Fig. 2.

Δ⁹-THC induces ER stress responses in HL-1 cells.

(A–D) Induction of ER stress genes. HL-1 cells were treated with the indicated concentrations of Δ⁹-THC and/or ethanol for 24 h. Levels of mRNAs of BIP, ATF4, ATF6 and CHOP were determined by qPCR. GAPDH was used as an internal control. Each graph shows mean ± S.D. (n = 3) **, p < 0.01; *, p < 0.05. (E–G) Increase in ER stress proteins and activation of caspases in HL-1 cells treated with the indicated concentrations of Δ⁹-THC and/or ethanol for 48 h. Levels of BIP, ATF4, ATF6 and CHOP (E and F) as well as the cleavage of caspase-12 and -3 (G) were determined by western blot analysis. Actin was also examined as a loading control. Each graph shows mean ± S.D. (n = 3) **, p < 0.01; *, p < 0.05. (H) ER stress inhibitor TUDCA alleviates Δ⁹-THC cytotoxicity. Cells were pretreated with 100 μM TUDCA, and further treated with the indicated concentrations of Δ⁹-THC and/or ethanol for 48 h. Cell viabilities were determined by modified MTT assay. Mean cell viability of control cells [Δ⁹-THC (-), ethanol (-)] was set to 100 %, and relative cell viabilities of Δ⁹-THC and/or ethanol treated cells are shown [mean ± S.D. (n = 5 or 6); **, p < 0.01].