Fig. 5

Inhibition of AKT1 signaling pathway sensitizing the cytotoxic effects of Dox and AD198 in human TCC cells. a T24 and UMUC3 cells were treated with Dox and AD198 (1 μM) with and without LY294002 (LY, 20 μM) for 48 h and compared to control groups. Cell proliferation was determined by MTS assay and relative cell growth rate was normalized to control counterpart. Values represent mean ± SE of four replicates from three independent experiments. Paired Student t-tests were used to compare DOX and AD198 treatments to control; ***p ≤ 0.001. Paired Student t-tests were used to compare Dox to Dox + Ly and AD198 to AD198 + LY treatment, ^### p ≤ 0.001. b T24 and UMUC3 cells were treated with DOX and AD198 (1 μM) with and without LY294002 (20 μM) for 24 h and caspase activities were measured using the Caspase-Glo 3/7 luminescence assay. Relative caspase activities were normalized to control. Values represent mean ± SE of three independent experiments. Paired Student t-tests were used to compare treatment to control **p ≤ 0.01, ***p ≤ 0.001. Paired Student t-tests were used to compare Dox to Dox + Ly and AD198 to AD198 + LY treatments, ^### p ≤ 0.001. c T24 and UMUC3 cells were treated with Dox and AD198 (1 μM) with and without LY294002 (20 μM) for 24 h. The expression of PARP (cleaved fragment) were evaluated by WB analysis. Actin was used as a loading control. Densitometry evaluation of PARP protein bands from WB analysis was done using ImageJ software. Values represent mean ± S.E. of measured densitometry of each protein’s band from three independent experiments. Paired Student t-tests were used to compare controls to Dox and AD198 treatments, *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001. Paired Student t-tests were used to compare Dox to AD198 treatment, ^# p ≤ 0.05, ^## p ≤ 0.05, and ^### p ≤ 0.001 d T24 and UMUC3 cells were treated with Dox and AD198 (1 μM) with and without LY294002 (20 μM) for 24 h. The expression of p-AKT (T308), p-AKT (S473), AKT1 and p-GSK-3β proteins were evaluated by WB analysis. Actin was used as a loading control. Densitometry evaluation of p-AKT (T308), p-AKT (S473) protein bands from WB analysis was done using ImageJ software. Values represent mean ± S.E. of measured densitometry of each band from three independent experiments. Paired Student t-tests were used to compare controls to Dox and AD198 treatments, *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001. Paired Student t-tests were used to compare Dox to Dox + LY or AD198 to AD198 + LY treatments, ^# p ≤ 0.05, ^## p ≤ 0.01