Summary of Study ST002786

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 PR001736. The data can be accessed directly via it's Project DOI: 10.21228/M89Q7K 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 IDST002786
Study TitleMetabolic changes of the pig kidney during isolated normothermic perfusion with autologous whole blood - urine
Study TypeExperimental
Study SummaryThis study investigates how glucose, lactate and 20 amino acids in the urine of normothermically perfused pig kidneys changes over time. It also studies how type and severity of ischemia before perfusion (cold or warm ischemia) change this profile.
Institute
KU Leuven
DepartmentMicrobiology, Immunology and Transplantation
LaboratoryLab of Abdominal Transplantation
Last NameJochmans
First NameIna
AddressHerestraat 49, Leuven, 3000, Belgium
Emailina.jochmans@kuleuven.be
PhoneNone
Submit Date2023-07-17
Num Groups2
Total Subjects12
Num Males6 male pigs
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailLC-MS
Release Date2023-08-07
Release Version1
Ina Jochmans Ina Jochmans
https://dx.doi.org/10.21228/M89Q7K
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001736
Project DOI:doi: 10.21228/M89Q7K
Project Title:Metabolic changes during normothermic isolated kidney perfusion
Project Type:Experimental
Project Summary:Normothermic isolated kidney perfusion is being developed as a method to preserve donor kidneys and to assess their future function before they are transplanted. Donor kidneys can be exposed to different types and degrees of ischemic injury before they are preserved. This project studies how the metabolome changes in relation to different types and degrees of ischemia.
Institute:KU Leuven
Department:Microbiology, Immunology and Transplantation
Laboratory:Lab of Abdominal Transplantation
Last Name:Jochmans
First Name:Ina
Address:Herestraat 49, Leuven, 3000, Belgium
Email:ina.jochmans@kuleuven.be
Phone:None
Funding Source:FTBO, KOOR, FWO
Contributors:Julie De Beule, Bart Ghesquière, Ina Jochmans

Subject:

Subject ID:SU002893
Subject Type:Mammal
Subject Species:Sus scrofa
Taxonomy ID:9823
Age Or Age Range:~4 months
Weight Or Weight Range:35-45 kg
Gender:Male
Animal Animal Supplier:TOPIGS TN70, Tojapigs, the Netherlands
Animal Light Cycle:12 hours
Animal Water:ad libitum

Factors:

Subject type: Mammal; Subject species: Sus scrofa (Factor headings shown in green)

mb_sample_id local_sample_id Timepoint Group Urine_volume
SA29893929_T2120 CI 19
SA29894075_T2120 CI 27
SA29894130_T2120 CI 33
SA29894233_T2120 CI 45
SA29894371_T2120 CI 50
SA29894472_T2120 CI 85
SA29894534_T2120 Control 29
SA29894670_T2120 Control 47
SA29894731_T2120 Control 60
SA29894867_T2120 Control 8
SA29894976_T2120 Control 80
SA29895019_T2120 Control 90
SA29895130_T3180 CI 20
SA29895271_T3180 CI 24
SA29895375_T3180 CI 3
SA29895429_T3180 CI 33
SA29895572_T3180 CI 60
SA29895633_T3180 CI 80
SA29895719_T3180 Control 100
SA29895876_T3180 Control 110
SA29895967_T3180 Control 3
SA29896031_T3180 Control 42
SA29896134_T3180 Control 60
SA29896270_T3180 Control 90
SA29896375_T4240 CI 0
SA29896471_T4240 CI 10
SA29896530_T4240 CI 15
SA29896633_T4240 CI 26
SA29896729_T4240 CI 32
SA29896872_T4240 CI 50
SA29896976_T4240 Control 105
SA29897067_T4240 Control 12
SA29897131_T4240 Control 17
SA29897234_T4240 Control 28
SA29897370_T4240 Control 42
SA29897419_T4240 Control 50
SA29897575_T160 CI 120
SA29897633_T160 CI 21
SA29897771_T160 CI 30
SA29897872_T160 CI 35
SA29897929_T160 CI 67
SA29898030_T160 CI 84
SA29898176_T160 Control 105
SA29898219_T160 Control 46
SA29898331_T160 Control 57
SA29898434_T160 Control 67
SA29898567_T160 Control 70
SA29898670_T160 Control 80
Showing results 1 to 48 of 48

Collection:

Collection ID:CO002886
Collection Summary:Urine production over 1 hour of perfusion was diverted to a collection bag. At the end of the hour, urine samples were taken in dry tubes from this collection bag. One milliliter perfusate aliquots (centrifuged 1000g, 10 minutes, 4°C) were snap frozen in liquid nitrogen and stored at -80°C until analysis.
Sample Type:Urine
Collection Frequency:Every hour during perfusion, so T1 a sample from urine produced during the first hour of perfusion, T2 for the second hour, T3 for the third hour, and T4 for the fourth hour
Storage Conditions:-80℃
Additives:None

Treatment:

