Summary of study ST001526

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 PR001027. The data can be accessed directly via it's Project DOI: 10.21228/M8Z41B 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.

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Study IDST001526
Study TitleMitochondrial health is enhanced in rats with higher vs. lower intrinsic exercise capacity and extended lifespan
Study SummaryThe intrinsic aerobic capacity of an organism is thought to play a role in aging and longevity. Maximal respiratory rate capacity, a metabolic performance measure, is one of the best predictors of cardiovascular- and all-cause mortality. Rats selectively bred for high-(HCR) vs. low-(LCR) intrinsic running-endurance capacity have up to 31% longer lifespan. We found that positive changes in indices of mitochondrial health in cardiomyocytes (respiratory reserve, maximal respiratory capacity, resistance to mitochondrial permeability transition, autophagy/mitophagy, higher lipids-over-glucose utilization) are uniformly associated with the extended longevity in HCR vs. LCR female rats. Cross-sectional heart metabolomics revealed pathways from lipid metabolism in the heart which were significantly enriched by a select group of strain dependent metabolites, consistent with enhanced lipids utilization by HCR cardiomyocytes. Heart-liver-serum metabolomics further revealed shunting of lipidic substrates between liver and heart via serum during aging. Thus, mitochondrial health in cardiomyocytes is associated with extended longevity in rats with higher intrinsic exercise capacity, and likely these findings can be translated to other populations as predictors of outcomes of health and survival.
Institute
National Institute on Aging
DepartmentCardioprotection Section
LaboratoryLaboratory of Cardiovascular Science
Last NameSollott
First NameSteven
AddressLaboratory of Cardiovascular Science, National Institute on Aging, NIH, Baltimore, MD 21224, USA
Emailsollotts@grc.nia.nih.gov
Phone410-558-8657
Submit Date2020-10-23
Raw Data AvailableYes
Raw Data File Type(s).cdf
Analysis Type DetailGC-MS
Release Date2020-12-01
Release Version1
Steven Sollott Steven Sollott
https://dx.doi.org/10.21228/M8Z41B
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR001027
Project DOI:doi: 10.21228/M8Z41B
Project Title:Mitochondrial health is enhanced in rats with higher vs. lower intrinsic exercise capacity and extended lifespan
Project Type:Untargeted analysis of primary metabolism by GCTOF
Project Summary:The intrinsic aerobic capacity of an organism is thought to play a role in aging and longevity. Maximal respiratory rate capacity, a metabolic performance measure, is one of the best predictors of cardiovascular- and all-cause mortality. Rats selectively bred for high-(HCR) vs. low-(LCR) intrinsic running-endurance capacity have up to 31% longer lifespan. We found that positive changes in indices of mitochondrial health in cardiomyocytes (respiratory reserve, maximal respiratory capacity, resistance to mitochondrial permeability transition, autophagy/mitophagy, higher lipids-over-glucose utilization) are uniformly associated with the extended longevity in HCR vs. LCR female rats. Cross-sectional heart metabolomics revealed pathways from lipid metabolism in the heart which were significantly enriched by a select group of strain dependent metabolites, consistent with enhanced lipids utilization by HCR cardiomyocytes. Heart-liver-serum metabolomics further revealed shunting of lipidic substrates between liver and heart via serum during aging. Thus, mitochondrial health in cardiomyocytes is associated with extended longevity in rats with higher intrinsic exercise capacity, and likely these findings can be translated to other populations as predictors of outcomes of health and survival.
Institute:National Institute on Aging
Department:Cardioprotection Section
Laboratory:Laboratory of Cardiovascular Science
Last Name:Sollott
First Name:Steven
Address:Laboratory of Cardiovascular Science, National Institute on Aging, NIH, Baltimore, MD 21224, USA
Email:sollotts@grc.nia.nih.gov
Phone:410-558-8657
Funding Source:Intramural Research Program of the National Institutes of Health, National Institute on Aging.

