Middle cerebral artery occlusion (MCAO) model

C57BL/6 mice (male, weighing 25-30 g) were anesthetized with 2.5% isofurane (RWD Life Science, China) and maintained in 2.5% isoflurane with 70% N₂ and 30% O₂. Under an operating microscope, a midline neck incision was made with scissors between the manubrium and the jaw. The right common carotid artery (CCA), external carotid artery (ECA), internal carotid artery (ICA), and vagus nerve were exposed. The filament was introduced into the right internal carotid artery up to the anterior cerebral artery to occlude the MCA and the anterior choroideal arteries. After the operation, the surgical site was sutured and re-perfused. Rectal temperature was maintained at 37 ± 0.5 °C with an electric blanket. For the sham groups, all procedures were identical except that the occluding monofilament was not inserted.