Fig. 5

Autophagy and occludin degradation in brain capillaries from rats with ischemic stroke by MCAO. a–c Ischemic stroke was induced in rats by middle cerebral artery occlusion (MCAO). At 24 h after MCAO, rats were sacrificed by transcardial perfusion, and brain capillaries were isolated from both the ischemic and non-ischemic hemispheres. 3-MA (0.3 mg/kg) or sterile saline solution was administrated to rats at 30 min prior to MCAO. N = 5–9. a Cerebral blood flow (CBF) changes were monitored before and after MCAO (Right). TTC-stained brain sections confirmed the ischemic brain damage (Middle). The protein levels of endothelial and neuronal marker (CD31 and NeuN, respectively) in the whole brain homogenates and isolated brain capillary were measured (Left). b Autophagic activation in brain capillaries was examined based on the level of LC3-II. The conversion ratio of LC3-II/I was calculated. c The amount of occludin protein in the brain capillaries was detected by western blot. Protein levels were normalized with the corresponding β-actin levels. d 2% Evans blue (4 mL/kg) was intravenously injected at 20 h after MCAO. The rats were transcardially perfused to remove any remaining Evans blue in blood and sacrificed at 24 h after MCAO. The amount of Evans blue was determined in ipsilateral or contralateral hemisphere. N = 5–7. Data are expressed as the mean ± SEM. *p < 0.05 and **p < 0.01 vs. sham-operated rats, ^#p < 0.05 vs. rats with MCAO without 3-MA treatment