Palmitic and linoleic acids induce ER stress and apoptosis in hepatoma cells

Objectives: Hepatic inflammation and degeneration induced by lipid depositions may be the major cause of nonalcoholic fatty liver disease. In this study, we tried to investigate the effects of saturated and unsaturated fatty acids on hepatoma cell apoptosis. Methods: H4IIE liver cells were treated with palmitic acid, linoleic acid, or both with or without the calcium-specific chelator BAPTA-AM after which the expression of proteins associated with endoplasmic reticulum (ER) stress, apoptosis, caspase-3 levels, and calcium flux were measured. Results: Palmitic or linoleic acid (250 mu M) induced H4IIE cell apoptosis, which required calcium flux but not caspase-3. Apoptosis was not observed when cells were co-treated with linoleic acid (125 mu M) and palmitic acid (250 mu M). Importantly, the release of cytochrome C from mitochondria into cytoplasm during cell apoptosis was specifically detected only when linoleic acid (125 mu M), but not palmitic acid (250 mu M), was added to the cells. Depletion of intracellular calcium flux by the calcium-specific chelator, BAPTA-AM, abolished linoleic acid-induced apoptosis. Moreover, in the presence of BAPTA-AM, expression of the unfolded protein response (UPR)-associated genes, CHOP, GRP78, and GRP94, was induced by linoleic acid, but not palmitic acid. Conclusions: The results suggest that linoleic acid promotes cell apoptosis through the release of cytochrome C, only if the intracellular calcium flux is unperturbed and intact. These results confirm that ER stress contributes to fatty acid-induced liver cell apoptosis.