Figure 1
(A) A representative autoradiogram showing NFκB DNA-binding activity levels in MCF-7 cells exposed to hypoxia and/or radiation with or without EF24, NLE, CUR, GEN, RES or RSE pre-treatment. The nuclear extracts were analyzed by EMSA using γ-32p [ATP] labeled NFκB-specific probe. Compared to the normoxia controls, hypoxic cells showed a significant induction of NFκB-DNA binding activity. IR-exposure further enhanced hypoxia induced NFκB activity. Hypoxic cells treated with EF24, NLE, CUR, GEN, RES or RSE showed a significant inhibition of IR-induced NFκB-DNA binding activity. (B) Semi-quantitative densitometric analysis showing the effect of EF24, NLE, CUR, GEN, RES or RSE on IR-induced NFκB-DNA binding activity in hypoxic MCF-7 cells. (C) Western blot analysis showing modulation in the IκBα phosphorylation, nuclear translocation of NFκB p50 and p65 in hypoxic MCF-7 cells exposed to IR with or without EF24, NLE, CUR, GEN, RES or RSE treatment. Total cell extracts for pIκBα and nuclear extracts for p50 and p65 were used.α-tubulin expression was determined to validate equal sample loading. (D) Histograms of densitometric analysis normalized to α-tubulin expression showing complete inhibition of IR-induced IκBα phosphorylation and nuclear translocation of p50/p65 in hypoxic BCa cells with EF24, NLE, CUR, GEN, RES or RSE treatment. Group-wise comparisons were made using two-way ANOVA with Tukey’s post-hoc correction.