Fig. 3.

BMP-2 dose-dependently triggered rapid metabolic reprogramming, which played a decisive role in subsequent osteogenic differentiation. A) Seahorse assay of the OCRs and ECARs of BMSCs during BMP-2 induction. B) Charts depicting the primary parameters calculated from the above Seahorse assay curves. C) Heatmap showing the gene expression profile during BMP-2-mediated osteogenic differentiation. (* and ▲ represent p < 0.05 in comparison with the control and 3d groups, respectively; **, ## and ▲▲ represent p < 0.01 in comparison with the control, 1d and 3d groups, respectively). D, E) Inhibition of glycolysis and the TCA cycle by 2-DG and LW6 substantially impeded the expression of the Ocn protein and biomineralization. F) BMP-2 stimulated significant phosphorylation of the Akt protein. G, H) ATP synthesis was disrupted and Alp activity was reduced after the treatment of BMSCs with BMP-2 and the Akt inhibitor MK2206 (Akti) (** represents p < 0.01 in comparison with the BMP + Akti group). I) BMP-2 activated the Akt pathway in a dose-dependent manner. J) Evaluation of metabolic enzymes in BMSCs treated with different doses of BMP-2 by immunofluorescence staining. K) JC-1 analysis of the mitochondrial membrane potential (ΔΨm) as an indicator of the cellular metabolic state. J-agg indicates cells with aggregated JC-1 (red fluorescence) and high ΔΨm, while j-free indicates cells with free JC-1 (green fluorescence) and low ΔΨm.