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Hypoxic–ischemic brain damage (HIBD) is a predominant cause of neuronal injury and mortality in newborns. Current preventive and therapeutic interventions demonstrate limited clinical efficacy. Emerging evidence reveals ferroptosis as a critical mechanism within HIBD pathophysiology, positioning it as a promising therapeutic target. Dimethyl malonate (DMM), a competitive inhibitor of succinate dehydrogenase, has demonstrated neuroprotective properties across multiple models of neurological disorders. However, the impact of DMM on the neonatal HIBD has not been studied. To investigate the neuroprotective effects of DMM against neonatal HIBD and elucidate its mechanisms of action. We created a model of HIBD in neonatal male C57BL/6J mice and administered various doses of DMM or vehicle control. Quantitative assessments included cerebral infarct volume measurement, Nissl staining for neurons, neurological behavior, ferrous ion (Fe 2+ ), malondialdehyde (MDA) level, 4-hydroxynonenal (4-HNE) expression, and solute carrier family 7 member 11 (SLC7A11, system Xc − )/glutathione peroxidase 4 (GPX4) antioxidant axis expression level. Parallel studies in vitro employed oxygen-glucose deprivation/reperfusion-treated HT22 cells to investigate the effects of DMM on ferroptosis and its underlying mechanisms. Moreover, key factors of ferritinophagy, including nuclear receptor coactivator 4 (NCOA4), SQSTM1/p62, ferritin heavy chain 1 (FTH1), and microtubule-associated protein light 3 II (LC3II) were analyzed by western blotting. Molecular interactions between NCOA4 and FTH1 in brain cortical tissues of DMM-treated HIBD mice were analyzed by coimmunoprecipitation (Co-IP). Ferroptosis regulation by DMM was further investigated via Fth1 knockdown in cellular models. Immunofluorescence staining was used to evaluate the capacity of DMM to suppress ferritin degradation and lysosomal Fe 2+ accumulation at the organelle level. DMM treatment demonstrated its neuroprotective efficacy in HIBD models, as evidenced by a reduction in cerebral infarct volume, an increase in the number of Nissl-positive neurons, and improved cognitive and motor functions in neonatal mice compared with controls. Additionally, the DMM intervention significantly modulated ferroptosis-related biomarkers in brain cortical tissues and HT22 cells, decreasing ferrous ion (Fe 2+ ) accumulation, reducing lipid peroxidation products (MDA and 4-HNE), and enhancing SLC7A11/GPX4 antioxidant system activity. Importantly, DMM specifically regulated core ferritinophagy components: suppressing NCOA4 and LC3II expression while upregulating FTH1 and p62 levels. Co-IP revealed that mechanistically, DMM disrupted the protein interaction between NCOA4 and FTH1, effectively inhibiting ferritinophagy progression. The effects of antiferroptosis were FTH1-dependent, as demonstrated by reversal of the DMM protective effect following Fth1 knockdown in vitro. Immunofluorescence analysis showed that DMM decreased the colocalization of FTH1-lysosome-associated membrane protein 2, and FerroOrange/LysoTracker Green dual staining confirmed its inhibition of lysosomal iron accumulation, collectively indicating regulation of DMM at the organelle level. DMM suppressed ferroptosis-induced neuronal death by specifically targeting FTH1 and disrupting the NCOA4-FTH1 interaction, thereby mitigating HIBD. These findings position DMM as a promising therapeutic candidate for the clinical management of neonatal hypoxic–ischemic encephalopathy. • Dimethyl malonate (DMM) protects against neonatal hypoxic–ischemic brain damage. • DMM alters ferroptosis in mouse and HT22 models of hypoxic–ischemic brain damage. • DMM reduces neuronal ferroptosis by suppressing ferritinophagy. • DMM inhibits ferroptosis by targeting FTH1 to disrupt NCOA4-FTH1.
Jin et al. (Tue,) studied this question.