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.
| Study ID | ST002143 |
| Study Title | Endo- and Exometabolome Crosstalk in Mesenchymal Stem Cells Undergoing Osteogenic Differentiation (Media Samples) |
| Study Summary | The 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 |
| 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 |
| d.bispo@ua.pt | |
| Phone | none |
| Submit Date | 2022-04-06 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | 1r |
| Analysis Type Detail | NMR |
| Release Date | 2022-05-02 |
| Release Version | 1 |
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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 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Experiment day |
|---|---|---|
| SA205745 | Medium_Blank_S1 | Blank |
| SA205746 | Medium_Blank_S4 | Blank |
| SA205747 | Medium_Blank_S3 | Blank |
| SA205748 | Medium_Blank_S2 | Blank |
| SA205749 | OI1_D0_S3_Medium | D0 |
| SA205750 | OI1_D0_S1_Medium | D0 |
| SA205751 | OI1_D0_S2_Medium | D0 |
| SA205752 | OI1_D1_S3_Medium | D1 |
| SA205753 | OI1_D1_S2_Medium | D1 |
| SA205754 | OI1_D1_S1_Medium | D1 |
| SA205755 | OI1_D12_S1_Medium | D12 |
| SA205756 | OI1_D12_S2_Medium | D12 |
| SA205757 | OI1_D12_S3_Medium | D12 |
| SA205758 | OI1_D14_S3_Medium | D14 |
| SA205759 | OI1_D14_S2_Medium | D14 |
| SA205760 | OI1_D14_S1_Medium | D14 |
| SA205761 | OI1_D16_S1_Medium | D16 |
| SA205762 | OI1_D16_S3_Medium | D16 |
| SA205763 | OI1_D16_S2_Medium | D16 |
| SA205764 | OI1_D21_S1_Medium | D21 |
| SA205765 | OI1_D21_S2_Medium | D21 |
| SA205766 | OI1_D21_S3_Medium | D21 |
| SA205767 | OI1_D6_S3_Medium | D6 |
| SA205768 | OI1_D6_S1_Medium | D6 |
| SA205769 | OI1_D6_S2_Medium | D6 |
| SA205770 | OI1_D9_S2_Medium | D9 |
| SA205771 | OI1_D9_S1_Medium | D9 |
| SA205772 | OI1_D9_S3_Medium | D9 |
| 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 |
