| Objective: To investigate the optimal time window of ischemic postconditioning (IP) against focal cerebral ischemia-reperfusion injury.Methods: To confirm the optimal time window of IP, 80 male SD (weighted 240~280g)were randomly assigned to 5 groups ( n = 16 each) : sham -operated group,control group and IP groups with different time intervals (15s,30s,1min). IP was performed by three cycles of reperfusion/ischemia (R/I). The neurological deficit scores (NDS) were evaluated 24h after reperfusion.Morris water maze was applied to 20 male SD(4 rats per group)to analyze the changes in learning and memory abilities. Infarct sizes were measured to 20 male SD(4 rats per group),was determined by 2,3,5-triPhenyltetrazolium(TTC).The others apoptosis of brain cells and expression levels of TNF-α,APP were evaluated by TUNEL and immune stained method respectively in hippocampal CA1 area.Results:1,NDS:There was no neurological deficit in the sham group, control and IP groups showed different degrees of neurological dysfunction. NDS in each of IP groups compared with control groups were decreased significantly(P <0.05), and NDS in IP = 15s,30s group were less than IP = 1min group(P <0.05).2,Ischemic postconditioning could improve behavioral deficits:Rats in the control group required more time to find the platform than those in the sham group and the IP groups. Rats in IP=15s,30s group required less time than IP=1min group(P<0.05). Analysis of escape latency revealed significant differences between groups, but there were no significant difference in the swimming speed among these groups. In the probe test, rats in the control and IP=1min groups spent significantly less time than the other groups in the quadrant where the platform had been, and there was significant difference between the IP=15s,30s group and the control group (P<0.05).3,IP reduced infarct size: Sham group without cerebral infarction , control group and IP groups can be observed cerebral infarction. IP groups significantly reduced infarction compared with control group (P<0.05). In IP = 15s,30s group infarct size is significantly smaller compared to infarct of IP = 1min group(P <0.05).4,HE staining: Sham group showed normal brain structure,in the sham group, cells displayed round, pale, stained nuclei. The control group showed a larger infarct, displayed the normal structure disappeared, disorder, the number of cells reduced, nuclear condensation.IP = 15s, IP = 30s group normal structure disappeared, had less pyknotic nucle, dark stained. IP = 1min group showed infarction (less than the control group), the normal structure can be seen, the number of cells reduced, showed pyknotic nuclei.5,Neuronal apoptosis: Sham group occasionally apoptotic cells, the number of apoptoic cells in hippocampal CA1 area increased obviously of control groups, compared with sham group and IP groups. The number of surviving neurons increased significantly in the I P groups as compared to the control group (P < 0.05).The level of apoptotic nerve cells in hippocampal CA1 area decreased obviously in IP=15s,30s group(P<0.05).6,IHC:(1) TNF-αThe sham group seldom expressed. The expression of TNF-αin hippocampal area CA1 increased obviously of control group. Control group versus sham group (P<0.01).IP groups compared with control group, showing that the level of TNF-αin hippocampal CA1 area decreased significantly. Furthermore, the level of TNF-αin hippocampal CA1 area was significantly increased in IP = 1min group, compared with the IP=15s,30s group (P<0.05).(2) APP The sham group seldom expressed, the amount of APP in both the control group and the IP groups increased significantly as compared with that of the sham group in hippocampal CA1 area (P<0.01).We observed that MOD of APP increased significantly in IP=1min group compared with IP=15s,30s group after reperfusion. The optimal time window of IP to induce reperfusion injury in brain is three cycles of IP=15s,30s group. |
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