Subject:

Subject ID:SU001600
Subject Type:Mammal
Subject Species:Rattus norvegicus
Taxonomy ID:10116

Factors:

Subject type: Mammal; Subject species: Rattus norvegicus (Factor headings shown in green)

mb_sample_id local_sample_id treatment Tissue
SA1286041-H_027control heart
SA1286052-H_028control heart
SA1286066-H_030control heart
SA12860725-H_038control heart
SA1286085-H_029control heart
SA12860926-H_039control heart
SA12861014-H_032control heart
SA12861124-H_037control heart
SA12861221-H_034control heart
SA12861315-H_033control heart
SA12861422-H_035control heart
SA12861513-H_031control heart
SA12861623-H_036control heart
SA12861726-L_024control liver
SA12861827-L_025control liver
SA12861924-L_022control liver
SA12862028-L_026control liver
SA12862125-L_023control liver
SA1286224-L_004control liver
SA1286236-L_006control liver
SA12862423-L_021control liver
SA1286255-L_005control liver
SA1286263-L_003control liver
SA1286272-L_002control liver
SA1286281-L_001control liver
SA12862921-L_019control liver
SA12863013-L_011control liver
SA12863114-L_012control liver
SA12863211-L_009control liver
SA12863310-L_008control liver
SA12863422-L_020control liver
SA12863515-L_013control liver
SA12863612-L_010control liver
SA12863716-L_014control liver
SA12863820-L_018control liver
SA12863919-L_017control liver
SA12864018-L_016control liver
SA12864117-L_015control liver
SA1286429-L_007control liver
SA128597_007control serum
SA128598_010control serum
SA128601_009control serum
SA128602_008control serum
SA128643Rat 1_040control serum
SA128644MA-9_062control serum
SA128645MA-8_061control serum
SA128646Rat 2_041control serum
SA128647Rat 5_044control serum
SA128648MA-7_060control serum
SA128649Rat 6_045control serum
SA128650Rat 4_043control serum
SA128651Rat 3_042control serum
SA128652HC-6_051control serum
SA128653HC-5_050control serum
SA128654HC-4_049control serum
SA128655C2-HC-2_047control serum
SA128656C2-HC-1_046control serum
SA128657HC-7_052control serum
SA128658HC-8_053control serum
SA128659MA-12_065control serum
SA128660MA-11_064control serum
SA128661MA-10_063control serum
SA128662MA-5_058control serum
SA128478Heart HCR18730_040HCR heart
SA128479HEART_HCR18895_016HCR heart
SA128480HEART_HCR18927_017HCR heart
SA128481HEART_HCR18947_018HCR heart
SA128482HEART_HCR18871_015HCR heart
SA128483HEART_HCR19139_061HCR heart
SA128484HEART_HCR19157_058HCR heart
SA128485HEART_HCR19158_059HCR heart
SA128486HEART_HCR18866_013HCR heart
SA128487Heart HCR18659_018HCR heart
SA128488HEART_HCR18883_014HCR heart
SA128489HEART_HCR19153_060HCR heart
SA128490HEART_HCR19143_063HCR heart
SA128491Heart HCR18731_014HCR heart
SA128492Heart HCR18730_015HCR heart
SA128493Heart HCR18698_017HCR heart
SA128494HEART_HCR19140_062HCR heart
SA128495Heart HCR18729_016HCR heart
SA128496LIVER_HCR18927_005HCR liver
SA128497LIVER_HCR18947_006HCR liver
SA128498LIVER_HCR19139_049HCR liver
SA128499LIVER_HCR19140_050HCR liver
SA128500LIVER_HCR18895_004HCR liver
SA128501LIVER_HCR18883_002HCR liver
SA128502Liver HCR18731_038HCR liver
SA128503LIVER_HCR18866_001HCR liver
SA128504LIVER_HCR18871_003HCR liver
SA128505LIVER_HCR19143_051HCR liver
SA128506LIVER_HCR19153_048HCR liver
SA128507_002HCR liver
SA128508_005HCR liver
SA128509_004HCR liver
SA128510Liver HCR18698_008HCR liver
SA128511Liver HCR18729_007HCR liver
SA128512Liver HCR18730_006HCR liver
SA128513LIVER_HCR19157_046HCR liver
SA128514LIVER_HCR19158_047HCR liver
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Collection:

Collection ID:CO001595
Collection Summary:Samples were flash-frozen.
Sample Type:Blood;Liver;Heart

Treatment:

Treatment ID:TR001615
Treatment Summary:Information not provided.

