(1)H NMR Metabolite Monitoring during the Differentiation of Human Induced Pluripotent Stem Cells Provides New Insights into the Molecular Events That Regulate Embryonic Chondrogenesis

The integration of cell metabolism with signalling pathways, transcription factor networks and epigenetic mediators is critical in coordinating molecular and cellular events during embryogenesis. Induced pluripotent stem cells (IPSCs) are an established model for embryogenesis, germ layer specification and cell lineage differentiation, advancing the study of human embryonic development and the translation of innovations in drug discovery, disease modelling and cell-based therapies. The metabolic regulation of IPSC pluripotency is mediated by balancing glycolysis and oxidative phosphorylation, but there is a paucity of data regarding the influence of individual metabolite changes during cell lineage differentiation. We used (1)H NMR metabolite fingerprinting and footprinting to monitor metabolite levels as IPSCs are directed in a three-stage protocol through primitive streak/mesendoderm, mesoderm and chondrogenic populations. Metabolite changes were associated with central metabolism, with aerobic glycolysis predominant in IPSC, elevated oxidative phosphorylation during differentiation and fatty acid oxidation and ketone body use in chondrogenic cells. Metabolites were also implicated in the epigenetic regulation of pluripotency, cell signalling and biosynthetic pathways. Our results show that (1)H NMR metabolomics is an effective tool for monitoring metabolite changes during the differentiation of pluripotent cells with implications on optimising media and environmental parameters for the study of embryogenesis and translational applications.