Ferroptosis, a kind of programmed cell death, is characterized with iron-dependent lipid ROS buildup, which is considered as an important cellular immunity in resisting intracellular bacterial infection in mammalian macrophages. In this process, lipid ROS oxidizes the bacterial biofilm to inhibit intracellular bacteria. However, the function of ferroptosis in invertebrate remains unknown. In this study, the existence of ferroptosis in Apostichopus japonicus coelomocytes was confirmed, and its antibacterial mechanism was investigated. First, our results indicated that the expression of glutathione peroxidase (AjGPX4) was significantly inhibited by 0.21-fold (p < 0.01) after injecting A. japonicus with the ferroptosis inducer RSL3, and the contents of MDA (3.93-fold, p < 0.01), ferrous iron (1.40-fold, p < 0.01), and lipid ROS (3.10-fold, p < 0.01) were all significantly increased under this condition and simultaneously accompanied with mitochondrial contraction and disappearance of cristae, indicating the existence of ferroptosis in the coelomocytes of A. japonicus. Subsequently, the contents of ferrous iron (1.40-fold, p < 0.05), MDA (2.10-fold, p < 0.01), ROS (1.70-fold, p < 0.01), and lipid ROS (2.50-fold, p < 0.01) were all significantly increased, whereas the mitochondrial membrane potential and GSH/GSSG were markedly decreased by 0.68-fold (p < 0.05) and 0.69-fold (p < 0.01) under Vibrio splendidus (AJ01) infection. This process could be reversed by the iron-chelating agent deferoxamine mesylate, which indicated that AJ01 could induce coelomocytic ferroptosis. Moreover, the results demonstrated that the intracellular AJ01 load was clearly decreased to 0.49-fold (p < 0.05) and 0.06-fold (p < 0.01) after treating coelomocytes with RSL3 and ferrous iron, which indicated that enhanced ferroptosis could inhibit bacterial growth. Finally, subcellular localization demonstrated that ferrous iron efflux protein ferroportin (AjFPN) and intracellular AJ01 were co-localized in coelomocytes. After AjFPN interference (0.58-fold, p < 0.01), the signals of ferrous iron and lipid ROS levels in intracellular AJ01 were significantly reduced by 0.38-fold (p < 0.01) and 0.48-fold (p < 0.01), indicating that AjFPN was an important factor in the introduction of ferroptosis into intracellular bacteria. Overall, our findings indicated that ferroptosis could resist intracellular AJ01 infection via AjFPN. These findings provide a novel defense mechanism for aquatic animals against intracellular bacterial infection. Copyright © 2024 Elsevier Ltd. All rights reserved.
Chengyang Wang, Yangxi Xiang, Yina Shao, Chenghua Li. Ferroptosis resists intracellular Vibrio splendidus AJ01 mediated by ferroportin in sea cucumber Apostichopus japonicus. Fish & shellfish immunology. 2024 Jun;149:109585
PMID: 38663462
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