Sample Preparation:

Sampleprep ID:SP001608
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 1-200μl and 100-1000μl 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 MΩ 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 500μl 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. Sample preparation of blood plasma or serum samples for GCTOF analysis Purpose: This SOP describes sample extraction and sample preparation for primary metabolism profiling by gas chromatography/time-of-flight mass spectrometry (GCTOF). References: Fiehn O, Kind T (2006) Metabolite profiling in blood plasma. In: Metabolomics: Methods and Protocols. Weckwerth W (ed.), Humana Press, Totowa NJ. Fiehn, O. Metabolomics by gas chromatography - mass spectrometry: combined targeted and untargeted profiling. 2016. Curr. Protoc. Mol. Biol. 114:30.4.1-30.4.32. doi: 10.1002/0471142727.mb3004s114. Starting material: Plasma/serum: 30 µL sample volume or aliquot Equipment: Centrifuge Eppendorf 5415 D Calibrated pipettes 1-200µL and 100-1000µL Multi-Tube Vortexer (VWR VX-2500) Orbital Mixing Chilling/Heating Plate (Torrey Pines Scientific Instruments) Speed vacuum concentration system (Labconco Centrivap cold trap) Nitrogen line with Pasteur pipette Chemicals and consumables: Product Manufacturer & Part Number Eppendorf tubes 1.5 mL, uncolored Eppendorf 022363204 Crushed ice UC Davis Water, LC/MS Grade Fisher Optima W6-4 Acetonitrile, LC/MS Grade Fisher Optima A955-4 Isopropanol, LC/MS Grade Fisher A461-4 pH paper 5-10 Millipore Sigma 1095330001 Bioreclamation human plasma (disodium EDTA) Bioreclamation HMPLEDTA Sample Preparation: Preparation of extraction solvent For 1 L of extraction solvent, combine 375 mL of acetonitrile, 375 mL of isopropanol, and 250 mL water in a 1 L bottle conditioned with the aforementioned chemicals. If a different total volume of extraction solvent is needed, simply mix acetonitrile, isopropanol, and water in volumes in proportion 3:3:2. Purge the extraction solution mix for 5 min with nitrogen with small bubbles. Make sure that the nitrogen line is flushed out of air before using it for degassing the extraction solvent solution. Store at -20°C until use. Note: if solvent freezes, sonicate until thawed and mix before use. Extraction Thaw raw samples at room temperature (or in the refrigerator at 4˚C) and vortex 10 sec at low speed to homogenize. Aliquot 30 μL of plasma sample into a 1.5 mL Eppendorf tube. Keep all samples on ice. Add 1 mL 3:3:2 (v/v/v) ACN:IPA:H2O extraction solvent (prechilled in a -20°C freezer). Vortex the sample for 10 sec. Shake for 5 min at 4°C using the Orbital Mixing Chilling/Heating Plate. Continue to keep all extracted samples on ice. Centrifuge samples for 2 min at 14000 rcf. Aliquot two 450 μL portions of the supernatant into 1.5 mL Eppendorf tubes (one for analysis and one as a backup sample). Transfer 100 μL of the remaining supernatant from each sample to a 2, 15, or 50 mL tube for pools, depending on number of samples in the study. Evaporate one 450 μL aliquot of the sample in the Labconco Centrivap cold trap concentrator to complete dryness. Proceed with cleanup or store tubes at -20°C until cleanup. Pooling Transfer multiple 475 µL aliquots of pooled samples to 1.5 mL Eppendorf tubes, one aliquot for every 10 samples in the study. If there is still pool remaining, prepare additional aliquots for backup. Centrifuge pool samples for 2 min at 14000 rcf. Remove 450 µL supernatant to new 1.5 mL Eppendorf tube. Evaporate to complete dryness in the Labconco Centrivap cold trap concentrator. Proceed with cleanup or store tubes at -20°C until cleanup. Cleanup Resuspend the dried aliquot with 500 μL 50:50 (v/v) ACN:H2O (degassed as given above) and vortex for about 10 sec. Centrifuge for 2 min at 14000 rcf. Remove 475 μL supernatant to a new 1.5 mL Eppendorf tube. Evaporate the transferred supernatant to complete dryness in the Labconco Centrivap cold trap concentrator. Submit to derivatization (see SOP “Derivatization of GC Samples & Standards”) or store at -20°C until ready for analysis. Quality assurance For every 50 samples, perform one method blank negative control extraction by applying the total procedure (i.e. all materials and plastic ware) without biological sample. If no combined pool was made from the extracted samples, use one commercial plasma/serum pool sample per 10 authentic subject samples as control instead. Disposal of waste Collect all chemicals in appropriate bottles and follow the disposal rules. Collect residual plasma/serum samples in specifically designed red ‘biohazard’ waste bags.

