Excessive phospholipid peroxidation distinguishes ferroptosis from other cell death modes including pyroptosis

Abstract

Lipid peroxidation (LPO) drives ferroptosis execution. However, LPO has been shown to contribute also to other modes of regulated cell death (RCD). To clarify the role of LPO in different modes of RCD, we studied in a comprehensive approach the differential involvement of reactive oxygen species (ROS), phospholipid peroxidation products, and lipid ROS flux in the major prototype modes of RCD viz. apoptosis, necroptosis, ferroptosis, and pyroptosis. LC-MS oxidative lipidomics revealed robust peroxidation of three classes of phospholipids during ferroptosis with quantitative predominance of phosphatidylethanolamine species. Incomparably lower amounts of phospholipid peroxidation products were found in any of the other modes of RCD. Nonetheless, a strong increase in lipid ROS levels was detected in non-canonical pyroptosis, but only during cell membrane rupture. In contrast to ferroptosis, lipid ROS apparently was not involved in non-canonical pyroptosis execution nor in the release of IL-1β and IL-18, while clear dependency on CASP11 and GSDMD was observed. Our data demonstrate that ferroptosis is the only mode of RCD that depends on excessive phospholipid peroxidation for its cytotoxicity. In addition, our results also highlight the importance of performing kinetics and using different methods to monitor the occurrence of LPO. This should open the discussion on the implication of particular LPO events in relation to different modes of RCD.

Lipid Metabolism and Ferroptosis

“…Withaferin A also induces ferroptosis with a general increase in the oxygenated form of most classes of phospholipids. Notably, PG and PI are increased the most among other classes [27].…”

“…In addition to PE, other phospholipids are also oxidized during ferroptosis [26,27]. In addition to PE, oxidized PS and PI are also elevated in ML162-induced ferroptosis in bone marrow-derived macrophages (BMDMs) [27]. Withaferin A also induces ferroptosis with a general increase in the oxygenated form of most classes of phospholipids.…”

The emerging role of ferroptosis in intestinal disease

“…Measuring lipid peroxidation is important for evaluating the presence of ferroptosis in specific contexts. Oxidative lipidomics is the gold standard for identifying specific oxidized lipids 28 , 29 . Probes such as C11-BODIPY and Liperfluo provide indirect but efficient means to detect lipid ROS 2 , 30 .…”

Role of Ferroptosis in Lung Diseases

“…17 The levels of lipid ROS can be determined using BODIPY-C11 dye. 18,19 Lipoxygenases (LOXs) may be involved in the formation of iron-dependent lipid ROS by catalysing the oxidation of PUFAs into lipid hydroperoxides and forming toxic lipidfree radicals in the presence of a large number of iron ions in the cytoplasm, which may eventually lead to cell damage. 20 Meanwhile, protons beside PUFAs can be transferred by these toxic lipid-free radicals and then start a new round of lipid oxidation reactions, which eventually lead to more severe oxidative damage.…”