#METABOLOMICS WORKBENCH nurul777_20210113_083827 DATATRACK_ID:2396 STUDY_ID:ST001654 ANALYSIS_ID:AN002701 PROJECT_ID:PR001060 VERSION 1 CREATED_ON January 19, 2021, 4:30 pm #PROJECT PR:PROJECT_TITLE Investigation of infection-tolerance phenomenon in mammalian reservoir of Lyme PR:PROJECT_TITLE disease, Peromyscus leucopus PR:PROJECT_TYPE Comparative study of Peromyscus leucopus and Mus musculus, BALB/c, metabolic PR:PROJECT_TYPE response to endotoxin treatment PR:PROJECT_SUMMARY Infection-tolerance mechanism were investigated in a natural reservoir of Lyme PR:PROJECT_SUMMARY disease, Peromyscus leucopus, white-footed deermice following exposure of a PR:PROJECT_SUMMARY single dose of lipopolysaccharide endotoxin. The multiparametric responses of PR:PROJECT_SUMMARY this treatment were compared to widely used mouse model, Mus musculus BALB/c. PR:PROJECT_SUMMARY Following LPS and saline treatment, samples from blood, spleen and liver tissues PR:PROJECT_SUMMARY were collected postmortem from both animals and subjected to RNA-seq, untargeted PR:PROJECT_SUMMARY metabolomics and specific RT-qPCR. PR:INSTITUTE Colorado State University PR:DEPARTMENT Microbiology, Immunology and Pathology PR:LABORATORY Belisle Lab PR:LAST_NAME Nurul PR:FIRST_NAME Islam PR:ADDRESS 3185 Rampart Road, Fort Collins, CO 80521 PR:EMAIL nurul.islam@colostate.edu PR:PHONE 970 492 4249 PR:FUNDING_SOURCE NIH and the Office of the Assistant Secretary of Defense for Health Affairs PR:FUNDING_SOURCE through the Tick Borne Disease Research Program PR:PROJECT_COMMENTS N/A PR:PUBLICATIONS N/A PR:CONTRIBUTORS Gabriela Balderrama Gutierrez, Ana Milovic, Vanessa J. Cook, M. Nurul Islam, PR:CONTRIBUTORS Youwen Zhang, Hippokratis Kiaris, John T. Belisle, Ali Mortazavi and Alan G. PR:CONTRIBUTORS Barbour. #STUDY ST:STUDY_TITLE Metabolic profiling of plasma collected from Peromyscus leucopus and Mus ST:STUDY_TITLE musculus following LPS treatment ST:STUDY_TYPE Untargeted Metabolomic profiling ST:STUDY_SUMMARY Small molecule metabolites were extracted from plasma from Peromyscus leucopus ST:STUDY_SUMMARY and Mus musculus BALB/c and analyzed by liquid chromatography couple with mass ST:STUDY_SUMMARY spectrometry (LC-MS). XCMS software was used for peak picking, grouping and ST:STUDY_SUMMARY retention time correction from LC-MS data and generated a molecular feature ST:STUDY_SUMMARY spreadsheet. Pathway way enrichment analysis was carried out using Mummichog ST:STUDY_SUMMARY pathway analysis software using data from feature table. ST:INSTITUTE Colorado State University ST:DEPARTMENT Microbiology, Immunology and Pathology ST:LABORATORY Belisle Lab ST:LAST_NAME Nurul ST:FIRST_NAME Islam ST:ADDRESS 3185 Rampart Road, Fort Collins, CO 80521 ST:EMAIL nurul.islam@colostate.edu ST:PHONE 970 492 4249 ST:NUM_GROUPS 4 ST:TOTAL_SUBJECTS 40 ST:NUM_MALES 20 ST:NUM_FEMALES 20 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus;Peromyscus leucopus SU:TAXONOMY_ID 10090 SU:GENDER Male and female #FACTORS #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - Peromyscus_22275_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22275_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22277_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22277_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22282_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22282_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22279_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22279_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22321_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22321_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22335_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22335_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22341_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22341_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22315_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Peromyscus_22315_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22258_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22258_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22281_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22281_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22278_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22278_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22289_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22289_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22292_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22292_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22284_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22284_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22319_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22319_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22316_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22316_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22325_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22325_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22332_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22332_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22327_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22327_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Peromyscus_22339_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Peromyscus_22339_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_102_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_102_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_120_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_120_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_F101_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_F101_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_F302_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_F302_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_106_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_106_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_115_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_115_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_116_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_116_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_M404_Control_Plasma