Figure 5

Effects of XPF or XPG downregulation on the repair of cisplatin-induced intrastrand crosslinks and cisplatin sensitivity in cisplatin-resistant cancer cells. A, B) CP70 and CDDP cells were transfected with control siRNA, XPF siRNA or XPG siRNA for 48 and 72 h. Whole cell lysates were prepared and subjected to Western blotting for detecting the expression of XPF and XPG. C, D) CP70 and CDDP cells were transfected with siRNA described above for 24 h, pretreated with HU for 24 h, and then treated with cisplatin for 1 h. The cells were then incubated in HU containing medium for 0, 8 and 24 h. The genomic DNA was isolated and analyzed by ISB assay for cisplatin-induced DNA intrastrand crosslinks with anti-Pt-GG antibody. The relative percentage of remaining Pt-GG at different time points is an average of three independent repeats (n = 3, Bar indicates SD). E, F) CP70 and CDDP cells growing in 96-well plates were transfected with above-described siRNA for 24 h. Cells were then treated with cisplatin at the indicated dose for 1 h, and further cultured in drug-free medium for 72 h. Methylene blue stained cells in the treated samples compared to untreated control cells were used to obtain relative cell survival. (n = 4, Bar indicates SD).