Treatment ID:TR002902
Treatment Summary:Three experimental conditions were investigated (n=6/group): (a) Controls; (b) warm ischemia (WI) simulating hypoxic acute kidney injury; (c) cold ischemia (CI) replicating clinical cold storage. Control kidneys were retrieved, flushed with cold IGL-1, and immediately reperfused. CI group, kidneys were exposed to 22h of CI by submerging them in IGL- 1 (a clinical preservation solution for cold storage) and storing them on ice. In WI, the renal artery and vein were clamped for 60 min before retrieval. In the All kidneys were flushed with 200 ml of Ringer’s solution before mounting them on the ex situ circuit to wash out IGL-1. Because little urine was produced by WI kidneys, too few samples were available for a meaningful analysis.
Animal Anesthesia:Pigs were sedated by an intramuscular injection of Tiletamine/Zolazepam (8 mg/kg, Zoletil®, Virbac, Belgium) and Xylazine (2 mg/kg, Xylazine®, VMD pharma, Belgium) to allow orotracheal intubation. Anesthesia was maintained by inhalation of isoflurane (1% Isovet®, Piramal Critical Care B.V., Belgium) and continuous infusion of fentanyl (8 µg/kg, Fentanyl®, Janssen Pharmaceutica, Belgium).
Animal Acclimation Duration:Minimum of 2 days
Animal Fasting:Overnight
Animal Endp Euthanasia:Non-recovery study

Sample Preparation:

Sampleprep ID:SP002899
Sampleprep Summary:Samples were extracted in an 80% methanol (80:20 methanol:water) (Methanol ≥99.9%, HiPerSolv CHROMANORM®, ULTRA for LC-MS, suitable for UPLC/UHPLC-MS instruments, VWR, Belgium) extraction buffer containing 1 µM of deuterated D27 myristic acid, 5 µM D12 glucose, 3 µM 13C5-D5-15N Glutamic acid and 3 µM D7-15N4-Arginine as internal standards. 10 µl of sample was added to 990 µl of the extraction buffer and stored overnight at -80 °C. Insolubilities and precipitated proteins were removed by centrifugation at 20.000 g, for 15 min at 4 °C. 200 µL of the supernatant was transferred to an appropriate mass-spectrometry vial. 50 µl of samples was added to 950 µl of the extraction buffer and stored overnight at -80°C.
Extract Storage:-80℃

Combined analysis:

Analysis ID AN004533
Analysis type MS
Chromatography type Reversed phase
Chromatography system Thermo Dionex Ultimate 3000
Column Waters ACQUITY UPLC BEH C18 (150 x 2.1mm,1.7um)
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Q Exactive Focus
Ion Mode NEGATIVE
Units Peak area

Chromatography:

Chromatography ID:CH003406
Chromatography Summary:10 µl of each sample was loaded into a Dionex UltiMate 3000 LC System (Thermo Scientific Bremen, Germany) equipped with a Waters ACQUITY UPLC BEH C18 (150 x 2.1mm,1.7um) column coupled to a Q Exactive Orbitrap mass spectrometer (Thermo Scientific) operating in negative ion mode. A step gradient was carried out using solvent A (10 mM TBA and 15 mM acetic acid) and solvent B (100% methanol). The gradient started with 5% of solvent B and 95% solvent A and remained at 5% B until 2 min post injection. A linear gradient to 37% B was carried out until 7 min and increased to 41% until 14 min. Between 14 and 26 minutes the gradient increased to 95% of B and remained at 95% B for 4 minutes. At 30 min the gradient returned to 5% B. The chromatography was stopped at 40 min. The flow was kept constant at 0.25 mL/min and the column was placed at 40°C throughout the analysis. The MS operated in full scan mode (m/z range: [70.0000-1050.0000]) using a spray voltage of 4.80 kV, capillary temperature of 300°C, sheath gas at 40.0, auxiliary gas at 10.0. The AGC target was set at 3.0E+006 using a resolution of 140000, with a maximum IT fill time of 512 ms. Data collection was performed using the Xcalibur software (Thermo Scientific). The data analyses were performed by integrating the peak areas (El-Maven - Polly - Elucidata).
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Waters ACQUITY UPLC BEH C18 (150 x 2.1mm,1.7um)
Column Temperature:40°C
Flow Gradient:The gradient started with 5% of solvent B and 95% solvent A and remained at 5% B until 2 min post injection. A linear gradient to 37% B was carried out until 7 min and increased to 41% until 14 min. Between 14 and 26 minutes the gradient increased to 95% of B and remained at 95% B for 4 minutes. At 30 min the gradient returned to 5% B. The chromatography was stopped at 40 min.
Flow Rate:0.25 ml/min
Solvent A:100% water; 10 mM TBA; 15 mM acetic acid
Solvent B:100% methanol
Chromatography Type:Reversed phase

MS:

MS ID:MS004280
Analysis ID:AN004533
Instrument Name:Thermo Q Exactive Focus
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Raw MS data were converted into mzML using the MSConvert tool of Proteowizard (version 3.0.20247). Peak picking was performed with El_Maven (El_Maven, Elucidata, Massachusetts, USA, version 0.12.0). Metabolites were identified using an in-house library containing exact mass and retention time. The mass accuracy during data processing in El Maven was set at 10 ppm. Calculation of abundances was done in the LC-MS Workflow of El_Maven. Raw abundances (peak area values) for each metabolite were corrected for internal standard (Myristic acid d27). These were corrected for urine volume.
Ion Mode:NEGATIVE
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