Fig. 5.

The effect of HZ08 on IR-induced RelB nuclear translocation and MnSOD expression. a, PC-3 and DU-145 cells were treated with different concentrations of HZ08. The levels of nuclear RelB and relating cytosolic MnSOD were quantified by Western blots. PCNA and β-actin served as the loading controls. b, the two cell lines were pretreated with HZ08 and then followed by IR treatment as indicated. The amounts of RelB and MnSOD in the fractions of nuclei and cytosols were quantified accordingly. c, PC-3 cells were treated with HZ08 and IR as described in Fig. 4c. Chromatin was isolated from the treated cells and pulled down using a RelB antibody or IgG control. An enhancer fragment of the SOD2 gene containing a NF-κB element was amplified by qPCR with specific primers. The amount of pulled down fragment was quantified by normalizing amplified from unprecipitated chromatin (input control). d, after treatment, the cell extracts were subjected to measure MnSOD activity. e-g, PC-3 and DU-145 cells were transfected with a MnSOD expression construct, and then treated with HZ08 and IR. The increased level of MnSOD mRNA was confirmed by qRT-PCR with β-actin normalization (e). Cell survival was quantified by colony formation (f and g). Mean ± SD was representative of three independent experiments carried out in duplication. *(P < 0.05), **(P < 0.01) show the significances between two groups as indicated