Effects of Dox alone and Y-27632 Plus Dox on Key Apoptotic Factors in CSCs.

(A) The dose-dependent expressions of Akt/p-Akt/Bcl-2/Caspase-3/C-Caspase-3 in CSCs under Dox treatments. Briefly, CSCs were incubated with Dox at 0.0, 0.2, 0.4, 0.6, 0.8, and 1.0μM for 48hrs followed by Western blot analysis. The means of fold changes by Western blot shows a “Gaussian-type” response for pAkt/Akt and Bcl-2 but a “Sigmoid-type” for C-Caspase-3 with increasing concentrations of Dox. n = 3-4.ns: no significance; *: ppp(B) Pretreatment of Y-26732 significantly decreased the activity of cleaved Caspase-3 induced by Dox-treatment. In the experiment, CSCs were incubated with or without Y-27632 (10μM) for 48hr followed by Dox (0.6μM) for 48 hrs. Western blot was performed at the end of the treatments. The means of fold changes are shown in the right bar graphs except for those (Akt, Bcl-xl, and Bax) without statistical significance. For all panels, n = 3–6; ns: no significance; *: ppDox: Doxorubicin; Akt: protein Kinase-B (PKB); p-Akt: phospho-Akt; Bcl-2: B-cell lymphoma 2; Bcl-xl: Bcl-extra large (or Bcl-2-like 1 isoform 1); Bax: Bcl-2-associated X protein; C-Caspase-3: cleaved Caspase-3.