Figure 1
EGCG regulated the expression of JWA and topoisomerase IIα (topo IIα) in vitro (NCI-H460 and A549 cells) and in vivo.
(a) Cells transfected with Flag-JWA (4 μg) or without transfection were incubated with EGCG (20–40 μM) for 24 h. JWA protein expression was assessed by Western blot analysis. β-actin expression served as a loading control. (b) NCI-H460 cells were treated with EGCG (20–40 μM) for 24 h and total cellular RNA was extracted. mRNA level of JWA was detected by real-time PCR. GAPDH was used as an internal control. (c) Western blot analysis of the protein level of JWA in EGCG-treated A549 cells for 24 h. β-actin expression served as a loading control. (d) After A549 cells were incubated with EGCG (20–40 μM) for 24 h, total RNAs were prepared and real-time PCR was applied to measure the JWA mRNA level. GAPDH was used as an internal control. (e) NCI-H460 cells were transfected with or without Flag-topoisomerase IIα plasmid (4 μg) and then treated with EGCG (20–40 μM) for 24 h. Protein from cell was subjected to western blot analysis. β-actin expression was served as a loading control. (f) Total RNAs from NCI-H460 cells incubated with EGCG (20–40 μM) for 24 h were extracted and subjected to real-time PCR using primer for topoisomerase IIα. GAPDH was used as an internal control. (g) A549 cells were treated with EGCG (20–40 μM) for 24 h and then protein from cell lysate was subjected to western blot analysis. β-actin expression was served as a loading control. (h) A549 cells were incubated in the absence or presence of EGCG (20–40 μM) for 24 h. Then cells were lysed for the detection expression of topoisomerase IIα mRNA by real-time PCR. GAPDH was used as an internal control. The A549 xenograft nude mice model was established and treated with EGCG or normal saline. (i) Tumor size was checked twice per week. (j) Protein obtained from tumor tissues was subjected to western blot. β-tubulin expression was served as a loading control. Error bars represent the mean ± SD of triplicate experiments. Statistical differences to the controls were shown as *p < 0.05, **p < 0.01.
