Summary of Study ST001615

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 PR001039. The data can be accessed directly via it's Project DOI: 10.21228/M8D400 This work is supported by NIH grant, U2C- DK119886.

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Study IDST001615
Study TitleUnique metabolomic profile of skeletal muscle in chronic limb threatening ischemia (organic phase samples)
Study TypeNMR
Study SummaryThis project is focused on a cross-sectional analysis of non-PAD controls and CLTI patients undergoing either a vascular intervention or undergoing limb amputation was performed and involved a detailed assessment of the limb muscle metabolome using HR-MAS and solution state NMR spectroscopy. It was hypothesized that patients undergoing limb amputation would present with altered muscle metaolite features compared with non-PAD controls.
Institute
University of Florida
DepartmentApplied Physiology and Kinesiology
LaboratoryRm 42 and Rm 43
Last NameRyan
First NameTerence
AddressUniversity of Florida, Applied Physiology and Kinesiology, 1864 stadium RD, Gainesville, FL 32611
Emailryant@ufl.edu
Phone352-294-1700
Submit Date2020-11-30
Num Groups3
Total Subjects30
Num Males25
Num Females5
Study CommentsPAD metabolomic study via NMR. A detailed assessment of the limb muscle metabolome using semi-solid and solution state NMR spectroscopy
PublicationsJournal of Clinical Medicine
Raw Data AvailableYes
Raw Data File Type(s)wiff
Analysis Type DetailNMR
Release Date2021-05-30
Release Version1
Terence Ryan Terence Ryan
https://dx.doi.org/10.21228/M8D400
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001039
Project DOI:doi: 10.21228/M8D400
Project Title:Unique metabolomic profile of skeletal muscle in chronic limb threatening ischemia
Project Type:NMR
Project Summary:This project is focused on a cross-sectional analysis of non-PAD controls and CLTI patients undergoing either a vascular intervention or undergoing limb amputation was performed and involved a detailed assessment of the limb muscle metabolome using HR-MAS and solution state NMR spectroscopy. It was hypothesized that patients undergoing limb amputation would present with altered muscle metabolite features compared with non-PAD controls.
Institute:University of Florida
Department:Applied Physiology and Kinesiology
Laboratory:Rm 42 and Rm 43
Last Name:Ryan
First Name:Terence
Address:University of Florida, Applied Physiology and Kinesiology, 1864 stadium RD, Gainesville, FL 32611
Email:ryant@ufl.edu
Phone:352-294-1700
Funding Source:National Institutes of Health and the National Heart, Lung, and Blood, Institute numbers R01-HL149704 (to T.E.R.) and R01-HL148597 (to S.T.S.); as well as the American Heart Association grant number 18CDA34110044 (to T.E.R.). A portion of this work was supported by pilot funds (awarded to T.E.R.) from The University of Florida Claude D. Pepper Older Americans Independence Center P30AG028740. (if applicable)
Project Comments:PAD metabolomic study via NMR. A detailed assessment of the limb muscle metabolome using semi-solid and solution state NMR spectroscopy.
Publications:Journal of Clinical Medicine
Contributors:Ram B. Khattri, Kyoungrae Kim, Trace Thome, Zachary R. Salyers, Kerri A. O’Malley, Scott A. Berceli, Salvatore T. Scali, Terence E. Ryan*

Subject:

Subject ID:SU001692
Subject Type:Human
Subject Species:Homo sapiens
Taxonomy ID:9606
Age Or Age Range:55-82 years
Gender:Male and female
Human Race:unknown
Human Ethnicity:unknown
Human Medications:Asprin, ACE inhibitor, statins, cilostazol
Human Smoking Status:Controls = 4, CLTI Pre-surgery = 7, and CLTI Amputation = 9

Factors:

Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Group
SA137165CLTI_Amputation9aCLTI_Amputation
SA137166CLTI_Amputation10aCLTI_Amputation
SA137167CLTI_Amputation8aCLTI_Amputation
SA137168CLTI_Amputation1aCLTI_Amputation
SA137169CLTI_Amputation7aCLTI_Amputation
SA137170CLTI_Amputation2aCLTI_Amputation
SA137171CLTI_Amputation3aCLTI_Amputation
SA137172CLTI_Amputation4aCLTI_Amputation
SA137173CLTI_Amputation6aCLTI_Amputation
SA137174CLTI_Amputation5aCLTI_Amputation
SA137175CLTI_Pre_surgery7aCLTI_Pre_surgery
SA137176CLTI_Pre_surgery9aCLTI_Pre_surgery
SA137177CLTI_Pre_surgery5aCLTI_Pre_surgery
SA137178CLTI_Pre_surgery6aCLTI_Pre_surgery
SA137179CLTI_Pre_surgery1aCLTI_Pre_surgery
SA137180CLTI_Pre_surgery2aCLTI_Pre_surgery
SA137181Control8aControl
SA137182Control10aControl
SA137183Control7aControl
SA137184Control5aControl
SA137185Control2aControl
SA137186Control3aControl
SA137187Control4aControl
SA137188Control6aControl
Showing results 1 to 24 of 24

