CSE exposure potentiates oxidative stress responses in BBB endothelial cells. Briefly, confluent hCMEC/D3 cell monolayers were exposed to media containing nicotine only (100 μg/mL) or CSEs from full flavor (3R4F) or ULN tobacco products containing nicotine equivalent to 100 ng/mL). Fresh media without nicotine or mainstream TS served as controls. a Immunofluorescence analysis of cellular ROS levels (an indicator of oxidative stress load) were determined by using CellROX® Green reagent following exposure to CSE or nicotine treatment (3 h); b gene array based analysis of Nrf2 gene expression changes was determined after control, nicotine or TS exposure (n = 6); c Nrf2 mRNA expression was quantified by RT-PCR using primer specific sequences (n = 4); d Nrf2 expression and nuclear translocation following exposure to different conditions at early time point of 8 h as analyzed by western blotting. Representative western blots were shown with actin as a loading control. Data were expressed as mean ± SEM (fold change over control; n = 4 biological replicates). *p < 0.05 vs. control.