Knockdown of Gasdermin D protects hippocampal neurons by regulating both pyroptosis and ferroptosis in a rat model of status epilepticus.

BACKGROUND: Pyroptosis and ferroptosis may be implicated in status epilepticus (SE)-induced neuronal damage. This study aimed to elucidate the effects of Gasdermin D (GSDMD) on pyroptosis and ferroptosis after SE, and to determine whether GSDMD inhibition ameliorates SE-induced neuronal damage and cognitive dysfunction. METHODS: In the lithium-pilocarpine-induced SE rat model, we measured GSDMD and glutathione peroxidase 4 (GPX4) expression in the hippocampus via western blotting to evaluate the involvement of pyroptosis and ferroptosis after SE. After GSDMD knockdown by RNA interference, we performed Nissl staining to detect neuronal loss; quantitative reverse transcription polymerase chain reaction (qRT-PCR) to detect the hippocampal expressions of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α); Prussian blue staining to detect iron deposition; and western blotting and enzyme-linked immunosorbent assay to detect the expressions of the ferroptosis-related molecules GPX4, malonaldehyde (MDA), and glutathione (GSH). Finally, cognitive and behavioral functions were evaluated using the spontaneous-activity test and novel object-recognition test. RESULTS: Western blotting showed a marked increase in GSDMD expression (P < 0.050) and a notable decrease in GPX4 levels in (P < 0.050) the hippocampus of rats, providing evidence that both pyroptosis and ferroptosis are activated following SE. In addition, GSDMD knockdown significantly improved hippocampal neuronal survival and improved cognitive and behavioral functions of SE rats. Meanwhile, knockdown of GSDMD led to a significant reduction in the mRNA expression of IL-1β, IL-6, and TNF-α (pyroptosis products) (P = 0.021, 0.010, and 0.015, respectively), a decrease in iron deposition (P = 0.000) and MDA levels (ferroptosis product) (P = 0.000), and increased GSH and GPX4 expression (negative ferroptosis regulators) (P = 0.000 and 0.004, respectively). CONCLUSIONS: Pyroptosis and ferroptosis may jointly contribute to SE-induced neuronal damage. GSDMD may regulate both pyroptosis and ferroptosis, and inhibiting GSDMD could improve cognitive and behavioral function in SE rats.