methanol, Acetonitrile 2:1:1) before injection.
Storage Conditions:-80℃

Treatment:

Treatment ID:TR002480
Treatment Summary:LPS-primed BMDMs were stimulated with or without 5 mM ATP in the presence or absence of 10 uM JX06 for 30 min. 13C tracing started by replacing medium with glucose-free medium supplemented with 2.06 g/L [U-13C]-glucose (Cayman Chemical) for 90 min.
Treatment:in vitro cell culture treatment
Treatment Compound:LPS, ATP, JX06

Sample Preparation:

Sampleprep ID:SP002474
Sampleprep Summary:Macrophages were lysed, and polar metabolites were extracted using methanol and H2O (80:20; HPLC Grade; Sigma-Aldrich) (Liu et al., 2015). Extracts were dried in a vacuum concentrator at room temperature and stored at -80 freezer.

Chromatography:

Chromatography ID:CH002873
Chromatography Summary:Ultimate 3000 UHPLC (Dionex) is coupled to Q Exactive Plus-Mass spectrometer (QE-MS, Thermo Scientific) for metabolite profiling.
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Waters XBridge Amide (100 x 4.6mm,3.5um)
Chromatography Type:HILIC

Analysis:

Analysis ID:AN003877
Laboratory Name:Jason Locosale
Analysis Type:MS
Operator Name:Likun Duan
Chromatography ID:CH002873
Num Factors:4
Num Metabolites:224
Units:Fractional enrichment
  
Analysis ID:AN003878
Laboratory Name:Jason Locosale
Analysis Type:MS
Operator Name:Likun Duan
Chromatography ID:CH002873
Num Factors:4
Num Metabolites:298
Units:Fractional enrichment
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