Influence of Sevoflurane Postconditioning on Hypoxic-Ischemic Brain Injury via Nrf2-Regulated Ferroptosis in Neonatal Rats

BACKGROUND: The mechanisms by which sevoflurane protects the brain from hypoxic-ischemic brain injury (HIBI) are unknown. Ferroptosis occurs during HIBI and is regulated by the nuclear factor erythroid 2-related factor 2 (Nrf2). This study investigated the roles of Nrf2-regulated ferroptosis in sevoflurane postconditioning (SPC)-mediated neuroprotection during HIBI. METHODS: HIBI was induced in 7-day-old rats. SPC (2.5%, 30 minutes) was performed immediately after HIBI, and some rats were injected with ML385 (an Nrf2-inhibitor) 30 minutes before HIBI. Ferroptosis was evaluated by measuring glutathione peroxidase 4 (GPx4), solute carrier family 7 member 11 (SLC7A11, also known as xCT), glutathione (GSH), cysteine, iron, malondialdehyde (MDA) levels, and mitochondrial morphology. Nrf2 and heme oxygenase-1 (HO-1) expression were determined to explore the signaling pathways involved in SPC-mediated neuroprotection. Brain morphology, left/right hemisphere weight ratios, and Nissl staining were measured to assess brain damage. The Morris water maze was conducted to assess long-term learning and memory abilities. RESULTS: SPC alleviated HIBI-induced cysteine depletion-induced (HIBI versus SPC, xCT/β-tubulin ratio: −0.435 [95% CI, −0.727 to −0.143], P = .003; Cysteine (% of Sham): −29.8 [95% CI, −39.4 to −20.2], P < .001; GSH (% of Sham): −46.5 [95% CI, −54.6 to −38.4], P < .001) and GPx4 inhibition-induced ferroptosis (HIBI versus SPC, GPx4/β-tubulin ratio: −0.287 [95% CI, −0.514 to −0.0603], P = .01). Compared with the HIBI group, the SPC group showed improved learning and memory abilities (HIBI versus SPC, platform crossings: −4 times [95% CI, −7 to −1], P = .002; escape latency: 46 seconds [95% CI, 24 to 68], P < .001), reduced brain damage (HIBI versus SPC, weight ratio of left/right cerebral hemispheres: −13.1 [95% CI, −15.7 to −10.4], P < .001; neuronal density ratio: −0.450 [−0.620 to −0.280], P < .001), and increased Nrf2 and HO-1 protein levels (HIBI versus SPC, Nrf2/β-tubulin ratio: −1.89 [95% CI, −2.82 to −0.970], P < .001; HO-1/β-tubulin ratio: −1.08 [95% CI, −1.73 to −0.442], P < .001). Inhibiting Nrf2 via ML385 partly reversed SPC-mediated neuroprotection (SPC versus SPC+ML385, weight ratio of left/right cerebral hemispheres: 12.4 [95% CI, 9.73–15.1], P < .001; neuronal density ratio: 0.412 [95% CI, 0.242–0.582], P < .001), accompanied by decreased HO-1 expression (SPC versus SPC+ML385, HO-1/β-tubulin ratio: 1.70 [95% CI, 1.05–2.34], P < .001). CONCLUSIONS: SPC inhibits both cysteine depletion- and GPx4 inhibition-induced ferroptosis by regulating Nrf2/HO-1 signaling to protect against HIBI.