Programmed cell death contributes to neurological damage in ischemic stroke, especially during the reperfusion stage. Several cell death pathways have been tested preclinically and clinically, including ferroptosis, necroptosis, and apoptosis. However, the sequence and complex interplay between cell death pathways during ischemia/reperfusion remains under investigation. Here, we unbiasedly investigated cell death pathways during ischemia/reperfusion by utilizing RNA sequencing analysis and immunoblot assays and revealed that ferroptosis and necroptosis occurred early post-reperfusion, followed by apoptosis. Ferroptosis inhibitor Liproxstatin-1 effectively inhibited necroptosis during reperfusion, while the necroptosis inhibitor Necrostatin-1 suppressed protein expression consistent with ferroptosis activation. Protein–protein interaction analysis and iron chelation therapy by deferoxamine mesylate indicate that iron is capable of promoting both ferroptosis and necroptosis in middle cerebral artery occlusion/repression modeled mice. Treatment of cells with iron led to a disruption in redox balance with activated necroptosis and increased susceptibility to ferroptosis. Collectively, these data uncovered a complex interplay between ferroptosis and necroptosis during ischemic stroke and indicated that multiple programmed cell death pathways may be targeted co-currently.

中文翻译：

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铁在缺血性卒中再灌注早期促进铁死亡和坏死性凋亡

程序性细胞死亡会导致缺血性中风的神经损伤，尤其是在再灌注阶段。几种细胞死亡途径已在临床前和临床上进行了测试，包括铁死亡、坏死性凋亡和细胞凋亡。然而，缺血/再灌注期间细胞死亡途径之间的顺序和复杂的相互作用仍在研究中。在这里，我们利用 RNA 测序分析和免疫印迹分析公正地研究了缺血/再灌注期间的细胞死亡途径，并揭示了再灌注后早期发生铁死亡和坏死性凋亡，随后发生细胞凋亡。铁死亡抑制剂 Liproxstatin-1 有效抑制再灌注期间的坏死性凋亡，而坏死性抑制剂 Necrostatin-1 抑制与铁死亡激活一致的蛋白表达。蛋白质-蛋白质相互作用分析和甲磺酸去铁胺的铁螯合疗法表明，铁能够促进大脑中动脉闭塞/抑制模型小鼠的铁死亡和坏死性凋亡。用铁处理细胞会导致氧化还原平衡被破坏，从而激活坏死性凋亡并增加铁死亡的易感性。总的来说，这些数据揭示了缺血性中风期间铁死亡和坏死性凋亡之间复杂的相互作用，并表明多种程序性细胞死亡途径可能同时被靶向。