| Objective:To analyze the time-phase changes of Occludin,Claudin-5,ZO-1 and Caspase-3 after cerebral ischemia-reperfusion in rats and explore the protective effect of Netrin-1 on blood-brain barrier after cerebral ischemia-reperfusion in rats.Methods:Thirty six adult SD rats were randomly divided into 6 groups with 6 rats in each group.They were divided into sham operation group,MCAO12 h group,MCAO24 h group,MCAO48 h group,PBS group and NT-1 group.Modified Longa’s method was used to prepare a cerebral ischemia reperfusion model.After the operation,the neurological function and behavior of animals were scored by the Zea-Longa method.Identification model of HE staining for evaluating brain histopathological injury.The sham operation group only opened the carotid sheath without blocking treatment.Brain tissue samples were collected at 12 hours,24 hours and 48 hours after ischemia-reperfusion in MCAO group.The PBS group and NT-1group were injected with equal volume of PBS or Netrin-1 solution immediately after successful modeling.Brain tissue samples were collected 24 hours later.The Zea-Longa method was used to score the neurological functions and behaviors of animals,the wet and dry weight methods were used to measure the water content of brain tissue samples,and Western blot was used to detect the expression levels of Occludin,Claudin-5,ZO-1,and Caspase-3 in ischemic infarcted brain tissue specimens.Immunofluorescence was used to detect the expression and distribution of Occludin and Albumin in ischemic infarcted brain tissue specimens.Results:Compared with the sham group,the brain water content in the MCAO 12 h,24h,48 h groups showed an increasing trend.Brain water content in NT-1 group were lower than those in MCAO 24 h group.Compared with the MCAO 24 h,neurobehavioral score in NT-1 group were lower than those in MCAO 24 h group.The protein expressions of Occludin,Claudin-5and ZO-1 in MCAO groups decreased significantly compared with those in sham group,and Caspase-3 increased significantly.After successful building MCAO modles,Occludin,Claudin-5 and ZO-1 began to decrease with time,reaching the lowest at 48 h,Caspase-3increased significantly and reached the highest at 48 h.Compared with MCAO 24 h group,the expression of the Occludin,Claudin-5 and ZO-1 in NT-1 group decreased more slowly,.the expression of the Caspase-3 in NT-1 group increased more slowly.Compared with the proteins in the sham group,immunofluorescence showed that Occludin decreased significantly,Albumin increased significantly.Compared with MCAO 24 h group,the expression of the Occludin in NT-1 group decreased more slowly,the expression of the Albumin in NT-1 group increased more slowly.Conclusion:Occludin,Claudin-5 and ZO-1 showed a downward trend and Caspase-3 increased significantly after cerebral ischemia-reperfusion injury;Netrin-1 had a protective effect on blood-brain barrier permeability in cerebral ischemia-reperfusion injury. |
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