Summary of Study ST001702

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001089. The data can be accessed directly via it's Project DOI: 10.21228/M8XT42 This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST001702
Study Title	A cross-sectional study of functional and metabolic changes during aging through the lifespan in male mice (Heart) part-IV
Study Summary	Aging is associated with distinct phenotypical, physiological, and functional changes, leading to the onset of disease and death. The progression of aging-related traits varies widely among individuals, influenced by their environment, lifestyle, and genetics. In this study, we performed physiologic and functional tests cross-sectionally throughout the entire lifespan of male C57BL/6N mice. In parallel, metabolomics analyses in serum, brain, liver, heart, and skeletal muscle were also performed to identify signatures associated with frailty and age-dependent functional decline. Our findings indicate that the decline in gait speed as a function of age and frailty is associated with dramatic increases in the energetic cost of physical activity and decreases in working capacity. Aging and functional decline prompt organs to rewire their substrate selection and metabolism towards redox-related pathways, mainly in liver and heart. Collectively, the data provide a framework to further understand and characterize processes of aging at the individual and organ levels.
Institute	National Institutes of Health
Department	NIA
Laboratory	Experimental Gerontology Section and Translational Gerontology Branch
Last Name	de Cabo
First Name	Rafael
Address	251 Bayview Blvd. Suite 100/Room 5C214. Baltimore, MD 21224
Email	deCaboRa@grc.nia.nih.gov
Phone	1-410-558-8510
Submit Date	2021-02-11
Raw Data Available	Yes
Raw Data File Type(s)	cdf
Analysis Type Detail	GC-MS
Release Date	2021-03-01
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR001089
Project DOI:	doi: 10.21228/M8XT42
Project Title:	A cross-sectional study of functional and metabolic changes during aging through the lifespan in male mice
Project Type:	Untargeted metabolomics
Project Summary:	Aging is associated with distinct phenotypical, physiological, and functional changes, leading to the onset of disease and death. The progression of aging-related traits varies widely among individuals, influenced by their environment, lifestyle, and genetics. In this study, we performed physiologic and functional tests cross-sectionally throughout the entire lifespan of male C57BL/6N mice. In parallel, metabolomics analyses in serum, brain, liver, heart, and skeletal muscle were also performed to identify signatures associated with frailty and age-dependent functional decline. Our findings indicate that the decline in gait speed as a function of age and frailty is associated with dramatic increases in the energetic cost of physical activity and decreases in working capacity. Aging and functional decline prompt organs to rewire their substrate selection and metabolism towards redox-related pathways, mainly in liver and heart. Collectively, the data provide a framework to further understand and characterize processes of aging at the individual and organ levels.
Institute:	National Institutes of Health
Department:	Experimental Gerontology Section and Translational Gerontology Branch
Laboratory:	NIA
Last Name:	de Cabo
First Name:	Rafael
Address:	251 Bayview Blvd. Suite 100/Room 5C214. Baltimore, MD 21224
Email:	deCaboRa@grc.nia.nih.gov
Phone:	1-410-558-8510
Funding Source:	Intramural Research Program of the National Institute on Aging, NIH

Subject:

Subject ID:	SU001779
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id	local_sample_id	Age
SA158274	14_heart_014	mid age
SA158275	30_heart_030	mid age
SA158276	19_heart_019	mid age
SA158277	4_heart_004	mid age
SA158278	5_heart_005	mid age
SA158279	10_heart_010	mid age
SA158280	12_heart_012	mid age
SA158281	11_heart_011	old
SA158282	15_heart_015	old
SA158283	29_heart_029	old
SA158284	28_heart_028	old
SA158285	20_heart_020	old
SA158286	6_heart_006	old
SA158287	3_heart_003	old
SA158288	2_heart_002	old
SA158289	8_heart_008	old
SA158290	9_heart_009	old
SA158291	27_heart_027	old
SA158292	23_heart_023	young
SA158293	24_heart_024	young
SA158294	25_heart_025	young
SA158295	22_heart_022	young
SA158296	16_heart_016	young
SA158297	13_heart_013	young
SA158298	26_heart_026	young
SA158299	18_heart_018	young
SA158300	1_heart_001	young
SA158301	21_heart_021	young
SA158302	17_heart_017	young
SA158303	7_heart_007	young

Showing results 1 to 30 of 30

Showing results 1 to 30 of 30

Collection:

Collection ID:	CO001772
Collection Summary:	Tissues were collected from perfused mouse and flash frozen in liquid nitrogen. Samples stored at -80 degrees.
Sample Type:	Heart

Treatment:

Treatment ID:	TR001792
Treatment Summary:	Healthy mice are divided based on their ages. 3 groups are presented: - Young (n=12): <15 mo-old - Mid (n=7): 15 to 20 mo-old - Old (n=11): >20 mo-old

Sample Preparation:

