The Effect Of Ischemic Postcoditioning To P38MAPK/autophagy Signaling Way In Rats Hippocampus Following Cerebral Ischemia

Objectives 1 To investigate the effect of cerebral ischemic postconditioning on expression of hippocampus neurons autophagy in rats affter cerebral ischemia reperfusion. 2 To investigate the effect of cerebral ischemic postconditioning on activation of P38 MAPK.Methods 1 128 male healthy clean SD rats were randomly divided into four groups:Sham group, global cerebral ischemia group, cerebral ischemic postconditioning group and inhibitor group. 2 using improved pulsinelli four vessels block(4-VO) to establish cerebral ischemia model. Rats of ischemic postconditioning group were given reperfusion15s/ischemia 15 s before restore reperfusion completely, repeat 3 times. Inhibitor group were injected SB203580 into the rats by intraperitoneal 30 min before ischemic postconditioning, after building models every other day until the rats were executed. 3 At48 h, 72 h using water maze record the number of pass the platform and the escape latency time at the points after building the model, so as to evaluate the ability of learning and memory in the process to form conditioned reflex. 4 At 24 h, 48 h, 72 h using HE staining under light microscope to observe variation of neurons form and survival neuron numbers in rats hippocampus. 5 At 24 h, 48 h, 72 h using immunohistochemistry to test the positive cells of autophagy related genes LC3-II, Beclin1, phosphorylated P38 MAPK and using Western Blot method to test the protein level of autophagy related genes LC3-II, Beclin1,phosphorylated P38 MAPK.Results 1 Water maze experiment results: compared with Sham group, global cerebral ischemia group at the points(48h, 72h) measured the numbers of pass platform decreased and the escape latency time prolonged, the difference was statistically significant(P<0.05);Compared with cerebral ischemia group, ischemic postconditioning group at the points(48h, 72h) measured the numbers of passing platform increased and the escape latency time shortened, the difference was statistically significant(P<0.05); Compared with ischemic postconditioning group, inhibitor group at the points(48h, 72h) masured the numbers of passing platform increased and the escape latency time shortened, the difference was statistically significant(P<0.05). 2 HE staining result: compared with the Sham group, cerebral ischemia group visibled the neurons structure was damaged, the numbers of survival neurons decreased, the difference was statistically significant(P<0.05);Compared with the cerebral ischemia group, ischemic postconditioning group visibled the neurons structure damage ameliorated, and the numbers of survival neurons increased(P<0.05); Compared with the ischemic postconditioning group, inhibitor group visibled the neurons structure damage ameliorated further, and the numbers of survival neurons increased(P<0.05); 3 The Immunohistochemistry and Westernblot results of LC3-II and Beclin1: the positive cells presented brown-yellow, located in the cell nucleus. The sham group had low expression in hippocampal neurons of rats. Compared with the Sham group, the positive cells of LC3-II and Beclin1 increased at each time point(6h,24 h,48,72h) in cerebral ischemia group, the difference was statistically significant(P<0.05). Compared with the cerebral ischemia group, the positive cells of LC3-II and Beclin1 increased at each time point(6h,24 h,48,72h) in ischemic postconditioning group(P<0.05); Compared with the ischemic postconditioning group, the expression of LC3-II and Beclin1 increased more at each time point(6h,24 h,48,72h)in inhibitor group, the difference was statistically significant(P<0.05). Compared with the cerebral ischemia group, the protein expression of LC3-II and Beclin1 began to increase after restore reperfusion 6h, 24 h expressed most, 48 h,72h expression gradually decline(P<0.05); Compared with the cerebral ischemia group, the protein expression of LC3-II and Beclin1 increased at each time point(6h,24 h,48,72h) in ischemic postconditioning group(P<0.05); Compared with the ischemic postconditioning group, the protein expression of LC3-II and Beclin1 was more higher at each time pointin inhibitor group(P<0.05). 4 The Immunohistochemistry and Westernblot results of phosphorylated P38MAPK: the positive cells presented brown-yellow, located in the cell cytoplasm and nucleus. The sham group had low expression in hippocampal neurons of rats. Compared with the Sham group, the positive cells phosphorylated P38 MAPK increased at each time point in cerebral ischemia group, the difference was statistically significant(P<0.05); Compared with the cerebral ischemia group, the positive cells of phosphorylated P38 MAPK decreased at each time point(6h,24 h,48,72h) in ischemic postconditioning group(P<0.05); Compared with the ischemic postconditioning group, the positive cells of phosphorylated P38 MAPK decreased more at each time point(6h,24 h,48,72h) in inhibitor group, the difference was statistically significant(P<0.05).Compared with the Sham group, the protein expression of phosphorylated P38 MAPK began to increase after restore reperfusion 6h, 24 h expressed most, 48 h, 72 h expression gradually decline by Westernblot in cerebral ischemia group(P<0.05); Compared with the cerebral ischemia group, the protein expression of phosphorylated P38 MAPK decreased at each time point(6h,24 h,48,72h) in ischemic postconditioning group(P<0.05); Compared with the ischemic postconditioning group, the protein expression of phosphorylated P38 MAPK was more lower at each time point(6h,24 h,48,72h) in inhibitor group, the difference was statistically significant(P<0.05).Conclusions 1 Cerebral ischemic postconditioning promote the expression of autophagy related gene LC3-II and Beclin1 and relieve cerebral ischemia reperfusion injury. 2Cerebral ischemic postconditioning can inhibit P38 MAPK signal pathway, and promote the expression of autophagy related gene LC3-II and Beclin1, give play to endogenous nerve protective effect.