Summary of Study ST000413

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 PR000322. The data can be accessed directly via it's Project DOI: 10.21228/M8V312 This work is supported by NIH grant, U2C- DK119886.

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Study IDST000413
Study TitleMetabolic profiling during ex vivo machine perfusion of the human liver (part III)
Study SummaryAs donor organ shortages persist, functional machine perfusion is under investigation to improve preservation of the donor liver. The transplantation of donation after circulatory death (DCD) livers is limited by poor outcomes, but its application may be expanded by ex vivo repair and assessment of the organ before transplantation. Here we employed subnormothermic (21 °C) machine perfusion of discarded human livers combined with metabolomics to gain insight into metabolic recovery during machine perfusion. Improvements in energetic cofactors and redox shifts were observed, as well as reversal of ischemia-induced alterations in selected pathways, including lactate metabolism and increased TCA cycle intermediates. We next evaluated whether DCD livers with steatotic and severe ischemic injury could be discriminated from ‘transplantable’ DCD livers. Metabolomic profiling was able to cluster livers with similar metabolic patterns based on the degree of injury. Moreover, perfusion parameters combined with differences in metabolic factors suggest variable mechanisms that result in poor energy recovery in injured livers. We conclude that machine perfusion combined with metabolomics has significant potential as a clinical instrument for the assessment of preserved livers.
Institute
University of California, Davis
DepartmentGenome and Biomedical Sciences Facility
LaboratoryWCMC Metabolomics Core
Last NameFiehn
First NameOliver
Address1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, CA 95616
Emailofiehn@ucdavis.edu
Phone(530) 754-8258
Submit Date2016-07-01
Study CommentsThe first 4 samples were a test run to see how efficient the analysis was and were run on a lipidomics platform. The next 12 samples were the used in the paper and were the same as the original 4 samples, but they were split into 3 biological replicates and run on the GC platform.
Publicationsdoi:10.1038/srep22415
Raw Data AvailableYes
Raw Data File Type(s)d
Analysis Type DetailLC-MS
Release Date2016-09-23
Release Version1
Oliver Fiehn Oliver Fiehn
https://dx.doi.org/10.21228/M8V312
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000322
Project DOI:doi: 10.21228/M8V312
Project Title:Metabolic profiling during ex vivo machine perfusion of the human liver
Project Summary:As donor organ shortages persist, functional machine perfusion is under investigation to improve preservation of the donor liver. The transplantation of donation after circulatory death (DCD) livers is limited by poor outcomes, but its application may be expanded by ex vivo repair and assessment of the organ before transplantation. Here we employed subnormothermic (21 °C) machine perfusion of discarded human livers combined with metabolomics to gain insight into metabolic recovery during machine perfusion. Improvements in energetic cofactors and redox shifts were observed, as well as reversal of ischemia-induced alterations in selected pathways, including lactate metabolism and increased TCA cycle intermediates. We next evaluated whether DCD livers with steatotic and severe ischemic injury could be discriminated from ‘transplantable’ DCD livers. Metabolomic profiling was able to cluster livers with similar metabolic patterns based on the degree of injury. Moreover, perfusion parameters combined with differences in metabolic factors suggest variable mechanisms that result in poor energy recovery in injured livers. We conclude that machine perfusion combined with metabolomics has significant potential as a clinical instrument for the assessment of preserved livers.
Institute:University of California, Davis
Department:Genome and Biomedical Sciences Facility
Laboratory:WCMC Metabolomics Core
Last Name:Fiehn
First Name:Oliver
Address:1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, CA 95616
Email:ofiehn@ucdavis.edu
Phone:(530) 754-8258
Funding Source:NIH U24DK097154

Subject:

Subject ID:SU000434
Subject Type:Human
Subject Species:Homo sapiens
Taxonomy ID:9606
Species Group:Human

Factors:

Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Collection Time
SA020296Inj02_SA14_BruinCSH.d-
SA020297Inj11_SA15_BruinCSH.d-
SA020298Inj09_SA01_BruinCSH.d-
SA020299Inj04_SA13_BruinCSH.d-
SA020300Inj22_SA02_BruinCSH.d-
SA020301Inj14_SA03_BruinCSH.d-
SA020302Inj06_SA04_BruinCSH.d1 Hr
SA020303Inj20_SA06_BruinCSH.d1 Hr
SA020304Inj23_SA17_BruinCSH.d1 Hr
SA020305Inj03_SA18_BruinCSH.d1 Hr
SA020306Inj10_SA05_BruinCSH.d1 Hr
SA020307Inj15_SA16_BruinCSH.d1 Hr
SA020308Inj08_SA20_BruinCSH.d2 Hr
SA020309Inj19_SA21_BruinCSH.d2 Hr
SA020310Inj01_SA07_BruinCSH.d2 Hr
SA020311Inj17_SA09_BruinCSH.d2 Hr
SA020312Inj13_SA19_BruinCSH.d2 Hr
SA020313Inj05_SA08_BruinCSH.d2 Hr
SA020314Inj07_SA24_BruinCSH.d3 Hr
SA020315Inj24_SA11_BruinCSH.d3 Hr
SA020316Inj18_SA23_BruinCSH.d3 Hr
SA020317Inj16_SA22_BruinCSH.d3 Hr
SA020318Inj12_SA10_BruinCSH.d3 Hr
SA020319Inj21_SA12_BruinCSH.d3 Hr
Showing results 1 to 24 of 24

Collection:

Collection ID:CO000428
Collection Summary:Liver biopsies were snap frozen in liquid nitrogen and stored at ­80. Tissue was pulverized, while frozen and tissue was aliquoted into .6 ml tubes between 30 and 70 mg of liver tissue. 3 replicates are provided per sample.
Collection Protocol Filename:StudyDesign_BoteBruinsma_072414.pdf
Sample Type:Tissue
Storage Conditions:-80 C
Tissue Cell Quantity Taken:between 30 and 70 mg

Treatment:

Treatment ID:TR000448
Treatment Summary:2 livers (names 23 and 31) were perfused for 3 hours, providing 4 time­course (0, 1, 2, 3) biopsies. 3 replicates for each biopsy (A, B ,C). These were the only 2 run on the Lipidomics platform.
Treatment Protocol Filename:StudyDesign_BoteBruinsma_072414.pdf

Sample Preparation:

Sampleprep ID:SP000441
Sampleprep Summary:1. Weigh 4mg tissue sample in to a 2mL Eppendorf tube. 2. Add 1mL extraction solvent to the tissue sample and homogenize for 45 seconds ensuring that sample resembles a powder. In between samples, clean the homogenizer in solutions of methanol, acetone, water, and the extraction solvent in the order listed. 3. Vortex samples for 10 seconds, then 5 minutes on 4°C shaker. 4. Centrifuge the samples for 2 minutes at 14,000 rcf. Aliquot 500μL supernatant for analysis, and 500μL for a backup. Store backup aliquots in the -20°C freezer. 5. Evaporate one 500μl analysis aliquot in the Labconco Centrivap cold trap concentrator to complete dryness (typically overnight). 6. The dried aliquot is then re-suspended with 500l 50% acetonitrile (degassed as given) 7. Centrifuge for 2 minutes at 14,000 rcf using the centrifuge Eppendorf 5415. 8. Remove supernatant to a new Eppendorf tube. 9. Evaporate the supernatant to dryness in the the Labconco Centrivap cold trap concentrator. 10. Submit to derivatization.
Sampleprep Protocol Filename:SOP_Extraction_of_Liver_Tissue_Samples.pdf
Processing Method:Homogenization

Combined analysis:

Analysis ID AN000653 AN000654
Analysis type MS MS
Chromatography type Reversed phase Reversed phase
Chromatography system Agilent 6530 Agilent 6550
Column Waters Acquity CSH C18 (100 x 2.1mm, 1.7um) Waters Acquity CSH C18 (100 x 2.1mm, 1.7um)
MS Type ESI ESI
MS instrument type QTOF QTOF
MS instrument name Agilent 6530 QTOF Agilent 6550 QTOF
Ion Mode POSITIVE NEGATIVE
Units counts counts

Chromatography:

