FIG. 3.

Akt activation by PGE₂ is independent of the activation of PKA signaling. (A) MLO-Y4 cells were treated with vehicle, PGE₂ (5 μM), FSK (10 μM), or 8-Br-cAMP (2 mM) for 1 h. Cell lysates were immunoblotted with anti-Akt antibody specific for total Akt or Akt phosphorylated at position 308 or 473. The normalized ratio of pAkt to Akt from densitometric measurements of three separate Western blots is presented in right panel. pAkt308 or pAkt473 under PGE₂ treatment versus vesicle control or other treatments: **, P < 0.01. The data are presented as mean ± SEM (n = 3). (B) MLO-Y4 cells were pretreated with vehicle or PKI (0.4 μM) for 15 min, followed by treatment with PGE₂ (5 μM) for 1 h. Cell lysates were immunoblotted with anti-Akt antibody specific for total Akt or Akt phosphorylated at position 308 or 473. The normalized ratio of pAkt to Akt from densitometric measurements of three separate Western blots is presented in right panel. pAkt308 or pAkt473 under PGE₂ or PGE₂ plus PKI treatment versus vesicle control or other treatments: **, P < 0.01. The data are presented as mean ± SEM (n = 3).