#METABOLOMICS WORKBENCH AnnalauraM_20250521_083407 DATATRACK_ID:5932 STUDY_ID:ST003928 ANALYSIS_ID:AN006451 PROJECT_ID:PR002461
VERSION             	1
CREATED_ON             	May 27, 2025, 1:49 am
#PROJECT
PR:PROJECT_TITLE                 	Imidazole propionate is a driver and therapeutic target in atherosclerosis
PR:PROJECT_TYPE                  	Original research
PR:PROJECT_SUMMARY               	Atherosclerosis (AT) is the main underlying cause of cardiovascular diseases
PR:PROJECT_SUMMARY               	(CVDs). Its prevention is based on the detection and treatment of traditional
PR:PROJECT_SUMMARY               	cardiovascular risk factors (PMID:34120177) but often fails to identify
PR:PROJECT_SUMMARY               	individuals at risk for early vascular disease (PMID:25882487). Recent research
PR:PROJECT_SUMMARY               	has suggested new players in the pathophysiology of atherosclerosis (PMID:
PR:PROJECT_SUMMARY               	33883728), highlighting the need for alternative disease biomarkers and
PR:PROJECT_SUMMARY               	therapeutic targets to improve early diagnosis and therapy efficacy. Here, we
PR:PROJECT_SUMMARY               	identified that microbially produced imidazole propionate (ImP) is associated
PR:PROJECT_SUMMARY               	with the extent of atherosclerosis in mice and in two independent human cohorts.
PR:PROJECT_SUMMARY               	Furthermore, ImP administration to atherosclerosis-prone mice fed chow diet was
PR:PROJECT_SUMMARY               	sufficient to induce atherosclerosis without altering the lipid profile, and it
PR:PROJECT_SUMMARY               	was linked to activation of both systemic and local innate and adaptive immunity
PR:PROJECT_SUMMARY               	and inflammation. Specifically, we found that ImP caused atherosclerosis through
PR:PROJECT_SUMMARY               	Imidazoline-1 receptor (I1R) expressed in myeloid cells. Blocking this ImP/I1R
PR:PROJECT_SUMMARY               	axis inhibited the development of atherosclerosis induced by ImP as well as by
PR:PROJECT_SUMMARY               	high cholesterol diet in mice. Identification of the strong association of ImP
PR:PROJECT_SUMMARY               	with active atherosclerosis, along with the discovery of the contribution of the
PR:PROJECT_SUMMARY               	ImP/I1R axis to disease progression opens new avenues for improving the early
PR:PROJECT_SUMMARY               	diagnosis and personalized therapy of atherosclerosis.
PR:INSTITUTE                     	Centro Nacional de Investigaciones Cardiovasculares Carlos III
PR:LAST_NAME                     	Mastrangelo
PR:FIRST_NAME                    	Annalaura
PR:ADDRESS                       	Calle de Melchor Fernández Almagro, 3 – 28029, Madrid (Spain)
PR:EMAIL                         	amastrangelo@cnic.es
PR:PHONE                         	(+34) 914531202
#STUDY
ST:STUDY_TITLE                   	Untargeted metabolomics links imidazole propionate and atherosclerosis
ST:STUDY_SUMMARY                 	Atherosclerosis (AT) is the primary underlying cause of cardiovascular diseases
ST:STUDY_SUMMARY                 	(CVDs). Current preventive strategies rely on the detection and management of
ST:STUDY_SUMMARY                 	traditional cardiovascular risk factors (PMID: 34120177), yet they often fail to
ST:STUDY_SUMMARY                 	identify individuals at risk for early-stage vascular disease (PMID: 25882487).
