Fig. 5

Esculetin disrupts Nrf2-KEAP1 interaction: a and b Western Blot analysis of PANC-1 protein extract immuno-precipitated using (a) Nrf2 and (b) KEAP1 antibody and probed with the other, showing loss of their interaction in esculetin treated cells. Input lane represents western blot analysis of 10 % of total protein extract used in CoIP indicating endogenous level of probed protein. c Western Blot analysis of total protein and nuclear extract from esculetin treated PANC-1 cells showing increase in phosphorylated form of Nrf2 and its nuclear accumulation. d Confocal microscopy view of esculetin treated PANC-1 cells incubated with Nrf2 antibody and probed with FITC (green) labeled secondary antibody showing an increase in nuclear accumulation of Nrf2. Nuclear staining was done with DAPI (blue). e Change in expression of NQO1, a target of Nrf2, measured using qPCR, showed about 5 fold increase upon 100 μM esculetin treatment. (C stands for control, VC stands for vehicle control, Iso stands for isotype Ab control, E stands for esculetin treated sample for indicated time). Data represents the mean ± SD of three independent experiments. The significance was determined using t test. Key:*p < 0.05; **p < 0.01; ***p < 0.001)