Collection:

Collection ID:CO001685
Collection Summary:Gastrocnemius muscle specimens were obtained from ten older adult non-PAD controls (Control), ten patients with critical limb ischemia (CLI) undergoing surgical intervention (CLTI Pre-surgery), and ten CLI patients undergoing limb amputation (CLTI Amputation). Five pre-surgery patients underwent bypass interventions and five underwent endovascular procedures. Muscle specimens were collected within the confines of the operating rooms (CLI patients) or via percutaneous muscle biopsy using sterile procedures. A portion of the muscle was quickly trimmed of fat/connective tissue and snap frozen in liquid nitrogen for metabolomics analysis.
Sample Type:Muscle
Collection Method:Muscle specimens were collected within the confines of the operating rooms (CLI patients) or via percutaneous muscle biopsy using sterile procedures. A portion of the muscle was quickly trimmed of fat/connective tissue and snap frozen in liquid nitrogen for metabolomics analysis.
Storage Conditions:-80℃
Storage Vials:cryovials

Treatment:

Treatment ID:TR001705
Treatment Summary:This was a prospective cohort study that examined the metabolomic profile of skeletal muscle from CLTI patients undergoing surgerical intervention or amputation, as well as a cohort of non-PAD controls. CLTI patients exhibited severe symptomology (Rutherford Classification 3-6), with high incidence of common PAD risk factors including hypertension, hyperlipidemia, coronary artery disease, and diabetes.Ten patients with critical limb ischemia (CLI) undergoing surgical intervention (CLTI Pre-surgery), and ten CLI patients undergoing limb amputation (CLTI Amputation). Five pre-surgery patients underwent bypass interventions and five underwent endovascular procedures. Muscle specimens were collected within the confines of the operating rooms (CLI patients) or via percutaneous muscle biopsy using sterile procedures.
Human Fasting:non-fasted
Human Endp Clinical Signs:N/A

Sample Preparation:

Sampleprep ID:SP001698
Sampleprep Summary:Extraction. Both polar and non-polar metabolites were extracted from the gastrocnemius muscle specimens using FOLCH extraction. In brief, wet weigh of the frozen tissues were determined and immediately homogenized in 1 mL of ice-cold methanol using a PowerLyzer 24 Homogenizer (QIAGEN Group, Hilden, Germany). All enzymatic activities are halted once the sample was homogenized in the methanol. Homogenization was followed by centrifugation (13.2K r.p.m., 4 oC, 30 minutes) and supernatant was transferred into a new glass vial consisting a mixture of 3 mL of ice-cold chloroform and methanol (2:1 v/v) ratio. The cold mixture was vortexed for several minutes and left in an ice bath 15 minutes to allow for phase separation. Next, 1 mL of ice-cold 0.9% saline was added to the mixture followed by vigorous mixing. The mixture was again left in the ice bath for 45 minutes for phase separation. The upper methanol/water layer was transferred to a new falcon tube. To the lower chloroform layer, 1 mL of ice-cold 0.9% saline was added and all steps were followed as mentioned above. Following a 45-minute incubation, upper methanol/water layer was again transferred to the previous falcon tube and dried using a Labconco freeze drier (Labconco Corporation, MO, USA). The chloroform layer was dried under a stream of nitrogen gas. The dried samples (both aqueous and organic phases) were stored at -80 oC until resuspension for NMR experiments. Lyophilized organic phase sample were re-suspended in 80 µL of CDCl3 along with 10 mM of pyrazine. All samples were loaded in 1.7 mm NMR tube to acquire spectra.
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:AN002650
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:3
Num Metabolites:21
Units:A.U.

NMR:

NMR ID:NM000187
Analysis ID:AN002650
Instrument Name:Bruker Avance Neo 600 MHz/54mm console
Instrument Type:FT-NMR
NMR Experiment Type:1D-1H
Field Frequency Lock:Deuterium
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
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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