Cancer cells and ischemic diseases exhibit unique metabolic responses and adaptations to energy stress. Forkhead box O 3a (FoxO3a) is a transcription factor that plays an important role in cell metabolism, mitochondrial dysfunction and oxidative stress response. Although the AMP-activated protein kinase (AMPK)/FoxO3a signaling pathway plays a pivotal role in maintaining energy homeostasis under conditions of energy stress, the role of AMPK/FoxO3a signaling in mitochondria-associated ferroptosis has not yet been fully elucidated. We show that glucose starvation induced AMPK/FoxO3a activation and inhibited ferroptosis induced by erastin. Inhibition of AMPK or loss of FoxO3a in cancer cells under the glucose starvation condition can sensitize these cells to ferroptosis. Glucose deprivation inhibited mitochondria-related gene expression, reduced mitochondrial DNA(mtDNA) copy number, decreased expression of mitochondrial proteins and lowered the levels of respiratory complexes by inducing FoxO3a. Loss of FoxO3a promoted mitochondrial membrane potential hyperpolarization, oxygen consumption, lipid peroxide accumulation and abolished the protective effects of energy stress on ferroptosis in vitro. In addition, we identified a FDA-approved antipsychotic agent, the potent FoxO3a agonist trifluoperazine, which largely reduced ferroptosis-associated cerebral ischemia-reperfusion (CIR) injuries in rats through AMPK/FoxO3a/HIF-1α signaling and mitochondria-dependent mechanisms. We found that FoxO3a binds to the promoters of SLC7A11 and reduces CIR-mediated glutamate excitotoxicity through inhibiting the expression of SLC7A11. Collectively, these results suggest that energy stress modulation of AMPK/FoxO3a signaling regulates mitochondrial activity and alters the ferroptosis response. The regulation of FoxO3a by AMPK may play a crucial role in mitochondrial gene expression that controls energy balance and confers resistance to mitochondria-associated ferroptosis and CIR injuries.

Microglial HO‐1 regulates iron metabolism in the brain. Intracerebral haemorrhage (ICH) shares features of ferroptosis and necroptosis; hemin is an oxidized product of haemoglobin from lysed red blood cells, leading to secondary injury. However, little is known about the underlying molecular mechanisms attributable to secondary injury by hemin or ICH. In this study, we first show that FoxO3a was highly co‐located with neurons and microglia but not astrocytes area of ICH model mice. Hemin activated FoxO3a/ATG‐mediated autophagy and HO‐1 signalling resulting in ferroptosis in vitro and in a mice model of brain haemorrhage. Accordingly, autophagy inhibitor Baf‐A1 or HO‐1 inhibitor ZnPP protected against hemin‐induced ferroptosis. Hemin promoted ferroptosis of neuronal cells via FoxO3a/ATG‐mediated autophagy and HO‐1 signalling pathway. Knock‐down of FoxO3a inhibited autophagy and prevented hemin‐induced ferroptosis dependent of HO‐1 signalling. We first showed that hemin stimulated microglial FoxO3a/HO‐1 expression and enhanced the microglial polarisation towards the M1 phenotype, while knockdown of microglial FoxO3a inhibited pro‐inflammatory cytokine production in microglia. Furthermore, the microglia activation in the striatum showed significant along with a high expression level of FoxO3a in the ICH mice. We found that conditional knockout of FoxO3a in microglia in mice alleviated neurological deficits and microglia activation as well as ferroptosis‐induced striatum injury in the autologous blood‐induced ICH model. We demonstrate, for the first time, that FoxO3a/ATG‐mediated autophagy and HO‐1 play an important role in microglial activation and ferroptosis‐induced striatum injury of ICH, identifying a new therapeutic avenue for the treatment of ICH.

Parkinson’s disease (PD) pathogenesis remains incompletely understood, with the causal link between gut microbiota dysbiosis and central neurodegeneration unclarified. This study aimed to unravel this complex interplay by integrating Mendelian randomization (MR) analysis, animal models, cellular experiments, and transcriptomic profiling. MR analysis identified Clostridium sensu stricto 1 and Candidatus Soleaferrea as novel genetic risk factors for PD, providing causal evidence for gut microbiota-driven PD pathogenesis. Using a chronic MPTP-induced PD mouse model, we confirmed that gut microbiota dysbiosis is accompanied by elevated tumor necrosis factor-α (TNF-α) in the intestine, plasma, and midbrain, along with dopaminergic neuron loss and motor deficits. Fecal microbiota transplantation (FMT) from healthy mice reversed these phenotypes, restoring gut homeostasis, reducing TNF-α levels, and preserving dopaminergic neurons. Transcriptomic analysis of mouse midbrain and human PD substantia nigra (GEO datasets) revealed ferroptosis as a key enriched pathway, with activating transcription factor 4 (ATF4) significantly upregulated in both PD models and patients. Mechanistically, TNF-α induced ATF4 overexpression in a concentration- and time-dependent manner, triggering dopaminergic neuron ferroptosis. This effect was mediated by TNF-α binding to TNFR1 (not TNFR2), activating the NF-κB pathway, and subsequently upregulating ATF4. Knockdown of ATF4 or inhibition of NF-κB reversed TNF-α-induced ferroptosis, as evidenced by restored glutathione (GSH) levels, reduced malondialdehyde (MDA) accumulation, and improved cell viability. Our findings establish a novel gut-brain axis-mediated pathogenic pathway: gut microbiota dysbiosis drives TNF-α-TNFR1-NF-κB-ATF4 signaling, leading to dopaminergic neuron ferroptosis and PD progression. This study bridges the gap between peripheral microbial dysbiosis and central neurodegeneration, identifies potential therapeutic targets, and highlights FMT as a promising intervention for PD.