Fig. 5.
NF-κB is required for tmTNF-α-mediated DOX-resistance. a Representative IHC images of phosphorylation of NF-κB p65 in paratumor and breast cancer tumors, and histogram showing co-expression rate of phosphorylated NF-κB p65 and tmTNF-α in tumors of 105 patients. (b-i) Parental or shRNA-transfected MDA-MB-231 cells and parental or NTF-transfected MCF-7 cells were treated with or without 3 μM DOX for 24 h. (b and d) Representative immunoblotting of levels of IκB-α, cIAP1, cIAP2, and XIAP in total protein and translocation of NF-κB p65 from cytoplasmic fraction to nuclear fraction. LamB1 or β-actin served as a nuclear protein or a cytoplasmic/total protein loading control. c, e NF-κB activity was detected by ELISA. f–i Real-time PCR analysis of Bcl-XL (f and g) and BAX (h and i). j, k A 24 h-DOX (3 μM)-induced apoptosis was determined in parental or shRNA-transfected MDA-MB-231 cells (j) and in parental or NTF-transfected MCF-7 cells (k) pretreated with 10 μM BAY117082 (BAY) for 30 min using Annexin V/PI staining. Data are represented as mean ± SEM of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001