FIG. 5.

Osteoinduction activates Akt and increases phosphorylation of mitochondrial proteins. (A) Western blots of phospho-Akt and total Akt protein following 5-, 15-, 30-, or 60-min incubation with OBM (left), BMP2 (center), or Wnt3a (right). (B) (top) Western blot using anti-Akt phosphorylation sequence (RXXSp/Tp) to detect Akt target proteins in mitochondrial protein extracts following 48 h of osteoinduction. Quantification of total lane intensity shown below expressed as fold change over control lane. (bottom) No-Stain™ Protein Labeling Reagent used as total protein loading control. (C) Western blot of phospho-Akt and total Akt protein following incubation with pan 1/2/3 Akt inhibitor (iAkt) for 24 h, and then OBM (left), BMP2 (center), or Wnt3a (right) addition for 12 h for ST2 (C) and MC3T3-E1 (D) cells. Western blots and stains are representative images from n = 3 technical replicates. *P < 0.05 versus uninduced control as determined by t-test.”