FIG. 6.

Effect of ORAI1 knockdown on usnic acid-induced ATP depletion and ER stress in HepG2 cells. HepG2 cells stably expressing doxycycline-inducible sh-ORAI1 or a scramble control were used. These two cell lines were incubated with doxycycline for 72h followed by continued culture for another 24 h without doxycycline; then the effect of ORAI1 silencing was assessed by real-time PCR (A) and Western blotting (B). Values were mean6SD of three individual experiments *P<0.05 as compared with the SC. C, sh-ORAI1 and scramble control cells were incubated with doxycycline for 72h and then treated with usnic acid at indicated concentration for another 6h without doxycycline. Treated cells were then analyzed for ATP content. The bar graph represents the mean6SD of three separate experiments.*P<0.05 compared with the SCs. D, sh-ORAI1 and scramble control cells were treated with usnic acid at 25 µM for 24 h. Total cellular proteins were extracted. Western blotting was performed using antibodies for CHOP and GAPDH.