Summary of Study ST002143

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 PR001357. The data can be accessed directly via it's Project DOI: 10.21228/M89Q66 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 IDST002143
Study TitleEndo- and Exometabolome Crosstalk in Mesenchymal Stem Cells Undergoing Osteogenic Differentiation (Media Samples)
Study SummaryThe holistic nature of NMR enabled the time-course evolution of cholesterol, mono- and polyunsaturated fatty acids (including ω-6 and ω-3 fatty acids), several phospholipids (phosphatidylcholine, phosphatidylethanolamine, sphingomyelins, and plasmalogens), and mono- and triglycerides to be followed. Lipid changes occurred almost exclusively between days 1 and 7, followed by a tendency for lipidome stabilization after day 7. On average, phospholipids and longer and more unsaturated fatty acids increased up to day 7, probably related to plasma membrane fluidity. Articulation of lipidome changes with previously reported polar endometabolome profiling and with exometabolome changes reported here in the same cells, enabled important correlations to be established during hAMSC osteogenic differentiation. Our results supported hypotheses related to the dynamics of membrane remodelling, anti-oxidative mechanisms, protein synthesis, and energy metabolism. Importantly, the observation of specific up-taken or excreted metabolites paves the way for the identification of potential osteoinductive metabolites useful for optimized osteogenic protocols.
Institute
University of Aveiro
DepartmentDepartment of Chemistry
LaboratoryCICECO - Aveiro Institute of Materials
Last NameBispo
First NameDaniela S.C.
AddressCampus Universitário de Santiago, Aveiro, Aveiro, Aveiro, 3810-193, Portugal
Emaild.bispo@ua.pt
Phonenone
Submit Date2022-04-06
Raw Data AvailableYes
Raw Data File Type(s)1r
Analysis Type DetailNMR
Release Date2022-05-02
Release Version1
Daniela S.C. Bispo Daniela S.C. Bispo
https://dx.doi.org/10.21228/M89Q66
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001357
Project DOI:doi: 10.21228/M89Q66
Project Title:A Metabolomics-guided Bioreactor for Improved Engineered Bone Implants (BioImplant)
Project Type:NMR-based untargeted metabolomics
Project Summary:In an “omic” era, metabolomics offers exquisite insight into the complex metabolic network of living organisms and their adaptation mechanisms towards disease, therapy and environment. Metabolic markers (sets of metabolites) are emerging as new means of diagnostics, personalized follow-up and prediction of patient health status. Here, metabolomics is used for the first time to guide the development of a stem cell (SC) bioreactor to produce improved quality bone tissue for implantation. 3D porous scaffolds will be produced using biodegradable polymer poly-L-lactic acid (PLLA), both bare and collagen-coated to improve cell adhesion. These scaffolds will be 3D-printed with controlled architectures (both internally and externally), namely with struts alignment of 90o, with and without offset, expected to trigger distinct biological behaviors, particularly under mechanical cues. Scaffolds will be tested in a bioreactor for growth and differentiation of human mesenchymal SCs (hMSCs) into osteogenic lineage. hMSCs obtained from adipose tissue or bone marrow will be compared, as they have shown secretome differences and possible different potentials for osteogenic differentiation. The bioreactor will allow the application of controlled compression, to help mimic bone physiological conditions, and scaffold piezoelectricity will be studied in the same context. Scaffolds and physical cues will be tested in vitro (in osteogenic media) and monitored by biological measurements (proliferation, viability, differentiation indicators) and, for the first time, by cell metabolomics to identify the impact of each variable (scaffold composition, morphology, piezoelectricity and compression) on hMSC metabolism and define metabolic markers of hMSC function. Untargeted Nuclear Magnetic Resonance (NMR) metabolomics of cell extracts will identify dynamic metabolic cellular profiles associated to i) hMSC self-renewal and differentiation mechanisms, and their adaptations to ii) scaffold characteristics and iii) physical cues (compression and/or piezoelectricity). Statistical correlation of metabolic profiles with scaffold/bioreactor features and biological parameters will unveil metabolic markers of bioreactor performance and novel knowledge on SC osteogenic metabolism. Key metabolites will be identified as potentially osteogenesis-inducing, a role to be demonstrated using metabolite-tailored cell media to potentially substitute osteogenic growth factors and, thus, tackle related implantation challenges. Putative hypotheses of responsive hMSCs metabolic pathways will be validated through pathway network analysis, isotope-labeled tracers (NMR) and specific protein and genetic measurements. To our knowledge, this project proposes the first use of metabolomics to guide in vitro bone tissue engineering, building on recent proposals to exploit “omics” to understand, monitor and guide SC behavior for effective tissue engineering and implantation.
Institute:University of Aveiro
Department:Department of Chemistry
Laboratory:CICECO - Aveiro Institute of Materials
Last Name:Bispo
First Name:Daniela S.C.
Address:Campus Universitário de Santiago, Aveiro, Aveiro, Aveiro, 3810-193, Portugal
Email:d.bispo@ua.pt
Phone:none
Funding Source:The authors acknowledge the Portuguese Foundation for Science and Technology (FCT) for co-funding the BIOIMPLANT project (PTDC/BTM-ORG/28835/2017) through the COMPETE2020 program and European Union fund FEDER (POCI-01-0145-FEDER-028835). CSHJ is grateful to the same project for funding her contract with the University of Aveiro. DSB acknowledges the Sociedade Portuguesa de Química and FCT for her PhD grant SFRH/BD/150655/2020. This work was developed within the scope of the project CICECO-Aveiro Institute of Materials, UIDB/50011/2020, UIDP/50011/2020 & LA/P/0006/2020, financed by national funds through the FCT/MEC (PIDDAC). The NMR spectrometer used in this work is part of the National NMR Network (PTNMR) and, partially supported by Infrastructure Project Nº 022161 (co-financed by FEDER through COMPETE 2020, POCI and PORL and FCT through PIDDAC).

