abstract
Conventional methods involving the measurement of established biochemical indicators are usually used to assess/characterize the osteogenic differentiation of mesenchymal stem cells. These methods commonly entail the quantification of osteocalcin, calcium levels, and alkaline phosphatase (ALP) activity throughout differentiation. Although important, these approaches are time-consuming, significantly operator-dependent and require the full 21-day follow-up of differentiation. We propose that metabolomics can rapidly and efficiently assess differentiation through a single analysis from stages as early as day 7, by measuring donor-independent metabolite markers of differentiation. We present an untargeted Nuclear Magnetic Resonance (NMR) metabolomics study of 3 independent cell donors and the characterization of their cellular metabolic profile as a function of differentiation, to identify and refine a potential marker set of metabolites for early prediction of osteogenic capacity. This work builds on previous hypotheses primarily founded on data from 2 donors alone [1], which advanced, e.g., glutathione, phosphocreatine, adonitol, ethanolamine, choline and phosphocholine as donor-independent marker metabolites. The present study focuses on a 3rd independent donor to validate/discard previous hypotheses. As a sideline, the effect of DMSO (required for dexamethasone – one of the traditional osteogenic inductor compounds – dissolution) on cells behavior was investigated, as previous reports [2,3] suggest that it affects proliferation and osteogenesis.
authors
Correia MC, Bispo DSC, Jesus CSH, Rodrigues JEA, Oliveira MB, Mano JF, Gil AM
our authors
Ana Maria Pissarra Coelho Gil
Associate Professor with Aggregation
Catarina Sofia Henriques de Jesus
Junior Researcher
Daniela Bispo
PhD Student
João A. Rodrigues
Junior Researcher
João Mano
Full professor
Mariana Braga de Oliveira
Assistant Researcher
Marlene Correia
PhD StudentProjects
Metabolite-activated 3D stem cell differentiation into bone (BetterBone)
A Metabolomics-guided Bioreactor for Improved Engineered Bone Implants (BioImplant)
CICECO - Aveiro Institute of Materials (UIDB/50011/2020)
CICECO - Aveiro Institute of Materials (UIDP/50011/2020)
Associated Laboratory CICECO-Aveiro Institute of Materials (LA/P/0006/2020)
acknowledgements
BetterBone project (2022.04286.PTDC) and the BIOIMPLANT project (PTDC/BTM-ORG/28835/2017) through COMPETE2020 and European Union fund FEDER (POCI-01-0145-FEDER-028835); CICECO – Aveiro Institute of Materials project (UIDB/50011/2020, UIDP/50011/2020 & LA/P/0006/2020), financed by national funds through the FCT/MCTS and co-financed by FEDER under the PT2020. DSB acknowledges the Sociedade Portuguesa de Química and FCT for her PhD grant SFRHBD/150655/2020. The NMR spectrometer used in this work is part of the National NMR Network, partially supported by Infrastructure Project No. 022161 (co-financed by FEDER through COMPETE 2020, POCI and PORL and FCT through PIDDAC).