Skip to main content
. Author manuscript; available in PMC: 2017 Aug 1.
Published in final edited form as: Toxicol Appl Pharmacol. 2016 May 12;304:90–98. doi: 10.1016/j.taap.2016.05.007

Fig. 4. Participation of Nrf2 in MRP2 modulation by BZL in HepG2 cells.

Fig. 4

Purity of nuclear and cytosolic fractions was checked assessing H1 Histone and GAPDH enrichment, respectively (a). Nrf2 expression was quantified in nuclear extracts (b) and cytosolic fraction (c) of BZL treated HepG2 cells (200 μM; 3, 24 and 48 h). Equal amounts of protein were loaded in the gels. Nrf2 O.D. was normalized to histone H1 and GAPDH O.D. in nuclear and cytosolic fractions, respectively. Uniformity of loading and transfer from gel to PVDF membrane was also determined by Ponceau S staining. Firefly luciferase activity was assessed in HepG2 cells transfected with pGL3-ARE and exposed to BZL (200 μM; 3, 24 and 48 h) (d). Results (mean ± S.D., n = 3-4) are expressed as percentage of control values (C). * different from C, p<0.05. For a final confirmation of Nrf2 participation, an Nrf2 knock-down model was established. e: Nrf2 expression was assessed in total lysates of Nrf2+ (transfected with a non-silencing siRNA) and Nrf2− HepG2 cells (transfected with a pool of specific siRNAs against human Nrf2) and normalized to GAPDH expression. Results (mean ± S.D., n = 3) are expressed as percentage of the Nrf2/GAPDH ratio in Nrf2+ cells. * different from Nrf2+, p<0.05. f: MRP2 expression was assessed in control and BZL-treated (200 μM, 24 h) Nrf2+ and Nrf2− cells. Results (mean ± S.D., n = 3) are expressed as percentage of the Nrf2/GAPDH ratio in the corresponding control cells. * different from control (C), p<0.05.