Shear stress increases Nrf2 protein expression and induces Nrf2 translocation into nuclei. (A) Shear stress increased the total amount of Nrf2 protein. Human umbilical vein endothelial cells (HUVECs) were exposed to laminar shear stress at a magnitude of 12 dyne/cm2 for the indicated times. Total cell lysates were recovered and subjected to Western blot analysis with anti-Nrf2 and anti-tubulin (internal control, used to indicate equal loading of protein in each lane) antibodies. (B) Shear stress triggered the translocation and accumulation of Nrf2 protein in nuclei. HUVECs were exposed to laminar shear stress (12 dyne/cm2) for the indicated times. Nuclear extracts were isolated and subjected to Western blotting with anti-Nrf2 and anti-nucleolin (internal control) antibodies. (C) Shear stress increased Nrf2 mRNA. HUVECs were exposed to laminar shear stress (12 dyne/cm2) for the indicated times. Total RNA was isolated, and Nrf2 and GAPDH (internal control) mRNA levels were detected by RT-PCR. Results are representative of three independent experiments.