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

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.Palmitic or linoleic acid (250 μM) 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 μM) and palmitic acid (250 μM). Importantly, the release of cytochrome C from mitochondria into cytoplasm during cell apoptosis was specifically detected only when linoleic acid (125 μM), but not palmitic acid (250 μM), 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.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.Nonalcoholic fatty liver disease (NAFLD) is a multifactorial disease [1] that manifests as symptoms, ranging from mild steatosis, to nonalcoholic steatohepatitis and cirrhosis in the liver. Although NAFLD affects millions of people worldwide, its etiology remains unclear. However, hepatic inflammation and degeneration induced by the deposition of lipid droplets in the organ has been established as one of the major causes of the disease [1-3]. In particular, certain saturated fatty acids, such as palmitic acid, can induce endoplasmic reticulum (ER) stress and apoptosis in rat and human liver cell lines [4-9], leading to inflammation and/or degeneration in the liver. This hypothesis was further supported by the