{
"METABOLOMICS WORKBENCH":{"VERSION":"1","CREATED_ON":"November 15, 2016, 8:44 am"},

"PROJECT":{"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","PUBLICATIONS":"Metabolic profiling during ex vivo machine perfusion of the human liver","DOI":"10.1038/srep22415"},

"STUDY":{"STUDY_TITLE":"Metabolic profiling during ex vivo machine perfusion of the human liver (part III)","STUDY_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","STUDY_COMMENTS":"The 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.","PUBLICATIONS":"doi:10.1038/srep22415"},

"SUBJECT":{"SUBJECT_TYPE":"Human","SUBJECT_SPECIES":"Homo sapiens","TAXONOMY_ID":"9606"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"23.0.A_01",
"Sample ID":"Inj09_SA01_BruinCSH.d",
"Factors":{"Collection Time":"-"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"23.0.B_02",
"Sample ID":"Inj22_SA02_BruinCSH.d",
"Factors":{"Collection Time":"-"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"23.0.C_03",
"Sample ID":"Inj14_SA03_BruinCSH.d",
"Factors":{"Collection Time":"-"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"23.1.A_04",
"Sample ID":"Inj06_SA04_BruinCSH.d",
"Factors":{"Collection Time":"1 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"23.1.B_05",
"Sample ID":"Inj10_SA05_BruinCSH.d",
"Factors":{"Collection Time":"1 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"23.1.C_06",
"Sample ID":"Inj20_SA06_BruinCSH.d",
"Factors":{"Collection Time":"1 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"23.2.A_07",
"Sample ID":"Inj01_SA07_BruinCSH.d",
"Factors":{"Collection Time":"2 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"23.2.B_08",
"Sample ID":"Inj05_SA08_BruinCSH.d",
"Factors":{"Collection Time":"2 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"23.2.C_09",
"Sample ID":"Inj17_SA09_BruinCSH.d",
"Factors":{"Collection Time":"2 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"23.3.A_10",
"Sample ID":"Inj12_SA10_BruinCSH.d",
"Factors":{"Collection Time":"3 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"23.3.B_11",
"Sample ID":"Inj24_SA11_BruinCSH.d",
"Factors":{"Collection Time":"3 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"23.3.C_12",
"Sample ID":"Inj21_SA12_BruinCSH.d",
"Factors":{"Collection Time":"3 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"31.0.A_13",
"Sample ID":"Inj04_SA13_BruinCSH.d",
"Factors":{"Collection Time":"-"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"31.0.B_14",
"Sample ID":"Inj02_SA14_BruinCSH.d",
"Factors":{"Collection Time":"-"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"31.0.C_15",
"Sample ID":"Inj11_SA15_BruinCSH.d",
"Factors":{"Collection Time":"-"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"31.1.A_16",
"Sample ID":"Inj15_SA16_BruinCSH.d",
"Factors":{"Collection Time":"1 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"31.1.B_17",
"Sample ID":"Inj23_SA17_BruinCSH.d",
"Factors":{"Collection Time":"1 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"31.1.C_18",
"Sample ID":"Inj03_SA18_BruinCSH.d",
"Factors":{"Collection Time":"1 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"31.2.A_19",
"Sample ID":"Inj13_SA19_BruinCSH.d",
"Factors":{"Collection Time":"2 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"31.2.B_20",
"Sample ID":"Inj08_SA20_BruinCSH.d",
"Factors":{"Collection Time":"2 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"31.2.C_21",
"Sample ID":"Inj19_SA21_BruinCSH.d",
"Factors":{"Collection Time":"2 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
},
{
"Subject ID":"31.3.A_22",
"Sample ID":"Inj16_SA22_BruinCSH.d",
"Factors":{"Collection Time":"3 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"1"}
},
{
"Subject ID":"31.3.B_23",
"Sample ID":"Inj18_SA23_BruinCSH.d",
"Factors":{"Collection Time":"3 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"2"}
},
{
"Subject ID":"31.3.C_24",
"Sample ID":"Inj07_SA24_BruinCSH.d",
"Factors":{"Collection Time":"3 Hr"},
"Additional sample data":{"Treatment":"steatotic liver","Replicate":"3"}
}
],
"COLLECTION":{"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_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"},

"SAMPLEPREP":{"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"},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Agilent 6530","COLUMN_NAME":"Waters Acquity CSH C18 (100 x 2.1mm, 1.7um) 1.7um Pre-Column","FLOW_GRADIENT":"15% B to 99%B","FLOW_RATE":"0.6 mL/min","COLUMN_TEMPERATURE":"65 C","METHODS_FILENAME":"Data_Dictionary_Fiehn_laboratory_CSH_QTOF_lipidomics_05-29-2014.pdf","SOLVENT_A":"60:40 Acetonitrile:Water +10mM Ammonium Formate +10mM Formic Acid","SOLVENT_B":"9:1 Isopropanol:Acetonitrile +10mM Ammonium Formate +10mM Formic Acid","COLUMN_PRESSURE":"450-850 bar","INTERNAL_STANDARD":"See data dictionary","RETENTION_TIME":"See data dictionary","SAMPLE_INJECTION":"1.67 uL","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"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS","LABORATORY_NAME":"WCMC Metabolomics Core","SOFTWARE_VERSION":"MassHunter","DATA_FORMAT":".d"},

"MS":{"MS_COMMENTS":"-","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_METABOLITE_DATA":{
"Units":"counts",

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