ST:STUDY_SUMMARY                 	Emerging research has revealed novel contributors to the pathophysiology of
ST:STUDY_SUMMARY                 	atherosclerosis (PMID: 33883728), underscoring the need for alternative
ST:STUDY_SUMMARY                 	biomarkers and therapeutic targets to enhance early diagnosis and treatment
ST:STUDY_SUMMARY                 	efficacy. In this study, we identified the microbial metabolite imidazole
ST:STUDY_SUMMARY                 	propionate (ImP) as being strongly associated with atherosclerosis in both mice
ST:STUDY_SUMMARY                 	and two independent human cohorts. To uncover microbial metabolites that
ST:STUDY_SUMMARY                 	influence atherosclerosis progression, we employed an untargeted metabolomics
ST:STUDY_SUMMARY                 	approach in atherosclerosis-prone ApoE⁻/⁻ mice subjected to different diets,
ST:STUDY_SUMMARY                 	with or without antibiotic treatment to deplete the gut microbiota. This
ST:STUDY_SUMMARY                 	analysis revealed substantial remodeling of the plasma metabolome in response to
ST:STUDY_SUMMARY                 	dietary and microbiota changes. Among the differentially abundant metabolites,
ST:STUDY_SUMMARY                 	ImP emerged as a microbiota-dependent molecule closely linked to the extent of
ST:STUDY_SUMMARY                 	atherosclerosis.
ST:INSTITUTE                     	Centro Nacional de Investigaciones Cardiovasculares Carlos III
ST:LAST_NAME                     	Mastrangelo
ST:FIRST_NAME                    	Annalaura
ST:ADDRESS                       	Calle de Melchor Fernández Almagro, 3 – 28029, Madrid (Spain)
ST:EMAIL                         	a.mastrangelo@cnic.es
ST:PHONE                         	(+34) 914531202
#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           	26C	H_P_26C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_26C.mzML
SUBJECT_SAMPLE_FACTORS           	27C	H_P_27C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_27C.mzML
SUBJECT_SAMPLE_FACTORS           	28C	H_P_28C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_28C.mzML
SUBJECT_SAMPLE_FACTORS           	29C	H_P_29C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_29C.mzML
SUBJECT_SAMPLE_FACTORS           	30C	H_P_30C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_30C.mzML
SUBJECT_SAMPLE_FACTORS           	66C	H_P2_66C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_66C.mzML
SUBJECT_SAMPLE_FACTORS           	67C	H_P2_67C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_67C.mzML
SUBJECT_SAMPLE_FACTORS           	68C	H_P2_68C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_68C.mzML
SUBJECT_SAMPLE_FACTORS           	70C	H_P2_70C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_70C.mzML
SUBJECT_SAMPLE_FACTORS           	61C	H_P2_61C	Abx:Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_61C.mzML
SUBJECT_SAMPLE_FACTORS           	10C	H_P_10C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_10C.mzML
SUBJECT_SAMPLE_FACTORS           	6C	H_P_6C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_6C.mzML
SUBJECT_SAMPLE_FACTORS           	7C	H_P_7C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_7C.mzML
SUBJECT_SAMPLE_FACTORS           	8C	H_P_8C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_8C.mzML
SUBJECT_SAMPLE_FACTORS           	9C	H_P_9C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_9C.mzML
SUBJECT_SAMPLE_FACTORS           	46C	H_P2_46C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_46C.mzML
SUBJECT_SAMPLE_FACTORS           	47C	H_P2_47C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_47C.mzML
SUBJECT_SAMPLE_FACTORS           	48C	H_P2_48C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_48C.mzML
SUBJECT_SAMPLE_FACTORS           	49C	H_P2_49C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_49C.mzML
SUBJECT_SAMPLE_FACTORS           	50C	H_P2_50C	Abx:Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_50C.mzML
SUBJECT_SAMPLE_FACTORS           	16C	H_P_16C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_16C.mzML
SUBJECT_SAMPLE_FACTORS           	17C	H_P_17C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_17C.mzML
SUBJECT_SAMPLE_FACTORS           	18C	H_P_18C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_18C.mzML
SUBJECT_SAMPLE_FACTORS           	19C	H_P_19C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_19C.mzML
