Figure 5.

Study on the RAD001 resistant cell lines. (A) Growth curves of RAD001 resistant cell lines and the parental untreated cell lines. A slowed down proliferation was observed in the resistant cell lines comparing to the parental untreated cell lines even being cultured in RAD001 free medium. (B) Cell viability assay was applied to RAD001 resistant cell lines and the parental untreated cell lines. WST-1 absorption was tested after 72 h treatment. (C) The GI₅₀s of the primary cell lines and RAD001 resistant cell lines (*p < 0.05: RAD001 resistant cell lines vs. primary cell lines, n = 3). Much higher GI₅₀s were achieved in the resistant cell lines. (D) RAD001 resistant cell lines and their parental untreated cell lines were analyzed for acidic senescence-associated β-galactosidase activity. Higher percentage of β-gal positive cells was observed in the resistant cell liens. Scale bars: 100 μm. *p < 0.05: RAD001 resistant group vs. control group. (E) The expression of molecular markers was compared between the RAD001 resistant cell lines kept in 400nM RAD001 and the parental untreated cell lines by western blot.