Figure 7

Basal ER stress determines sensitivity to CBD. (a) LX-2 cells were incubated in serum-free media with 0–4 μM CBD for 6 h in the presence or absence of tunicamycin (1 μg/mL) to induce ER stress. Cell death was detected by western blot analysis of cleaved PARP, expressed as fold over vehicle±S.E.M. for n=three independent experiments. (b) Total RNA was extracted from whole cell lysates of either ethanol-fed or control rat HSCs and reverse transcribed to cDNA. Real-time PCR was performed with gene-specific primers for ATF4, ATF6, and CHOP to determine the relative basal mRNA levels, normalized to GAPDH and expressed as fold over control. ^***P<0.001 (Student's t-test). (c) ER stress was induced in HSCs from control rats by incubation with the indicated concentrations of tunicamycin in the presence or absence of 5 μM CBD for 6 h in serum-free media. Cell death and ER stress were detected by western blot analysis of PARP and CHOP expression, respectively. Cleaved PARP is expressed as fold over control±S.E.M. for n=three independent experiments. ^*P<0.05; ^**P<0.01