Sampleprep ID:	SP001785
Sampleprep Summary:	Extraction of Mammalian Tissue Samples: Lungs/Muscle/Heart 1. References: Fiehn O, Kind T (2006) Metabolite profiling in blood plasma. In: Metabolomics: Methods and Protocols. Weckwerth W (ed.), Humana Press, Totowa NJ (in press) 2.Starting material: Mammalian tissue: Lung/Muscle/Heart: Whole tissue sample is prepared OR stein mill whole tissue sample and weigh 50mL aliquot. 3. Equipment: Centrifuge (Eppendorf 5415 D) Calibrated pipettes 1-200?l and 100-1000?l Eppendorf tubes 2ml, uncoloured (Cat. No. 022363204) Centrifuge tubes, various sizes, polypropylene Eppendorff Tabletop Centrifuge (Proteomics core Lab.) ThermoElectron Neslab RTE 740 cooling bath at –20°C MiniVortexer (VWR) Orbital Mixing Chilling/Heating Plate (Torrey Pines Scientific Instruments) Speed vacuum concentration system (Labconco Centrivap cold trap) Turex mini homogenizer 4. Chemicals Acetonitrile, LCMS grade (JT Baker; Cat. No.9829-02) Isopropanol, HPLC grade (JT Baker; Cat. No. 9095-02) Crushed ice pH paper 5-10 (EMD Chem. Inc.) Nitrogen line with pipette tip 18 M? pure water (Millipore) 5. Procedure Preparation of extraction mix and material before experiment: Switch on bath to pre-cool at –20°C (±2°C validity temperature range) Check pH of acetonitrile and isopropanol (pH7) using wetted pH paper Make the extraction solution by missing acetonitrile, isopropanol and water in proportions 3 : 3 : 2 Rinse the extraction solution for 5 min with nitrogen. Make sure that the nitrogen line was flushed out of air before using it for degassing the extraction solvent solution Sample Preparation Weigh 50 mg tissue sample in to a 25 ml conical polypropylene centrifuge tube. Add 2.5mL 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. Centrifuge the samples at 2500 rpm. for 5 minutes. Aliquot 2 X 500µl supernatant, one for analysis and one for a backup sample. Store backup aliquot in the -20°C freezer. Evaporate one 500µl aliquot of the sample in the Labconco Centrivap cold trap concentrator to complete dryness The dried aliquot is then re-suspended with 500?l 50% acetonitrile (degassed as given) Centrifuge for 2 min at 14000 rcf using the centrifuge Eppendorf 5415. Remove supernatant to a new Eppendorff tube. Evaporate the supernatant to dryness in the the Labconco Centrivap cold trap concentrator. Submit to derivatization. The residue should contain membrane lipids because these are supposedly not soluble enough to be found in the 50% acetonitrile solution. Therefore, this ‘membrane residue’ is now taken for membrane lipidomic fingerprinting using the nanomate LTQ ion trap mass spectrometer. Likely, a good solvent to redissolve the membrane lipids is e.g. 75% isopropanol (degassed as given above). If the ‘analysis’ aliquot is to be used for semi lipophilic compounds such as tyrosine pathway intermediates (incl. dopamine, serotonine etc, i.e. polar aromatic compounds), then these are supposedly to be found together with the ‘GCTOF’ aliquot. We can assume that this mixture is still too complex for Agilent chipLCMS. Therefore, in order to develop and validate target analysis for such aromatic compounds, we should use some sort of Solid Phase purification. We re-suspend the dried ‘GCTOF’ aliquot in 300 ?l water (degassed as before) to take out sugars, aliphatic amino acids, hydroxyl acids and similar logP compounds. The residue should contain our target aromatics .We could also try to adjust pH by using low concentration acetate or phosphate buffer. The residue could then be taken up in 50% acetonitrile and used for GCTOF and Agilent chipMS experiments. The other aliquot should be checked how much of our target compounds would actually be found in the ‘sugar’ fraction. 6. Problems To prevent contamination disposable material is used. Control pH from extraction mix. 7. Quality assurance For each sequence of sample extractions, perform one blank negative control extraction by applying the total procedure (i.e. all materials and plastic ware) without biological sample. 8. Disposal of waste Collect all chemicals in appropriate bottles and follow the disposal rules.

