A: expression of Akt isoforms in cultured canine airway smooth muscle cells. Total cell lysates were prepared from serum-fed (FBS) cells or serum-deprived cells for 7 days maintained in insulin (10 μg/ml), transferrin (5.5 μg/ml), and selenium (5 ng/ml) (ITS). Akt1 is the predominant isoform expressed in both serum-fed and serum-deprived cells. Akt, total Akt. B: transfection of cultured canine tracheal myocytes with a plasmid expressing constitutively active Akt1 causes progressive increase in cell size in both serum-fed and serum-free cultures. Cells were cotransfected with a plasmid encoding enhanced green fluorescent protein (EGFP), and only EGFP+ cells were analyzed (P < 0.0001 by 2-way ANOVA for each culture condition). myrAkt-HA, cells transfected with pCMV-myrAkt-HA; empty, cells transfected with pcDNA3.1.