The cerebral infarct volume and integrity of the BBB were assessed by TTC and Evans blue staining, respectively, as previously described (38 (link)). Briefly, Evans blue dye (2%; 4 ml/kg) was injected into the left femoral vein and allowed to circulate for 60 sec before the rats were euthanized with 1% pentobarbital (40 mg/kg) and perfused transcardially with PBS for 60 sec at 37°C. The brains were removed, washed with PBS and kept at −20°C for 30 min before sectioning. Coronal sections (2 mm) were cut in a rat brain matrix and stained with 2% TTC (Amresco, LLC) for 30 min at 37°C. Following staining, the brain slices were fixed with 4% paraformaldehyde for 10 min at 37°C to conserve the area stained by TTC. The stained brain sections were digitally photographed using a digital camera (PowerShot G7 X Mark II Canon, Inc.). The infarcted area (white) and BBB disruption (blue) of each brain section was measured using Image-Pro Plus 6.0 software (Media Cybernetics, Inc.). The infarcted volume and the BBB disrupted area were calculated according to the formula: 100% × (ipsilateral volume-contralateral volume)/contralateral volume.