Subject:

Subject ID:SU002228
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606
Species Group:Mammals

Factors:

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

mb_sample_id local_sample_id Experiment day
SA205745Medium_Blank_S1Blank
SA205746Medium_Blank_S4Blank
SA205747Medium_Blank_S3Blank
SA205748Medium_Blank_S2Blank
SA205749OI1_D0_S3_MediumD0
SA205750OI1_D0_S1_MediumD0
SA205751OI1_D0_S2_MediumD0
SA205752OI1_D1_S3_MediumD1
SA205753OI1_D1_S2_MediumD1
SA205754OI1_D1_S1_MediumD1
SA205755OI1_D12_S1_MediumD12
SA205756OI1_D12_S2_MediumD12
SA205757OI1_D12_S3_MediumD12
SA205758OI1_D14_S3_MediumD14
SA205759OI1_D14_S2_MediumD14
SA205760OI1_D14_S1_MediumD14
SA205761OI1_D16_S1_MediumD16
SA205762OI1_D16_S3_MediumD16
SA205763OI1_D16_S2_MediumD16
SA205764OI1_D21_S1_MediumD21
SA205765OI1_D21_S2_MediumD21
SA205766OI1_D21_S3_MediumD21
SA205767OI1_D6_S3_MediumD6
SA205768OI1_D6_S1_MediumD6
SA205769OI1_D6_S2_MediumD6
SA205770OI1_D9_S2_MediumD9
SA205771OI1_D9_S1_MediumD9
SA205772OI1_D9_S3_MediumD9
Showing results 1 to 28 of 28

Collection:

Collection ID:CO002221
Collection Summary:hAMSCs conditioned media samples were collected in triplicate at days 1, 6, 9, 12, 14, 16, and 21 after filtration through 40 μm pore strainers.
Sample Type:Stem cells

Treatment:

Treatment ID:TR002240
Treatment Summary:hAMSCs were detached, at passage 7, from the flasks by trypsinization, counted in a Neubauer chamber and seeded at a density of 0.5 × 10 6 cells/flask. Cells were maintained under basal conditions until reaching ~100% confluence, then the basal culture medium was exchanged and supplemented with osteoinductive factors, specifically 10 mM β-glycerophosphate (β-GP, Sigma-Aldrich G9422), 50 µg/mL L-ascorbic acid (Sigma A0278), and 10 nM Dexa (ACROS Organics™ 230300010).

Sample Preparation:

Sampleprep ID:SP002234
Sampleprep Summary:Both blank and conditioned media samples were subjected to a protein-precipitation procedure and dried under vacuum. Prior to NMR analysis, dried media samples were resuspended in 100 mM phosphate buffer at pH 7.4 (prepared in D2O) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP in D2O, Sigma-Aldrich 293040), homogenised and transferred to 5 mm NMR tubes.
Extract Storage:-80℃
Sample Resuspension:100 mM phosphate buffer at pH 7.4 (prepared in D2O) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid (TSP in D2O, Sigma-Aldrich 293040)

Analysis:

Analysis ID:AN003508
Analysis Type:NMR
Software Version:Topspin 4.0.8
Operator Name:Daniela Bispo
Results File:ST002143_AN003508_Results.txt
Units:ppm

NMR:

NMR ID:NM000238
Analysis ID:AN003508
Instrument Name:Bruker Avance III
Instrument Type:FT-NMR
NMR Experiment Type:1D-1H
Field Frequency Lock:D2O
Spectrometer Frequency:500 MHz
NMR Probe:TXI
NMR Solvent:100 mM phosphate buffer at pH 7.4 (prepared in D2O) containing 0.1 mM 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid
NMR Tube Size:5 mm
Shimming Method:Topshim
Receiver Gain:203
Temperature:298 K
Number Of Scans:256
Dummy Scans:4
Acquisition Time:2.3 s
Relaxation Delay:4 s
Spectral Width:7002.801 Hz
Num Data Points Acquired:32k
Zero Filling:131k
Baseline Correction Method:Manual
Chemical Shift Ref Std:TSP
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