Ferroptosis, triggered by discoordination of iron, thiols and lipids, leads to accumulation of 15-hydroperoxy-arachidonoyl-PE (15-HpETE-PE) generated by complexes of 15-lipoxygenase (15-LOX) and a scaffold protein, PEBP1. As Ca 2+ -independent phospholipase PLA 2 (iPLA 2 β, PLA2G6/PNPLA9 gene), can preferentially hydrolyze peroxidized phospholipids, it may eliminate ferroptotic 15-HpETE-PE death signal. Here we demonstrate that by hydrolyzing 15-HpETE-PE, iPLA 2 β averts ferroptosis whereas its genetic or pharmacological inactivation sensitizes cells to ferroptosis. Given that PLA2G6/PNPLA9 mutations relate to neurodegeneration, we examined fibroblasts from a patient with a Parkinson’s disease (PD)-associated mutation fPD R747W and found selectively decreased 15-HpETE-PE hydrolyzing activity, 15-HpETE-PE accumulation and elevated sensitivity to ferroptosis. CRISPR-CAS9-engineered PNPLA9 R748W/R748W mice exhibited progressive parkinsonian motor deficits and 15-HpETE-PE accumulation. Elevated 15-HpETE-PE levels were also detected in midbrains of rotenone-infused parkinsonian rats and α-synuclein mutant SNCA-A53T mice with decreased iPLA 2 β expression and PD-relevant phenotype. Thus, iPLA 2 β is a new ferroptosis regulator and its mutations may be implicated in PD pathogenesis.

Skin cells are vulnerable to oxidative stress-induced senescence, which may lead to abnormal aging or aging-related disorders. Therefore, strategies that can ameliorate oxidative stress-induced senescence are expected to protect skin from damage, holding the promise of treating skin diseases in the clinic. This study aims to investigate whether caffeine, a well-known purine alkaloid, is able to prevent skin from oxidative stress-induced senescence, and to explore the underlying molecular mechanisms.Methods: A free radical inducer 2,2'-Azobis (2-amidinopropane) dihydrochloride (AAPH) was used to induce oxidative stress and cellular senescence in both transformed skin cells and in normal human epidermal keratinocytes (NHEKs). Ultraviolet (UV) irradiation was established as the in vivo oxidative stress model in mouse skin tissues. Cellular senescence was determined by SA β-galactosidase staining, immunofluorescence and western blotting. Activation of autophagy was confirmed by western blotting, immunofluorescence, and transmission electron microscopy. Reactive oxygen species (ROS) detection by commercial kits, gene knockdown by RNA interference (RNAi) and receptor activation/inactivation by agonist/antagonist treatment were applied in mechanistic experiments.Results: We report that AAPH induced senescence in both transformed skin cells and in NHEKs. Similarly, UV irradiation induced senescence in mouse skin tissues. Remarkably, low dose of caffeine (<10 μM) suppressed cellular senescence and skin damage induced by AAPH or UV. Mechanistically, caffeine facilitated the elimination of ROS by activating autophagy. Using a combination of RNAi and chemical treatment, we demonstrate that caffeine activates autophagy through a series of sequential events, starting from the inhibition of its primary cellular target adenosine A2a receptor (A2AR) to an increase in the protein level of Sirtuin 3 (SIRT3) and to the activation of 5' adenosine monophosphate-activated protein kinase (AMPK). Oral administration of caffeine increased the protein level of SIRT3, induced autophagy, and reduced senescence and tissue damage in UV-irradiated mouse skin. On the other hand, co-administration with autophagy inhibitors attenuated the protective effect of caffeine on UV-induced skin damage in mice.Conclusion: The results reveal that caffeine protects skin from oxidative stress-induced senescence through activating the A2AR/SIRT3/AMPK-mediated autophagy. Our study not only demonstrated the beneficial effect of caffeine using both in vitro and in vivo models, but also systematically investigated the underlying molecular mechanisms. These discoveries implicate the potential of caffeine in the protection of skin disease.

During cancer therapy, phagocytic clearance of dead cells plays a vital role in immune homeostasis. The nonapoptotic form of cell death, ferroptosis, exhibits extraordinary potential in tumor treatment. However, the phagocytosis mechanism that regulates the engulfment of ferroptotic cells remains unclear. Here, we establish a novel pathway for phagocytic clearance of ferroptotic cells that is different from canonical mechanisms by using diverse ferroptosis models evoked by GPX4 dysfunction/deficiency. We identified the oxidized phospholipid, 1-steaoryl-2-15-HpETE-sn-glycero-3-phosphatidylethanolamine (SAPE-OOH), as a key eat-me signal on the ferroptotic cell surface. Enriching the plasma membrane with SAPE-OOH increased the efficiency of phagocytosis of ferroptotic cells by macrophage, a process that was suppressed by lipoprotein-associated phospholipase A2. Ligand fishing, lipid blotting, and cellular thermal shift assay screened and identified TLR2 as a membrane receptor that directly recognized SAPE-OOH, which was further confirmed by TLR2 inhibitors and gene silencing studies. A mouse mammary tumor model of ferroptosis verified SAPE-OOH and TLR2 as critical players in the clearance of ferroptotic cells in vivo. Taken together, this work demonstrates that SAPE-OOH on ferroptotic cell surface acts as an eat-me signal and navigates phagocytosis by targeting TLR2 on macrophages.

No abstract