Summary of Study ST001627

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 PR001042. The data can be accessed directly via it's Project DOI: 10.21228/M80X2Z This work is supported by NIH grant, U2C- DK119886.

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Study IDST001627
Study TitleTissue-specific 1H-NMR metabolomic profiling in mice with adenine-induced chronic kidney disease - organic phase Heart (part-IV)
Study TypeMetabolomic profiling of the heart, liver, kidney, and skeletal muscles obtained from mice with and without CKD via 1H NMR
Study SummaryThis project is focused on a metabolomic analyses of the heart, liver, kidney, and skeletal muscles obtained from mice with and without CKD. To accomplish this objective, we extracted tissues from mice with CKD induced by long-term (24 week) adenine-supplemented diet as well as their control-diet fed counterparts with normal kidney function. Metabolites were extracted from tissues and 1H nuclear magnetic resonance (NMR) was performed and coupled with multivariate statistical analysis.
Institute
University of Florida
DepartmentApplied Physiology and Kinesiology
LaboratoryRm 42 and Rm 43
Last NameRyan
First NameTerence
Address1864 Stadium Rd, FLG 114, Gainesville, FL, 32611, USA
Emailryant@ufl.edu
Phone352-294-1700
Submit Date2020-12-11
Num Groups2
Total Subjects17
Num MalesAll
Study CommentsCKD metabolomic study via NMR using mice model
PublicationsMDPI
Raw Data AvailableYes
Raw Data File Type(s)fid
Analysis Type DetailNMR
Release Date2021-06-30
Release Version1
Terence Ryan Terence Ryan
https://dx.doi.org/10.21228/M80X2Z
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001042
Project DOI:doi: 10.21228/M80X2Z
Project Title:Tissue-specific 1H-NMR metabolomic profiling in mice with adenine-induced chronic kidney disease
Project Type:Metabolomic profiling of the heart, liver, kidney, and skeletal muscles obtained from mice with and without CKD via 1H NMR
Project Summary:This project is focused on a metabolomic analyses of the heart, liver, kidney, and skeletal muscles obtained from mice with and without CKD. To accomplish this objective, we extracted tissues from mice with CKD induced by long-term (24 week) adenine-supplemented diet as well as their control-diet fed counterparts with normal kidney function. Metabolites were extracted from tissues and 1H nuclear magnetic resonance (NMR) was performed and coupled with multivariate statistical analysis.
Institute:University of Florida
Department:Applied Physiology and Kinesiology
Laboratory:Rm 42 and Rm 43
Last Name:Ryan
First Name:Terence
Address:1864 Stadium Rd, FLG 114, Gainesville, FL, 32611, USA
Email:ryant@ufl.edu
Phone:352-294-1700
Funding Source:This research was funded by grants from the National Institutes of Health and the National Heart, Lung, and Blood, Institute numbers R01-HL149704 (to T.E.R.) and the American Heart Association grant number 18CDA34110044 (to T.E.R.).
Contributors:Ram B. Khattri, Trace Thome, and Terence E. Ryan

Subject:

Subject ID:SU001704
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57BL/6J
Age Or Age Range:13 months
Weight Or Weight Range:(32.86±1.21 g (control mice) vs 23.57±1.27 g (CKD mice), P < 0.0001, N=7/group).
Gender:Male
Animal Animal Supplier:Jackson Labs (Stock # 000664)
Animal Housing:Housed in a temperature of 22 oC
Animal Light Cycle:12-hour light/12-hour dark
Animal Feed:Ad libitum (Casein control diet vs. adenine-supplemented diet to induce CKD)
Animal Water:free access to food and water (3-5 animals per cage).
Animal Inclusion Criteria:(3-5 animals per cage).

Factors:

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

mb_sample_id local_sample_id Group
SA137982CKD7Heart_OrgCKD
SA137983CKD1Heart_OrgCKD
SA137984CKD5Heart_OrgCKD
SA137985CKD6Heart_OrgCKD
SA137986CKD2Heart_OrgCKD
SA137987CKD3Heart_OrgCKD
SA137988CKD4Heart_OrgCKD
SA137989Con6Heart_OrgControl
SA137990Con7Heart_OrgControl
SA137991Con5Heart_OrgControl
SA137992Con1Heart_OrgControl
SA137993Con2Heart_OrgControl
SA137994Con3Heart_OrgControl
SA137995Con4Heart_OrgControl
Showing results 1 to 14 of 14

Collection:

