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A New Model for Raf Kinase Inhibitory Protein Induced Chemotherapeutic Resistance

Figure 2

Antioxidant treatments rescue KEAP 1 in RKIP –depleted HEK-499 cells.

A, Upper panels show confocal images of HEK-293 and 499 stained with dihydro-ethidium bromide (DHE) as an indicator of ROS before and after treatments with 5 mM N-Acetylcysteine (N-Ac). The lower bar chart represents the quantification of cell-number-normalized DHE intensities in depicted cells. B, Upper panel represents western blotting for KEAP 1 and β-actin loading control in HEK-499 cells before and after treatment of cells with the antioxidants N-Ac and/or ALA. The lower western panel depicts KEAP1 and RKIP protein levels in HEK-499 and HEK-293 after the indicated treatment. C, Densitometric measurements of β-actin normalized KEAP 1 expression in an additional independent experiment after treatment of cells with 5 mM of n-Ac for 16 hours, showing the rescue of KEAP 1 protein and concomitant reduction of total NRF2 in the treated HEK-499 cells. Western experiments were performed at least in duplicates and repeated twice. Asterisks indicate statistical significance (p<0.05 compared to equivalent controls).

Figure 2