| Background and objectiveAcute cerebral infarction is the most common form of cerebrovascular disease(CVD), accounts for about 70% of CVD. Cerebral ischemia reperfusion is the process of restoring blood. It has an important role which restores blood and improves function of ischemic penumbra, but cerebral ischemia reperfusion also causes certain brain damage.Excessive inflammation immune reaction is one of the important damage mechanisms of cerebral ischemia-reperfusion injury. Microglia (MG) is the main immune cells in the brain which plays an important role in the physiological and pathological process of central nervous system. Microglia can active quickly when internal environment change of brain and produce neural toxic factors, and aggravate brain damage. iNOS is a kind of important enzyme of body when cerebral ischemia reperfusion injury. Cromakalin(ATP-sensitive potassium channel opener) is a special potassium ion channel of voltage independence with cell ATP/ADP level as the door control factors. Many studies have showed that K-ATP channels exist widely in central nervous system. Its activation is the self protection mechanism of anoxic condition and could combat damage to the body. At present, the study whether cromakalin intervenes microglia and iNOS is less and its mechanism is not clear.In this research, to provide theory basis of K-ATP channels opener as applied to clinical treatment of ischemic cerebrovascular diseases, using rat MCAO model, the changes and the protective effects of cromakalin on rats following acute cerebral ischemia-reperfusion were observed. Materials and methods1. Experimental animals and groups:72 health male SD rats were randomly divided into three groups, that were sham-operated group (Group A) ischemia-reperfusion group (Group B),and cromakalin intervention group (Group C).and each group was divided into 4 various stages(6h,24h,72h,7d,according time).2. Animal models preparation:MCAO model was established according to Zea-Longa methods. Nerve behavioral score was established according to Zea-Longa 5 cents. Animals with subarachnoid hemorrhage after craniotomy and death after reperfusion within 24 hours were discarded.3. Animal treatment:Group C was induced by intraperitoneal injection of cromakalin (0.4mg/kg/d) 2ml after MCAO 2h. Group B was induced by intraperitoneal injection of physiological saline 2ml after MCAO 2h. Group A and group B was dealt with the same, besides group A did not insert line bolt. Reaching different time points, rats were induced by intraperitoneal injection of 10% hydration chlorine aldehyde (350mg/kg), then brain coronary paraffin sections (thickness 4μm) were made.4. Observing indexes:the expressions of OX42 and iNOS were examined by immunohistochemical staining. The numbers of neurons were examined by Nissl's staining.5. Statistic analysis:the experimental data was expressed by mean±standard deviation and the date was analyzed with the SPSS 10.0 software, the means of the samples were compared using one-way analysis of variance followed by Least Significant Difference (LSD) for multiple comparison. The significant difference was judged by a=0.05.Results1.Animal neurobehavioral scores:Group A had no obvious neurologic deficits symptoms after waking up. Group B and C had no obvious difference after MCAO ischemia reperfusion 6h (P>0.05). In other time point, group B and C had obvious difference (P<0.05). Group C had significantly reduced than group B (P<0.05).2. The expression of OX42 in CA1 of hippocampus:there were expressions of OX42 in group A,B and C of all time points. Expressions of OX42 in group A did not change significantly in 4 time points.OX42 in group B and C begun to express after ischemia-reperfusion 6h; increased significantly in time point of 24h; reached peak in time point of 72h, and still expressed in time point of 7d. The expressions of OX42 in group B and C were significantly higher than that in group A at each time point (P< 0.05).3. The expression of iNOS in CA1 of hippocampus:there were expressions of iNOS in group A, B and C at all time points. Expressions of iNOS in group A did not change significantly in 4 time points. iNOS in group B and C begun to express after ischemia-reperfusion 6h; increased significantly in time poin of 24ht, reached peak in time point of 72h, and still expressed in time point of 7d. The expression of iNOS in group B and C were significantly higher than that in group A at each time point (P< 0.05), but compared with group B, the expression of iNOS in group C was fewer at all time points (P<0.05).4.The number of neurons in CA1 of hippocampus:there were 60.66±6.62 in group A,11.50±3.08 in group B, and 29.33±4.88 in group C, there was a statistically significant difference among three groups (P<0.05)Conclusion1. Microglia in CA1 of hippocampus can be activated, the expression of iNOS can increase significantly and enhanced as reperfusion prolonged by acute cerebral ischemia-reperfusion injury.2. Cromakalin can reduce microglia activation; the expression of iNOS in CA1 of hippocampus. can reduce the brain injury.3. Cromakalin can protect neurons of CA1 of hippocampus, reduce the death of neurons, alleviate nerve defect symptoms, it has the protective effect on brain. Keywords:cerebral ischemia reperfusion, microglia, cromakalin, brain protection... |
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