SUBJECT_SAMPLE_FACTORS           	20C	H_P_20C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_20C.mzML
SUBJECT_SAMPLE_FACTORS           	59C	H_2P_59C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_2P_59C.mzML
SUBJECT_SAMPLE_FACTORS           	56C	H_P2_56C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_56C.mzML
SUBJECT_SAMPLE_FACTORS           	57C	H_P2_57C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_57C.mzML
SUBJECT_SAMPLE_FACTORS           	58C	H_P2_58C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_58C.mzML
SUBJECT_SAMPLE_FACTORS           	60C	H_P2_60C	Abx:Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_60C.mzML
SUBJECT_SAMPLE_FACTORS           	21C	H_P_21C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_21C.mzML
SUBJECT_SAMPLE_FACTORS           	22C	H_P_22C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_22C.mzML
SUBJECT_SAMPLE_FACTORS           	23C	H_P_23C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_23C.mzML
SUBJECT_SAMPLE_FACTORS           	24C	H_P_24C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_24C.mzML
SUBJECT_SAMPLE_FACTORS           	25C	H_P_25C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_25C.mzML
SUBJECT_SAMPLE_FACTORS           	69C	H_P2_69C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_69C.mzML
SUBJECT_SAMPLE_FACTORS           	62C	H_P2_62C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_62C.mzML
SUBJECT_SAMPLE_FACTORS           	63C	H_P2_63C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_63C.mzML
SUBJECT_SAMPLE_FACTORS           	64C	H_P2_64C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_64C.mzML
SUBJECT_SAMPLE_FACTORS           	65C	H_P2_65C	Abx:No Abx | Diet:HCHC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_65C.mzML
SUBJECT_SAMPLE_FACTORS           	11C	H_P_11C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_11C.mzML
SUBJECT_SAMPLE_FACTORS           	12C	H_P_12C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_12C.mzML
SUBJECT_SAMPLE_FACTORS           	13C	H_P_13C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_13C.mzML
SUBJECT_SAMPLE_FACTORS           	14C	H_P_14C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_14C.mzML
SUBJECT_SAMPLE_FACTORS           	15C	H_P_15C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_15C.mzML
SUBJECT_SAMPLE_FACTORS           	51C	H_P2_51C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_51C.mzML
SUBJECT_SAMPLE_FACTORS           	52C	H_P2_52C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_52C.mzML
SUBJECT_SAMPLE_FACTORS           	53C	H_P2_53C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_53C.mzML
SUBJECT_SAMPLE_FACTORS           	54C	H_P2_54C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_54C.mzML
SUBJECT_SAMPLE_FACTORS           	55C	H_P2_55C	Abx:No Abx | Diet:HC | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_55C.mzML
SUBJECT_SAMPLE_FACTORS           	1C	H_P_1C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_1C.mzML
SUBJECT_SAMPLE_FACTORS           	2C	H_P_2C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_2C.mzML
SUBJECT_SAMPLE_FACTORS           	3C	H_P_3C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_3C.mzML
SUBJECT_SAMPLE_FACTORS           	4C	H_P_4C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_4C.mzML
SUBJECT_SAMPLE_FACTORS           	5C	H_P_5C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P_5C.mzML
SUBJECT_SAMPLE_FACTORS           	41C	H_P2_41C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_41C.mzML
SUBJECT_SAMPLE_FACTORS           	42C	H_P2_42C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_42C.mzML
SUBJECT_SAMPLE_FACTORS           	43C	H_P2_43C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_43C.mzML
SUBJECT_SAMPLE_FACTORS           	44C	H_P2_44C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_44C.mzML
SUBJECT_SAMPLE_FACTORS           	45C	H_P2_45C	Abx:No Abx | Diet:Chow | Sample source:plasma	RAW_FILE_NAME(mzML file name)=H_P2_45C.mzML
#COLLECTION
CO:COLLECTION_SUMMARY            	ApoE-/- mice were fed different diets (chow, high cholesterol (HC), HC and high
CO:COLLECTION_SUMMARY            	choline (HC/HC)) for 8 weeks. At 4 weeks post diet initiation, mice were treated
CO:COLLECTION_SUMMARY            	or not with a cocktail of antibiotics (abx) in the drinking water. At sacrifice,
CO:COLLECTION_SUMMARY            	aorta, heart, cecal content and plasma samples were collected in EDTA-K tubes
CO:COLLECTION_SUMMARY            	and analyzed.
