#METABOLOMICS WORKBENCH bsmith8_20251017_071710 DATATRACK_ID:6566 STUDY_ID:ST004302 ANALYSIS_ID:AN007159 PROJECT_ID:PR002718
VERSION             	1
CREATED_ON             	October 21, 2025, 9:32 am
#PROJECT
PR:PROJECT_TITLE                 	Metabolomic analysis of wild-type and TauT-/- murine Mesenchymal Stromal Cells
PR:PROJECT_TITLE                 	(MSCs)
PR:PROJECT_SUMMARY               	Bone marrow mesenchymal stromal cells (BMSCs) are a type of multipotent stem
PR:PROJECT_SUMMARY               	cell found in the bone marrow stroma that can differentiate into various cell
PR:PROJECT_SUMMARY               	types, support the bone marrow's microenvironment, and regulate other stem
PR:PROJECT_SUMMARY               	cells. Their key functions include providing physical support to hematopoietic
PR:PROJECT_SUMMARY               	stem cells, secreting factors that control their homeostasis, and undergoing
PR:PROJECT_SUMMARY               	differentiation into bone, cartilage, and fat cells. BMSCs are also being
PR:PROJECT_SUMMARY               	explored for various cell therapies due to their ability to be expanded in
PR:PROJECT_SUMMARY               	vitro. To determine the role of taurine in regulating metabolism of bone marrow
PR:PROJECT_SUMMARY               	mesenchymal stromal cells (MSCs), we carried out untargeted metabolomic analysis
PR:PROJECT_SUMMARY               	of murine taurine transporter (TauT) wild-type and knockout MSCs. Briefly, 16
PR:PROJECT_SUMMARY               	week old TauT+/+ and TauT-/- mice were sacrificed, stromal populations were
PR:PROJECT_SUMMARY               	isolated and magnetically enriched or flow sorted for MSCs. Next, untargeted
PR:PROJECT_SUMMARY               	metabolomic analysis was carried out to identify metabolic changes induced by
PR:PROJECT_SUMMARY               	inhibiting taurine uptake in MSCs. This analysis identified inositol metabolism
PR:PROJECT_SUMMARY               	as a downstream effector of taurine in MSCs.
PR:INSTITUTE                     	University of Rochester Medical Center
PR:LAST_NAME                     	Smith
PR:FIRST_NAME                    	Bradley
PR:ADDRESS                       	575 Elmwood Ave, Rochester, NY, 14620
PR:EMAIL                         	bradley_smith@urmc.rochester.edu
PR:PHONE                         	585-275-1445
#STUDY
ST:STUDY_TITLE                   	Metabolomic analysis of wild-type and TauT-/- murine Mesenchymal Stromal Cells
ST:STUDY_TITLE                   	(MSCs)
ST:STUDY_SUMMARY                 	16 week old TauT+/+ and TauT-/- mice were sacrificed, stromal populations were
ST:STUDY_SUMMARY                 	isolated and magnetically enriched or flow sorted for MSCs (CD45-Ter119-). MSC
ST:STUDY_SUMMARY                 	cultures were washed with PBS and detached using TrypLE. Cells were washed with
ST:STUDY_SUMMARY                 	PBS containing 5 mM glucose and centrifuged at 3000g for 1 min, snap frozen, and
ST:STUDY_SUMMARY                 	processed for untargeted metabolomics.
ST:INSTITUTE                     	University of Rochester Medical Center
ST:LAST_NAME                     	Smith
ST:FIRST_NAME                    	Bradley
ST:ADDRESS                       	575 Elmwood Ave, Rochester, NY, 14620
ST:EMAIL                         	bradley_smith@urmc.rochester.edu
ST:PHONE                         	585-275-1445
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
#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           	-	KO_1a_7	Sample source:Mesenchymal Stromal Cells | Genotype:KO	RAW_FILE_NAME(Raw file name)=KO_1a_250214_7
SUBJECT_SAMPLE_FACTORS           	-	KO_1b_8	Sample source:Mesenchymal Stromal Cells | Genotype:KO	RAW_FILE_NAME(Raw file name)=KO_1b_250214_8
SUBJECT_SAMPLE_FACTORS           	-	KO_2a_3	Sample source:Mesenchymal Stromal Cells | Genotype:KO	RAW_FILE_NAME(Raw file name)=KO_2a_250214_3
SUBJECT_SAMPLE_FACTORS           	-	KO_2b_4	Sample source:Mesenchymal Stromal Cells | Genotype:KO	RAW_FILE_NAME(Raw file name)=KO_2b_250214_4
SUBJECT_SAMPLE_FACTORS           	-	KO_6a_10	Sample source:Mesenchymal Stromal Cells | Genotype:KO	RAW_FILE_NAME(Raw file name)=KO_6a_250214_10