Chromatography ID:CH000471
Methods Filename:Data_Dictionary_Fiehn_laboratory_CSH_QTOF_lipidomics_05-29-2014.pdf
Instrument Name:Agilent 6530
Column Name:Waters Acquity CSH C18 (100 x 2.1mm, 1.7um)
Column Pressure:450-850 bar
Column Temperature:65 C
Flow Gradient:15% B to 99%B
Flow Rate:0.6 mL/min
Internal Standard:See data dictionary
Retention Time:See data dictionary
Sample Injection:1.67 uL
Solvent A:60:40 Acetonitrile:Water +10mM Ammonium Formate +10mM Formic Acid
Solvent B:9:1 Isopropanol:Acetonitrile +10mM Ammonium Formate +10mM Formic Acid
Analytical Time:13 min
Capillary Voltage:3500 V
Time Program:15 min
Weak Wash Solvent Name:Isopropanol
Strong Wash Solvent Name:Isopropanol
Target Sample Temperature:Autosampler temp 4 C
Randomization Order:Excel generated
Chromatography Type:Reversed phase
  
Chromatography ID:CH000472
Methods Filename:Data_Dictionary_Fiehn_laboratory_CSH_QTOF_lipidomics_05-29-2014.pdf
Instrument Name:Agilent 6550
Column Name:Waters Acquity CSH C18 (100 x 2.1mm, 1.7um)
Column Pressure:450-850 bar
Column Temperature:65 C
Flow Gradient:15% B to 99%B
Flow Rate:0.6 mL/min
Internal Standard:See data dictionary
Retention Time:See data dictionary
Sample Injection:5 uL
Solvent A:60:40 Acetonitrile:Water +10mM Ammonium Acetate +10mM Acetic Acid
Solvent B:9:1 Isopropanol:Acetonitrile +10mM Ammonium Acetate +10mM Acetic Acid
Analytical Time:13 min
Capillary Voltage:3500 V
Time Program:15 min
Weak Wash Solvent Name:Isopropanol
Strong Wash Solvent Name:Isopropanol
Target Sample Temperature:Autosampler temp 4 C
Randomization Order:Excel generated
Chromatography Type:Reversed phase

MS:

MS ID:MS000579
Analysis ID:AN000653
Instrument Name:Agilent 6530 QTOF
Instrument Type:QTOF
MS Type:ESI
Ion Mode:POSITIVE
Capillary Voltage:3500 V
Collision Gas:Nitrogen
Dry Gas Flow:8 L/min
Dry Gas Temp:325 C
Fragment Voltage:120 V
Fragmentation Method:Auto MS/MS
Ion Source Temperature:325 C
Ion Spray Voltage:1000 V
Ionization:Pos
Precursor Type:Intact Molecule
Reagent Gas:Nitrogen
Source Temperature:325 C
Dataformat:.d
Desolvation Gas Flow:11 L/min
Desolvation Temperature:350 C
Nebulizer:35 psig
Octpole Voltage:750 V
Resolution Setting:extended dynamic range
Scan Range Moverz:60-1700 Da
Scanning Cycle:2 Hz
Scanning Range:60-1700 Da
Skimmer Voltage:65 V
  
MS ID:MS000580
Analysis ID:AN000654
Instrument Name:Agilent 6550 QTOF
Instrument Type:QTOF
MS Type:ESI
Ion Mode:NEGATIVE
Capillary Voltage:3500 V
Collision Gas:Nitrogen
Dry Gas Flow:13 L/min
Dry Gas Temp:200 C
Fragment Voltage:175 V
Fragmentation Method:Auto MS/MS
Ion Source Temperature:325 C
Ion Spray Voltage:1000 V
Ionization:Neg
Precursor Type:Intact Molecule
Reagent Gas:Nitrogen
Source Temperature:325 C
Dataformat:.d
Desolvation Gas Flow:11 L/min
Desolvation Temperature:350 C
Nebulizer:35 psig
Octpole Voltage:750 V
Resolution Setting:extended dynamic range
Scan Range Moverz:60-1700 Da
Scanning Cycle:2 Hz
Scanning Range:60-1700 Da
Skimmer Voltage:65 V
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