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Resolving the Role of Lipoxygenases in the Initiation and Execution of Ferroptosis.

ABSTRACT: Lipoxygenases (LOXs) have been implicated as central players in ferroptosis, a recently characterized cell death modality associated with the accumulation of lipid hydroperoxides: the products of LOX catalysis. To provide insight on their role, human embryonic kidney cells were transfected to overexpress each of the human isoforms associated with disease, 5-LOX, p12-LOX, and 15-LOX-1, which yielded stable cell lines that were demonstrably sensitized to ferroptosis. Interestingly, the cells could be rescued by less than half of a diverse collection of known LOX inhibitors. Furthermore, the cytoprotective compounds were similarly potent in each of the cell lines even though some were clearly isoform-selective LOX inhibitors. The cytoprotective compounds were subsequently demonstrated to be effective radical-trapping antioxidants, which protect lipids from autoxidation, the autocatalytic radical chain reaction that produces lipid hydroperoxides. From these data (and others reported herein), a picture emerges wherein LOX activity may contribute to the cellular pool of lipid hydroperoxides that initiate ferroptosis, but lipid autoxidation drives the cell death process.

SUBMITTER: Shah R

PROVIDER: S-EPMC5879472 | biostudies-literature | 2018 Mar

REPOSITORIES: biostudies-literature

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Resolving the Role of Lipoxygenases in the Initiation and Execution of Ferroptosis.

Shah Ron R Shchepinov Mikhail S MS Pratt Derek A DA

ACS central science 20180207 3

Lipoxygenases (LOXs) have been implicated as central players in ferroptosis, a recently characterized cell death modality associated with the accumulation of lipid hydroperoxides: the products of LOX catalysis. To provide insight on their role, human embryonic kidney cells were transfected to overexpress each of the human isoforms associated with disease, 5-LOX, p12-LOX, and 15-LOX-1, which yielded stable cell lines that were demonstrably sensitized to ferroptosis. Interestingly, the cells could ...[more]

PMID: 29632885

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Project description:Introduction: Frankincense preparations are frequently used as traditional anti-inflammatory remedies in folk medicine with increasing popularity. Boswellic acids (BAs), especially 3-O-acetyl-11-keto-βBA (AKBA), are unique anti-inflammatory principles of frankincense, with multiple pharmacological actions and target proteins. We recently showed that AKBA favorably impacts lipid mediator (LM) networks in innate immune cells, by modulation of lipoxygenase (LOX) activities. Thus, AKBA binds to allosteric sites in 5-LOX, shifting the regiospecificity to a 12/15-lipoxygnating enzyme, and to an analogous site in 15-LOX-1, leading to enzyme activation, which favors specialized pro-resolving mediator (SPM) formation at the expense of leukotriene production. Methods: Here, we investigated Boswellin super® (BSR), a commercially available frankincense extract with ≥30% AKBA, used as remedy that approved efficacy in osteoarthritis trials, for its ability to modulate LM pathways in human monocyte-derived macrophage (MDM) phenotypes, neutrophils, and neutrophil/platelet co-incubations. LM profiling was performed by using targeted ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS). Results: BSR concentration-dependently (10-100 μg/ml) suppressed formation of pro-inflammatory 5-LOX products including LTB4 in exotoxin-stimulated M1-MDM and neutrophils, and strongly elevated 12/15-LOX products and SPM in activated M2-MDM and neutrophil/platelet cocultures, starting at 10 μg/mL. Also, BSR (≥10 μg/mL) induced robust 12/15-LOX product and SPM generation in resting M2-MDM, which was further markedly elevated when exogenous docosahexaenoic acid (DHA) and eicosahexaenoic acid (EPA) were supplied, and induced translocation of 15-LOX from a soluble to a particulate locale in M2 MDM. Discussion: We conclude that BSR especially when co-added with DHA and EPA, promotes the LM class switch in innate immune cells from pro-inflammatory to pro-resolving mediators, which might be a plausible mechanism underlying the anti-inflammatory actions of BSR.

Project description:BackgroundApoptosis is an essential cell death process throughout the entire life span of all metazoans and its deregulation in humans has been implicated in many proliferative and degenerative diseases. Mitochondrial outer membrane permeabilisation (MOMP) and activation of effector caspases are key processes during apoptosis signalling. MOMP can be subject to spatial coordination in human cancer cells, resulting in intracellular waves of cytochrome-c release. To investigate the consequences of these spatial anisotropies in mitochondrial permeabilisation on subsequent effector caspase activation, we devised a mathematical reaction-diffusion model building on a set of partial differential equations.ResultsReaction-diffusion modelling suggested that even if strong spatial anisotropies existed during mitochondrial cytochrome c release, these would be eliminated by free diffusion of the cytosolic proteins that instantiate the apoptosis execution network. Experimentally, rapid sampling of mitochondrial permeabilisation and effector caspase activity in individual HeLa cervical cancer cells confirmed predictions of the reaction-diffusion model and demonstrated that the signalling network of apoptosis execution could efficiently translate spatial anisotropies in mitochondrial permeabilisation into a homogeneous effector caspase response throughout the cytosol. Further systems modelling suggested that a more than 10,000-fold impaired diffusivity would be required to maintain spatial anisotropies as observed during mitochondrial permeabilisation until the time effector caspases become activated.ConclusionsMulti-protein diffusion efficiently contributes to eliminating spatial asynchronies which are present during the initiation of apoptosis execution and thereby ensures homogeneous apoptosis execution throughout the entire cell body. For previously reported biological scenarios in which effector caspase activity was shown to be targeted selectively to specific subcellular regions additional mechanisms must exist that limit or spatially coordinate caspase activation and/or protect diffusing soluble caspase substrates from unwanted proteolysis.

Dataset Information

Resolving the Role of Lipoxygenases in the Initiation and Execution of Ferroptosis.

Publications

Resolving the Role of Lipoxygenases in the Initiation and Execution of Ferroptosis.

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