Sample Type:control (saline) RAW_FILE_NAME=Mus_M404_Control_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_101_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_101_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_103_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_103_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_104_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_104_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_105_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_105_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_118_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_118_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_F303_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_F303_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_107_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_107_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_108_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_108_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_109_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_109_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_M205_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_M205_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_M406_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_M406_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Mus_M407_LPS_Plasma Sample Type:lipopolysaccharide RAW_FILE_NAME=Mus_M407_LPS_Plasma.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC01 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC01.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC02 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC02.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC03 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC03.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC04 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC04.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC05 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC05.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC06 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC06.mzML SUBJECT_SAMPLE_FACTORS - Pooled_QC07 Sample Type:Pooled quality control sample RAW_FILE_NAME=Pooled_QC07.mzML #COLLECTION CO:COLLECTION_SUMMARY At 4.0 hr after LPS injection the animals were euthanized. After opening the CO:COLLECTION_SUMMARY chest, exsanguination was performed by cardiac puncture and blood was CO:COLLECTION_SUMMARY transferred to a heparin sulfate coated tubes (Becton-Dickinson Microtainer). CO:COLLECTION_SUMMARY Anticoagulated blood was centrifuged to pellet blood cells for 3 min at 4600 x g CO:COLLECTION_SUMMARY at 4° C. Plasma and blood pellet was kept separately at -80° C until further CO:COLLECTION_SUMMARY analysis. CO:SAMPLE_TYPE Blood (plasma) CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Animals were anesthetized with isoflurane and injected i.p. with a single dose TR:TREATMENT_SUMMARY of E. coli O111:B4 LPS at a concentration of 10 µg/gm body weight in a 50 µl TR:TREATMENT_SUMMARY volume as described above. The control group was anesthetized and then injected TR:TREATMENT_SUMMARY with the 0.9% saline alone. The experiment started at 0800 h with 10 min TR:TREATMENT_SUMMARY intervals between animals and with alteration of LPS and control injections. At TR:TREATMENT_SUMMARY 4.0 hr after their injection the animals were euthanized. After opening the TR:TREATMENT_SUMMARY chest, exsanguination was performed by cardiac puncture and blood was TR:TREATMENT_SUMMARY transferred to a heparin sulfate coated tubes (Becton-Dickinson Microtainer). TR:TREATMENT_SUMMARY Anticoagulated blood was centrifuged to pellet blood cells for 3 min at 4600 x g TR:TREATMENT_SUMMARY at 4° C. Plasma and blood pellet was kept separately at -80° C until further TR:TREATMENT_SUMMARY analysis. TR:TREATMENT_ROUTE i.p. TR:TREATMENT_DOSE 10 µg/gm body weight TR:TREATMENT_DOSEVOLUME 50 µl volume TR:TREATMENT_VEHICLE sterile, endotoxin-free 0.9% saline TR:ANIMAL_ANESTHESIA isoflurane TR:TREATMENT_COMPOUND E. coli O111:B4 LPS #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Metabolites were extracted from aliquots of 40 µL of plasma with 75% LC-MS SP:SAMPLEPREP_SUMMARY grade methanol, supernatants were dried, resuspended in 50% methanol, 10 µL SP:SAMPLEPREP_SUMMARY samples were subjected to liquid chromatography and mass spectrometry. Pooled SP:SAMPLEPREP_SUMMARY quality control sample was prepared by mixing small aliquot (10 µL) of each SP:SAMPLEPREP_SUMMARY plasma sample in the study. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Agilent 1260 CH:COLUMN_NAME Agilent Poroshell 120 EC-C8 (2.1 x 100mm, 2.7um) CH:FLOW_GRADIENT non-linear gradient CH:FLOW_RATE 0.25 mL/min CH:COLUMN_TEMPERATURE 50 CH:SOLVENT_A 0.1% formic acid in water CH:SOLVENT_B 0.1% formic acid in acetonitrile #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME Belisle Lab AN:OPERATOR_NAME Nurul Islam AN:DATA_FORMAT Agilent .d file #MS MS:INSTRUMENT_NAME Agilent 6520 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS An aliquot of sample was injected into LC-MS to acquire data in positive mode. MS:MS_COMMENTS Data acquisition and analysis were achieved using MassHunter software. MS:CAPILLARY_VOLTAGE 4000V MS:DRY_GAS_TEMP 300 degree C MS:FRAGMENT_VOLTAGE 120V MS:GAS_PRESSURE 45 MS:MS_RESULTS_FILE ST001654_AN002701_Results.txt UNITS:peak area Has m/z:Yes Has RT:Yes RT units:Seconds #END