Combined analysis:

Analysis ID AN002547
Analysis type MS
Chromatography type GC
Chromatography system Leco
Column Rtx-5Sil MS column
MS Type EI
MS instrument type GC-TOF
MS instrument name Leco Pegasus IV TOF
Ion Mode POSITIVE
Units normalized peak height

Chromatography:

Chromatography ID:CH001865
Chromatography Summary:A 30 m long, 0.25 mm i.d. Rtx-5Sil MS column (0.25 μm 95% dimethyl 5% diphenyl polysiloxane film) with additional 10 m integrated guard column is used (Restek, Bellefonte PA). 99.9999% pure Helium with built-in purifier (Airgas, Radnor PA) is set at constant flow of 1 ml/min. The oven temperature is held constant at 50°C for 1 min and then ramped at 20°C/min to 330°C at which it is held constant for 5 min.
Instrument Name:Leco
Column Name:Rtx-5Sil MS column
Chromatography Type:GC

MS:

MS ID:MS002365
Analysis ID:AN002547
Instrument Name:Leco Pegasus IV TOF
Instrument Type:GC-TOF
MS Type:EI
MS Comments:GC-TOF Method: Instruments: Gerstel CIS4 –with dual MPS Injector/ Agilent 6890 GC- Pegasus III TOF MS Injector conditions: Agilent 6890 GC is equipped with a Gerstel automatic liner exchange system (ALEX) that includes a multipurpose sample (MPS2) dual rail, and a Gerstel CIS cold injection system (Gerstel, Muehlheim, Germany) with temperature program as follows: 50°C to 275°C final temperature at a rate of 12 °C/s and hold for 3 minutes. Injection volume is 0.5 μl with 10 μl/s injection speed on a splitless injector with purge time of 25 seconds. Liner (Gerstel #011711-010-00) is changed after every 10 samples, (using the Maestro1 Gerstel software vs. 1.1.4.18). Before and after each injection, the 10 μl injection syringe is washed three times with 10 μl ethyl acetate. Gas Chromatography conditions: A 30 m long, 0.25 mm i.d. Rtx-5Sil MS column (0.25 μm 95% dimethyl 5% diphenyl polysiloxane film) with additional 10 m integrated guard column is used (Restek, Bellefonte PA). 99.9999% pure Helium with built-in purifier (Airgas, Radnor PA) is set at constant flow of 1 ml/min. The oven temperature is held constant at 50°C for 1 min and then ramped at 20°C/min to 330°C at which it is held constant for 5 min. Mass spectrometer settings: 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:POSITIVE
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