#METABOLOMICS WORKBENCH araskind_20150606_9816311_mwtab.txt DATATRACK_ID:275 STUDY_ID:ST000178 ANALYSIS_ID:AN000275 PROJECT_ID:PR000153 VERSION 1 CREATED_ON 01-15-2019 #PROJECT PR:PROJECT_TITLE Graft-Versus-Host Disease PR:PROJECT_SUMMARY T cell metabolism during graft-versus-host disease PR:INSTITUTE University of Michigan PR:DEPARTMENT Pediatrics PR:LABORATORY Ferrara lab PR:LAST_NAME Byersdorfer PR:FIRST_NAME Craig PR:ADDRESS Ann Arbor, MI PR:EMAIL craigbye@med.umich.edu PR:PHONE 734-000-0000 PR:DOI http://dx.doi.org/10.21228/M8B01C #STUDY ST:STUDY_TITLE T cell metabolism during graft-versus-host disease (CAB 307)-PART I ST:STUDY_TYPE Glycolysis/TCA/Nucleotide analysis (tissue/cells) ST:STUDY_SUMMARY T cells were injected into mice to model graft-versus-host disease. On day 7 ST:STUDY_SUMMARY after bone marrow transplantation (BMT), cells were recovered and CD8 T cells ST:STUDY_SUMMARY were purified to > 90% purity over magnetic columns. Naïve T cells were used ST:STUDY_SUMMARY as control. Cells were then plated at 5x10^6 cells/ml onto plates coated with ST:STUDY_SUMMARY anti-CD3/CD28 antibodies in the presence of 300µM 13C-palmitate conjugated to ST:STUDY_SUMMARY BSA (in PBS). Cells were incubated at 37oC for 60min, after which time they ST:STUDY_SUMMARY were removed from the plates, counted, and an equal number (2.5x10^6) were ST:STUDY_SUMMARY placed into a 1.7 ml micro-centrifuge tubes and spun down. Cells were washed ST:STUDY_SUMMARY once with ammonium chloride, residual volume was removed by a second brief spin, ST:STUDY_SUMMARY cell pellets were flash frozen in liquid nitrogen and stored at -80oC until ST:STUDY_SUMMARY analysis. Several mice lines were used in this study. ST:INSTITUTE University of Michigan ST:DEPARTMENT Biomedical Research Core Facilities ST:LABORATORY Metabolomics core ST:LAST_NAME Kachman ST:FIRST_NAME Maureen ST:ADDRESS 6300 Brehm Tower, 1000 Wall Street, Ann Arbor, MI 48105-5714 ST:EMAIL mkachman@umich.edu ST:PHONE - #SUBJECT SU:SUBJECT_TYPE Animal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:SPECIES_GROUP Mammal #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS SU0004282 S00015097 Treatment:Control SUBJECT_SAMPLE_FACTORS SU0004282 S00015098 Treatment:Control SUBJECT_SAMPLE_FACTORS SU0004282 S00015099 Treatment:Control SUBJECT_SAMPLE_FACTORS SU0004282 S00015100 Treatment:Control SUBJECT_SAMPLE_FACTORS SU0004281 S00015094 Treatment:GVHD SUBJECT_SAMPLE_FACTORS SU0004281 S00015095 Treatment:GVHD SUBJECT_SAMPLE_FACTORS SU0004281 S00015096 Treatment:GVHD #COLLECTION CO:SAMPLE_TYPE Cells CO:COLLECTION_SUMMARY - #TREATMENT TR:TREATMENT_SUMMARY - #SAMPLEPREP SP:SAMPLEPREP_PROTOCOL_FILENAME Gly-TCA-nucleotides_analysis_protocol-2015-03-09.docx SP:SAMPLEPREP_SUMMARY - #CHROMATOGRAPHY CH:METHODS_ID AQM011 CH:METHODS_FILENAME ALPHA_KETO_ACIDS-FULL.M.zip CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Agilent DB5-MS (30m × 0.25mm, 0.25um) CH:CHROMATOGRAPHY_TYPE GC #ANALYSIS AN:LABORATORY_NAME MRC2 (University of Michigan) AN:ANALYSIS_TYPE MS AN:ACQUISITION_PARAMETERS_FILE ALPHA_KETO_ACIDS-FULL.M.zip AN:PROCESSING_PARAMETERS_FILE EX00317-MassHunterQuant-GlyTCA-DataAnalysis-GCMS-Method.m.zip #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 5975C MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS pmol/µg protein MS_METABOLITE_DATA_START Samples S00015097 S00015098 S00015099 S00015100 S00015094 S00015095 S00015096 Factors Treatment:Control Treatment:Control Treatment:Control Treatment:Control Treatment:GVHD Treatment:GVHD Treatment:GVHD Alanine 1.1190 0.9812 1.4374 alpha-Ketoglutarate 0.6082 0.6201 0.6114 0.4602 0.4289 0.3928 0.4765 Aspartate 11.5010 12.0246 17.0131 8.5928 24.8296 24.0247 27.3494 fumarate 1.2111 1.2342 1.2184 0.9111 0.9026 0.8082 0.9775 Glutamate 0.9925 0.9029 0.8984 0.5906 1.8674 1.7041 2.1098 glutamine 2.2511 1.8603 1.8879 2.0947 Lactate 1.8413 1.5102 1.2362 2.8291 1.2722 Oxaloacetate pyruvate 3.0709 3.1656 3.1729 2.3696 2.4151 2.2261 2.8650 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name pubchem_id inchi_key kegg_id other_id other_id_type ri ri_type moverz_quant Alanine 5950 C00041 ALA UM_Target_ID alpha-Ketoglutarate 51 C00026 AKG UM_Target_ID Aspartate 5960 C00049 ASP UM_Target_ID fumarate 444972 C00122 FUM UM_Target_ID Glutamate 611 GLU UM_Target_ID glutamine 5961 C00064 GLN UM_Target_ID Lactate 107689 C00186 LAC UM_Target_ID Oxaloacetate 970 C00036 OAA UM_Target_ID pyruvate 1060 C00022 PYR UM_Target_ID METABOLITES_END #END