To explore the preventive effect of electroacupuncture pretreatment on stroke in rats by inhibiting ferroptosis and oxidative stress. Rats were randomly assigned to the sham, middle cerebral artery occlusion/reperfusion (MCAO/R), MCAO/R + EP, MCAO/R + EP + erastin, and MCAO/R + EP + ferrostatin 1 groups. Daily electroacupuncture was performed 2 weeks before establishing the MCAO/R model utilizing the modified Zea Longa suture method. Rats were sacrificed 1 day after reperfusion, and brain tissues were collected. They were prepared for hematoxylin and eosin staining, prussian blue staining, transmission electron microscope. Measurement of total iron levels using a commercial kit, detection of malondialdehyde (MDA) and superoxide dismutase (SOD) levels by ELISA, and examination of 15-lox2, GPX4, SLC7A11, ACSL4, and TFR1 by western blotting. Compared with sham rats, cerebral infarction size was dramatically larger in MCAO/R rats. Moreover, the MCAO/R group displayed damaged mitochondria with a disarranged structure of cristae; free iron, total iron levels, and oxidative stress were significantly higher. Cerebral pathological lesions, oxidative stress, total iron levels, and protein levels of ACSL4, TFR1, and 15-lox2 were significantly reduced in the MCAO/R + EP and MCAO/R + EP + ferrostatin 1 groups, while the protective effect of electroacupuncture pretreatment on cerebral ischemia-reperfusion injury was inhibited by treatment with the ferroptosis activator erastin. Electroacupuncture pretreatment can protect rats from cerebral ischemia-reperfusion injury by reducing the area of cerebral infarction and inhibiting ferroptosis and oxidative stress.

We aimed to explore the influence of ferroptosis on an oxygen-glucose deprivation/reoxygenation (OGD/R) model in primary rat microglia. Primary microglia were extracted from rats and cultured in vitro. The cells were subjected to a hypoxic environment for 6 h in a glucose-free medium, and then re-oxygenated for 24 h in DMEM/F12. Rat microglia were pretreated with the ferroptosis activator erastin and the ferroptosis inhibitor ferrostatin 1 for 24 h, followed by detection of cell cycle progression and apoptosis by flow cytometry. Intracellular total iron levels were measured. In addition, the relative levels of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD) were determined using enzyme-linked immunosorbent assay. The protein levels of 15-lox2, GPX4, SLC7A11, ACSL4, and TFR1 were examined by western blotting. Compared with rat microglia subjected to OGD/R, pretreatment with erastin did not influence cell apoptosis but significantly enhanced total iron levels, MDA, and ROS levels, whereas it reduced SOD levels. Moreover, it upregulated ACSL4, TFR1, and 15-lox2 and downregulated GPX4 and SLC7A11. Pretreatment with ferrostatin 1 significantly inhibited cell apoptosis and cell cycle arrest in the G0/G1 phase. It significantly reduced total iron levels, MDA, and ROS levels and enhanced SOD levels, which also downregulated ACSL4, TFR1, and 15-lox2, and upregulated GPX4 and SLC7A11. Our study showed that inhibition of ferroptosis is favorable against potential OGD/R-induced damage in rat microglia.