Sampleprep ID:

SP001785

Sampleprep Summary:

Extraction of Mammalian Tissue Samples: Lungs/Muscle/Heart 1. References: Fiehn O, Kind T (2006) Metabolite profiling in blood plasma. In: Metabolomics: Methods and Protocols. Weckwerth W (ed.), Humana Press, Totowa NJ (in press) 2.Starting material: Mammalian tissue: Lung/Muscle/Heart: Whole tissue sample is prepared OR stein mill whole tissue sample and weigh 50mL aliquot. 3. Equipment: Centrifuge (Eppendorf 5415 D) Calibrated pipettes 1-200?l and 100-1000?l Eppendorf tubes 2ml, uncoloured (Cat. No. 022363204) Centrifuge tubes, various sizes, polypropylene Eppendorff Tabletop Centrifuge (Proteomics core Lab.) ThermoElectron Neslab RTE 740 cooling bath at –20°C MiniVortexer (VWR) Orbital Mixing Chilling/Heating Plate (Torrey Pines Scientific Instruments) Speed vacuum concentration system (Labconco Centrivap cold trap) Turex mini homogenizer 4. Chemicals Acetonitrile, LCMS grade (JT Baker; Cat. No.9829-02) Isopropanol, HPLC grade (JT Baker; Cat. No. 9095-02) Crushed ice pH paper 5-10 (EMD Chem. Inc.) Nitrogen line with pipette tip 18 M? pure water (Millipore) 5. Procedure Preparation of extraction mix and material before experiment: Switch on bath to pre-cool at –20°C (±2°C validity temperature range) Check pH of acetonitrile and isopropanol (pH7) using wetted pH paper Make the extraction solution by missing acetonitrile, isopropanol and water in proportions 3 : 3 : 2 Rinse the extraction solution for 5 min with nitrogen. Make sure that the nitrogen line was flushed out of air before using it for degassing the extraction solvent solution Sample Preparation Weigh 50 mg tissue sample in to a 25 ml conical polypropylene centrifuge tube. Add 2.5mL 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. Centrifuge the samples at 2500 rpm. for 5 minutes. Aliquot 2 X 500µl supernatant, one for analysis and one for a backup sample. Store backup aliquot in the -20°C freezer. Evaporate one 500µl aliquot of the sample in the Labconco Centrivap cold trap concentrator to complete dryness The dried aliquot is then re-suspended with 500?l 50% acetonitrile (degassed as given) Centrifuge for 2 min at 14000 rcf using the centrifuge Eppendorf 5415. Remove supernatant to a new Eppendorff tube. Evaporate the supernatant to dryness in the the Labconco Centrivap cold trap concentrator. Submit to derivatization. The residue should contain membrane lipids because these are supposedly not soluble enough to be found in the 50% acetonitrile solution. Therefore, this ‘membrane residue’ is now taken for membrane lipidomic fingerprinting using the nanomate LTQ ion trap mass spectrometer. Likely, a good solvent to redissolve the membrane lipids is e.g. 75% isopropanol (degassed as given above). If the ‘analysis’ aliquot is to be used for semi lipophilic compounds such as tyrosine pathway intermediates (incl. dopamine, serotonine etc, i.e. polar aromatic compounds), then these are supposedly to be found together with the ‘GCTOF’ aliquot. We can assume that this mixture is still too complex for Agilent chipLCMS. Therefore, in order to develop and validate target analysis for such aromatic compounds, we should use some sort of Solid Phase purification. We re-suspend the dried ‘GCTOF’ aliquot in 300 ?l water (degassed as before) to take out sugars, aliphatic amino acids, hydroxyl acids and similar logP compounds. The residue should contain our target aromatics .We could also try to adjust pH by using low concentration acetate or phosphate buffer. The residue could then be taken up in 50% acetonitrile and used for GCTOF and Agilent chipMS experiments. The other aliquot should be checked how much of our target compounds would actually be found in the ‘sugar’ fraction. 6. Problems To prevent contamination disposable material is used. Control pH from extraction mix. 7. Quality assurance For each sequence of sample extractions, perform one blank negative control extraction by applying the total procedure (i.e. all materials and plastic ware) without biological sample. 8. Disposal of waste Collect all chemicals in appropriate bottles and follow the disposal rules.

Combined analysis:

Analysis ID	AN002773
Analysis type	MS
Chromatography type	GC
Chromatography system	Leco Pegasus IV GC
Column	Restek Rtx-5Sil (30m x 0.25mm,0.25um)
MS Type	EI
MS instrument type	GC-TOF
MS instrument name	Leco Pegasus IV TOF
Ion Mode	UNSPECIFIED
Units	normalized peak height

Chromatography:

Chromatography ID:	CH002053
Chromatography Summary:	Primary metabolism by GCTOF
Instrument Name:	Leco Pegasus IV GC
Column Name:	Restek Rtx-5Sil (30m x 0.25mm,0.25um)
Chromatography Type:	GC

MS:

MS ID:	MS002570
Analysis ID:	AN002773
Instrument Name:	Leco Pegasus IV TOF
Instrument Type:	GC-TOF
MS Type:	EI
MS Comments:	A Leco Pegasus IV time of flight mass spectrometer is controlled by the Leco ChromaTOF software vs. 2.32 (St. Joseph, MI). The transfer line temperature between gas chromatograph and mass spectrometer is set to 280°C. Electron impact ionization at 70V is employed with an ion source temperature of 250°C. Acquisition rate is 17 spectra/second, with a scan mass range of 85-500 Da.
Ion Mode:	UNSPECIFIED