FA showed strong cytotoxicity via inhibiting the cell growth and even causing cell death at certain concentrations. Toxicity of FA on cells depends on the concentration, solvent, cell type and culture condition, which lead to different FA concentrations that were used in different studies. In our study, the FA that used in stimulating HepG2 cells was prepared via saponification and albumin binding, so that the toxicity of DMSO or ethanol on cell viability was avoided. MTT cell viability assay revealed that the optimal FA concentrations for inducing HepG2 cells steatosis without significant inhibition in cell viability were 0.25, 0.5, and 1 mM (Figure 3A). Nrf2 and CYP2A6 expression induced by 18-carbon FA (SA, OA, LA, and ALA) in vitro (Figures 3B–F) showed the same tendency as the experiments in vivo (Figure 2), which indicated that the experiment in vitro reproduced the process in vivo to a certain extent. Interestingly, Nrf2 mRNA and protein expressions were dose-dependently increased in HepG2 cells exposed to SA, OA, and ALA, but dose-dependently decreased in HepG2 cells exposed to LA, although still much higher than