Collection ID:CO001697
Collection Summary:While under isoflurane anesthesia, tissues were rapidly dissected and snap frozen in liquid nitrogen and stored at -80°C until metabolite extraction. The following tissues were used in this study: kidney, liver, heart (left ventricle), skeletal muscle (quadriceps). Euthanasia was carried out by thoracotomy followed by cervical dislocation.
Sample Type:Heart
Collection Method:While under isoflurane anesthesia, tissues were rapidly dissected and snap frozen in liquid nitrogen and stored at -80°C until metabolite extraction
Collection Location:University of Florida, Applied Physiology and Kinesiology, 1864 stadium RD, Gainesville, FL 32611
Storage Conditions:-80℃
Storage Vials:cryovials

Treatment:

Treatment ID:TR001717
Treatment Summary:To induce CKD, we utilized an established adenine-diet model. Briefly, mice were assigned to a casein-base chow for 7-days, followed by induction of renal tubular injury by supplementing the diet with 0.2% adenine for 7-days, and subsequently maintained on a 0.15% adenine diet for 5 months and two weeks. CKD mice were then placed back on casein control diet for 2-weeks prior to euthanasia and terminal experiments, allowing for a washout period of adenine based chow. Control mice received casein diet for the duration of the study. Total duration of CKD encompassed 6-months.
Animal Vet Treatments:none
Animal Anesthesia:isoflurane
Animal Fasting:non-fasted
Animal Endp Euthanasia:Euthanasia was carried out by thoracotomy followed by cervical dislocation.
Animal Endp Tissue Coll List:kidney, liver, heart (left ventricle), skeletal muscle (quadriceps)

Sample Preparation:

Sampleprep ID:SP001710
Sampleprep Summary:A modified form of FOLCH extraction protocol was used to extract metabolites from the tissues. Wet weights of all tissue samples were recorded prior to extraction. Tissue samples were immediately homogenized to prevent any possible enzymatic action using 1 mL of ice-cold methanol in a PowerLyzer 24 Homogenizer (QIAGEN Group, Hilden, Germany). The mixture was centrifuged using 13,200 rpm at 4oC for 30 minutes and the resulting supernatant was transferred to a new glass vial consisting 3 mL of ice cold chloroform:methanol (2:1, v/v) mixture. The homogenate was vortexed and left in an ice bath for 15 minutes to allow for phase separation. Next, 1 mL of 0.9% of saline was added, vortexed it for couple of minutes followed by a second incubation in an ice bath for 30-45 min for complete phase separation. The upper aqueous layer was transferred to a new falcon tube. To the remaining organic phase sample, 1 mL of 0.9% of saline was added again followed by vigorous mixing and letting it stand in ice bath (15 minutes) for a second phase separation. This second aqueous phase was combined with the first. The resulting aqueous and organic layers were dried separately. The aqueous layer was dried overnight with a Labconco freezer dryer (Labconco Corporation, MO, USA) and the organic layer was dried via inert nitrogen gas. These two dried powders (aqueous and organic phases) were stored at -80oC until performing NMR experiments.
Processing Method:Lyophilization and Homogenization
Processing Storage Conditions:-80℃
Extraction Method:Modified FOLCH extraction
Extract Storage:-80℃
Sample Resuspension:Deuterated chloroform (80 microliter) with 10 mM pyrazine was used to re-suspend organic phase samples.
Sample Spiking:10 mM of pyrazine for organic phase samples.

Analysis:

Analysis ID:AN002662
Laboratory Name:Terence lab, UF
Analysis Type:NMR
Acquisition Date:09/28/2020
Software Version:Bruker Topspin
Operator Name:Ram Khattri
Detector Type:Bruker
Data Format:fid, 1r
Num Factors:2
Num Metabolites:19
Units:A.U.

NMR:

NMR ID:NM000194
Analysis ID:AN002662
Instrument Name:Bruker Avance Neo 600 MHz/54mm console
Instrument Type:FT-NMR
NMR Experiment Type:1D-1H
Field Frequency Lock:Deuterated chloroform
Standard Concentration:10mM pyrazine
Spectrometer Frequency:600.2328273 MHz
NMR Probe:1.7 mm TXI CryoProbe
NMR Solvent:Deuterated chloroform
NMR Tube Size:1.7 mm O.D.
Shimming Method:Topshim
Pulse Sequence:noesypr1d
Water Suppression:none
Pulse Width:90-degree
Receiver Gain:101
Offset Frequency:2827.31 Hz
Chemical Shift Ref Cpd:CDCl3 at 7.26 ppm and pyrazine at 8.61 ppm
Temperature:25 o C
Number Of Scans:128 scans
Dummy Scans:8
Acquisition Time:4 s
Relaxation Delay:1 s
Spectral Width:7142.9 Hz
Num Data Points Acquired:28571
Real Data Points:65536
Line Broadening:0.22 Hz
Zero Filling:65,536 points
Apodization:Exponential
Baseline Correction Method:Spline
Chemical Shift Ref Std:7.26ppm for CDCl3
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