MCAO Model

Transient middle cerebral artery occlusion (MCAO) model in rat was performed as described previously [23]. Briefly, 3 days after injection, the rats were anesthetized with 4% isoflurane in 70% N₂O/30% O₂ using a mask. A 4-0 nylon suture with silicon was inserted into the internal carotid artery and gently advanced to occlude the middle cerebral artery (MCA). The body temperature was carefully monitored during the surgery and maintained at 37.0 ± 0.5 °C using a heating pad. After 2 h of MCA occlusion (MCAO), the suture was carefully removed to restore blood flow, and rats score <1 according to Zea-Longa five-point scale (0, no deficit; 1, failure to extend right paw; 2, circling to the right; 3, falling to the right; and 4, unable to walk spontaneously) were excluded from the study.