The Effect Of Ischemic Postconditioning On The Expression Of Notch1/Cell Cycle Signaling In The Hippocampus Of Global Cerebral Ischemia Rats

Objectives 1 To investigate the effect of ischemic postconditioning on the cell cycle in the hippocampus of rats with global cerebral ischemia; 2 To investigate the effect of ischemic postconditioning on the activity of Notch1.Methods 128 healthy male SD rats were divided into four groups randomly: sham group(n=32), global cerebral ischemia group(n=32), ischemic postconditioning group(n=32),inhibitor group(n=32). The improved four-vessel occlusion was applied to manufacture the global cerebral ischemia model; Ischemic postconditioning was produced by repeated three cycles of release/occlusion of the blood vessels before thorough reperfusion.The rats in inhibitor group were intraperitoneal inject DAPT at 3h by 10mg/(kg�d) before the preparation of ischemic postconditioning. The learning and memory abilities of rats were evaluated by Morris water-maze test at 48 h and 72 h after manufacturing model successfully, morphological changes were observed by HE staining, the expressions of Cyclin D1,CDK4 and Notch1 at 6h,24 h,48h,72 h were measured by immunohistochemical staining and Western blotting.Results 1 The result of the Morris water-maze test: Compared with sham group, the escape latency time of rats prolonged, and the number of crossing the platform decreased in global cerebral ischemia group at 48 h, 72h(P<0.05). Compared with global cerebral ischemia group, the escape latency time shortened and the number of crossing the platform increased of the rat in ischemic postconditioning group(P < 0.05). Compared with ischemic postconditioning group, the above changes were further serious in inhibitor group(P < 0.05).2 The result of morphological changes(HE staining): Compared with sham group,the nerve cell survival rate in hippocampus was decreased in global cerebral ischemia group at 6h,24 h,48h,72h(P < 0.05). Compared with global cerebral ischemia group, the nerve cell survival rate in hippocampus was increased in ischemic postconditioning group(P<0.05). Compared with ischemic postconditioning group, the nerve cell survival rate was further decreased in inhibitor group(P < 0.05).3 Results of Cyclin D1, CDK4 by immunohistochemistry and Western blotting: Immunohistochemistry: Cyclin D1, CDK4 peaked at 72 h. Compared with sham group, the numbers of Cyclin D1, CDK4 positive cell were significantly increased in global cerebral ischemia group at 6h,24 h,48h,72h(P<0.05). Compared with global cerebral ischemia group, the numbers of Cyclin D1, CDK4 positive cell were significantly decreased in ischemic postconditioning group(P<0.05).Compared with ischemic postconditioning group,the numbers were increased in inhibitor group(P < 0.05). Western blotting: The proteins of Cyclin D1 and CDK4 peaked at72 h.Compared with sham group, the expression level of Cyclin D1, CDK4 proteins were significantly increased in global cerebral ischemia group at 6h,24 h,48h,72h(P < 0.05).Compared with global cerebral ischemia group, the expression level of Cyclin D1, CDK4 proteins were significantly decreased in ischemic postconditioning group(P < 0.05).Compared with ischemic postconditioning group, the expression levels were increased in inhibitor group(P<0.05). 4 Results of Notch1 by immunohistochemistry and Western blotting: Immunohistochemistry: Compared with sham group, the number of Notch1 positive cell was significantly increased in global cerebral ischemia group at6 h,24h,48 h,72h, and the 48 h reached the peak in the group(P< 0.05). Compared with global cerebral ischemia group, the number of Notch1 positive cell was further increased in ischemic postconditioning group(P<0.05). Compared with ischemic postconditioning group,the number was decreased in inhibitor group(P < 0.05). Western blotting:Compared with sham group, the expression level of Notch1 protein was significantly increased in global cerebral ischemia group at 6h,24 h,48h,72 h, and the 48 h reached the peak in the group(P < 0.05). Compared with global cerebral ischemia group, the expression level of Notch1 protein was further increased in ischemic postconditioning group(P < 0.05). Compared with ischemic postconditioning group,the expression level was decreased in inhibitor group(P<0.05).Conclusions1 Ischemic postconditioning reduced the expressions of Cyclin D1 and CDK4 in hippocampus of global cerebral ischemia rat by which it could regulate the cell cycle to protect neurons; 2 Ischemic postconditioning regulated the cell cycle by promoting the activity of Notch1 pathway through which it could inhibit the expression of Cyclin D1 and CDK4.