Mitochondrial ROS Inhibit Unfolded Protein Response Induced by Inflammatory Mediators in Liver Cells

MK886, an inhibitor of 5-lipoxygenase activating protein (FLAP), and the lipoxygenase (LOX) inhibitors baicalein and nordihydroguaiaretic acid (NDGA), induce apoptosis by mechanisms independent of both LOX and FLAP. One possible mechanism for these agents is through an effect on the binding of fatty acids to LOX and fatty acid binding proteins resulting in increased intracellular levels of unbound fatty acids, particularly arachidonic acid (AA), that in turn, activate apoptosis signaling pathways either directly or following oxidation. In FL5.12 murine pro-B lymphocytic cells, exogenous fatty acids induced apoptosis proportional to their degree of unsaturation. MK886, baicalein, and NDGA significantly enhanced the release of [3H]-AA two to threefold within 2 h and induced apoptosis by 8 h. Neither MK886-induced AA release, nor apoptosis were affected by quinacrine, a phospholipase A2 inhibitor. The presence of peroxides 1 h after treatment of FL5.12 cells with these agents was evident by a two to threefold increase in the ferrous oxidation–xylenol orange (FOX) assay as well as dichlorofluorescein fluorescence measured with flow cytometry. Isoprostane formation, an additional index of lipid peroxidation, was increased threefold by 2 h, and fourfold at 4 h after MK886 or baicalein, but not after NDGA. Antioxidants were able to protect against NDGA-induced apoptosis but had no effect on baicalein and resulted in enhanced apoptosis with MK886. These data support the hypothesis that release of fatty acids and generation of oxidized species contribute to apoptosis induced by these LOX inhibitors, but that more complex mechanisms are likely involved.