CO:SAMPLE_TYPE                   	Blood (plasma)
#TREATMENT
TR:TREATMENT_SUMMARY             	Plasma samples were prepared from whole blood collected in EDTA-K tubes. After
TR:TREATMENT_SUMMARY             	gentle inversion to mix with the anticoagulant, samples were kept on ice and
TR:TREATMENT_SUMMARY             	processed within 30 minutes to minimize metabolic alterations. Blood was
TR:TREATMENT_SUMMARY             	centrifuged at 1,500–2,000 × g for 10–15 minutes at 4°C to separate
TR:TREATMENT_SUMMARY             	plasma, which was carefully collected without disturbing the buffy coat and
TR:TREATMENT_SUMMARY             	stored in aliquots at –80°C until analysis.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Plasma samples (50 µL) were prepared as previously (PMID: 27163744). Briefly,
SP:SAMPLEPREP_SUMMARY            	proteins were removed by adding a cold (-20°C) mixture of Methanol:Ethanol
SP:SAMPLEPREP_SUMMARY            	(1:1, v/v) in a ratio 5:1 (solvent:sample) and by storing the samples on ice for
SP:SAMPLEPREP_SUMMARY            	20 minutes. The samples were then centrifuged at 15700 g for 20 minutes at 4 °C
SP:SAMPLEPREP_SUMMARY            	and the supernatant was collected and placed in a LC vial for the subsequent
SP:SAMPLEPREP_SUMMARY            	analysis.
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE           	HILIC
CH:INSTRUMENT_NAME               	Thermo Dionex Ultimate 3000
CH:COLUMN_NAME                   	Merck SeQuant ZIC-HILIC column (150 × 1 mm, 3.5 µm)
CH:SOLVENT_A                     	100% Water; 0.01% formic acid
CH:SOLVENT_B                     	100% Acetonitrile; 0.01% formic acid
CH:FLOW_GRADIENT                 	gradient started from 90 % to 25 % of B in 15 min, keeping constant for 3 min
CH:FLOW_GRADIENT                 	and returned to starting conditions in 0.1 min, finally by keeping the
CH:FLOW_GRADIENT                 	re-equilibration at 90 % of B for 11.9 min.
CH:FLOW_RATE                     	180 µL/min
CH:COLUMN_TEMPERATURE            	45℃
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	LTQ Orbitrap XL™ Hybrid Ion Trap-Orbitrap Mass Spectrometer
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	MS was operating in full scan mode from 70 to 1000 m/z at 60000 resolution.
MS:MS_COMMENTS                   	MS/MS spectra were collected in data-dependent mode via collision induced
MS:MS_COMMENTS                   	dissociation (CID) in the ion trap. Samples were analysed in a randomized order.
MS:MS_COMMENTS                   	Generated data were firstly aligned using Compound Discoverer (ThermoFisher
MS:MS_COMMENTS                   	Scientific); signals were extracted and grouped into features (isotopic traces
MS:MS_COMMENTS                   	from a single analyte at a particular charge state) using the Metaboprofiler
MS:MS_COMMENTS                   	node in Compound discoverer (the open source plug-in freely available from
MS:MS_COMMENTS                   	OpenMS, http://www.openms.de/downloads/).
MS:MS_RESULTS_FILE               	ST003928_AN006451_Results.txt	UNITS:Relative abundance	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END