OBJECTIVES: To observe the effects of acupuncture on the KAT3B/ACSL4 pathway in the cerebral cortex of cerebral ischemia-reperfusion injury (CIRI) rats, and to explore the mechanisms by which acupuncture mitigates ferroptosis and ferritinophagy in CIRI.
METHODS: Forty male SD rats were randomly divided into sham operation, model, acupuncture, and western medicine groups, with 10 rats in each group. The CIRI rat model was established by middle cerebral artery occlusion. The rats in the acupuncture group received acupuncture at "Baihui" (GV20), "Sishencong" (EX-HN1), and "Shuigou" (GV26) acupoints, with the needles retained for 30 min once daily for 7 consecutive days. The rats in the western medicine group received intraperitoneal injections of 0.29 mL·100 g·dedaravone dexborneol for 7 consecutive days. Neurological deficit was assessed using the neurological deficit score. Infarct volume was measured by 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. Blood-brain barrier permeability was assessed by Evans blue assay. Hematoxylin-eosin (HE) staining and Nissl staining were applied to assess histopathological alterations in the ischemic cerebral cortex. TUNEL staining was used for apoptosis detection in the ischemic cerebral cortex. Immunofluorescence staining was performed to detect the expression levels of neuronal nuclei (NeuN), nuclear receptor coactivator 4 (NCOA4), and autophagy-related protein light chain 3B (LC3B) in the ischemic cerebral cortex. Dihydroethidium (DHE) fluorescence probe was used to measure reactive oxygen species (ROS) levels in the ischemic cerebral tissue. Colorimetric assay was used to quantify the contents of ferrous iron (Fe), lipid peroxidation (LPO), malondialdehyde (MDA), and glutathione (GSH), along with superoxide dismutase (SOD) activity in the ischemic cerebral tissue. Transmission electron microscopy was used to assess mitochondrial damage in neurons of the ischemic cerebral cortex. Western blot was performed to evaluate the protein expression levels of KAT3B, ACSL4, glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), NCOA4, transferrin receptor 1 (TfR1), ferritin heavy chain 1 (FTH1), and ferroportin-1 (FPN-1) in the ischemic cerebral tissue. -1 -1 2+
RESULTS: Compared with the sham operation group, the model group showed significant increases in neurological deficit score, cerebral infarction volume, and Evans blue permeability (<0.01);neuronal structure damage, with cell necrosis, nuclear shrinkage, loss of cytoplasm, and a decrease in Nissl bodies (<0.01). TUNEL-positive cells were increased (<0.01). Mitochondrial integrity was lost, mitochondria enlarged with cristae rupture and vacuolization. Expression of NeuN was decreased, while NCOA4 and LC3B expressions were increased (<0.01). In the ischemic cerebral tissue, contents of ROS, Fe, LPO, and MDA were increased, as well as the protein expression levels of KAT3B, ACSL4, NCOA4, and TfR1 were elevated (<0.01). By contrast, GSH content, SOD activity, and the expressions of GPX4, SLC7A11, FTH1, and FPN-1 were reduced (<0.01). Compared with the model group, the acupuncture and western medicine groups demonstrated reversal of these indicators (<0.05). There was no statistically significant difference between the acupuncture and western medicine groups. P P P P P P P2+
CONCLUSIONS: Acupuncture can improve cerebral injury in CIRI rats and reduce ferroptosis and ferritinophagy, potentially through inactivation of the KAT3B/ACSL4 pathway.