SUBJECT_SAMPLE_FACTORS           	-	WT_1a_1	Sample source:Mesenchymal Stromal Cells | Genotype:WT	RAW_FILE_NAME(Raw file name)=WT_1a_250214_1
SUBJECT_SAMPLE_FACTORS           	-	WT_1b_2	Sample source:Mesenchymal Stromal Cells | Genotype:WT	RAW_FILE_NAME(Raw file name)=WT_1b_250214_2
SUBJECT_SAMPLE_FACTORS           	-	WT_2a_5	Sample source:Mesenchymal Stromal Cells | Genotype:WT	RAW_FILE_NAME(Raw file name)=WT_2a_250214_5
SUBJECT_SAMPLE_FACTORS           	-	WT_2b_6	Sample source:Mesenchymal Stromal Cells | Genotype:WT	RAW_FILE_NAME(Raw file name)=WT_2b_250214_6
SUBJECT_SAMPLE_FACTORS           	-	WT_6a_9	Sample source:Mesenchymal Stromal Cells | Genotype:WT	RAW_FILE_NAME(Raw file name)=WT_6a_250214_9
#COLLECTION
CO:COLLECTION_SUMMARY            	16 week old TauT+/+ and TauT-/- mice were sacrificed, stromal populations were
CO:COLLECTION_SUMMARY            	isolated and magnetically enriched or flow sorted for MSCs (CD45-Ter119-). MSC
CO:COLLECTION_SUMMARY            	cultures were washed with PBS and detached using TrypLE. Cells were washed with
CO:COLLECTION_SUMMARY            	PBS containing 5mM glucose and centrifuged at 3000g for 1 min, snap frozen, and
CO:COLLECTION_SUMMARY            	processed for untargeted metabolomics.
CO:SAMPLE_TYPE                   	Mesenchymal stromal cells
#TREATMENT
TR:TREATMENT_SUMMARY             	N/A
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Frozen cell pellets were resuspended at 1 million cells per 1 mL of 80% MeOH via
SP:SAMPLEPREP_SUMMARY            	vortexing, transferred to -80°C for 30 min and then regular ice for 30 minutes
SP:SAMPLEPREP_SUMMARY            	with vortexing every 10 minutes. Next, samples were centrifuged at 17,000x g for
SP:SAMPLEPREP_SUMMARY            	10 minutes and 90% of supernatant was dried down in a vacuum evaporator
SP:SAMPLEPREP_SUMMARY            	(Thermo). Samples were reconstituted in 50% acetonitrile (A955, Fisher
SP:SAMPLEPREP_SUMMARY            	Scientific), at a volume equal to 10% of dried down volume, and transferred to
SP:SAMPLEPREP_SUMMARY            	glass vials for LC/MS analysis.
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE           	HILIC
CH:INSTRUMENT_NAME               	Thermo Vanquish
CH:COLUMN_NAME                   	Waters XBridge XP BEH Amide (150 mm x 2.1 mm, 2.5 µm)
CH:SOLVENT_A                     	100% water; 10 mM ammonium formate; 0.125% formic acid
CH:SOLVENT_B                     	90% acetonitrile/10% water; 10 mM ammonium formate; 0.125% formic acid
CH:FLOW_GRADIENT                 	0 minutes, 100% B; 2 minutes, 100% B; 3 minutes, 90% B; 5 minutes, 90%
CH:FLOW_GRADIENT                 	B; 6 minutes, 85% B; 7 minutes, 85% B; 8 minutes, 75% B; 9 minutes, 75%
CH:FLOW_GRADIENT                 	B; 10 minutes, 55% B; 12 minutes, 55% B; 13 minutes, 35%, 20 minutes,
CH:FLOW_GRADIENT                 	35% B; 20.1 minutes, 35% B; 20.6 minutes, 100% B; 22.2 minutes, 100% B; 28
CH:FLOW_GRADIENT                 	minutes, 100% B.
CH:FLOW_RATE                     	150 μL/min for first 22.7 minutes, then 300 μL/min for 22.7 to 28 minutes
CH:COLUMN_TEMPERATURE            	25℃
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Thermo Orbitrap Exploris 240
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	The H-ESI source was operated in positive mode at spray voltage 3500 with the
MS:MS_COMMENTS                   	following parameters: sheath gas 35 au, aux gas 7 au, sweep gas 0 au, ion
MS:MS_COMMENTS                   	transfer tube temperature 320°C, vaporizer temperature 275°C, mass range 70 to
MS:MS_COMMENTS                   	1000 m/z, full scan MS1 mass resolution of 120,000 FWHM, RF lens at 70%, and
MS:MS_COMMENTS                   	standard automatic gain control (AGC). Data acquisition software: Thermo
MS:MS_COMMENTS                   	XCalibur Data processing and feature assignment software: Thermo Compound
MS:MS_COMMENTS                   	Discoverer
MS:MS_RESULTS_FILE               	ST004302_AN007159_Results.txt	UNITS:area	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END