| Cerebral ischemia is a cerebral vascular disease with a reduction in cerebral circulation blood flow,characterized by high morbidity,mortality and mutilation rate,was became a global health problem.China’s Ministry of Health issued “China stroke declaration” in 2011,it identified that stroke has become China’s first deadly disease,the morbidity is the highest in the world.The expenses on medical,nursing and others every year have become a heavy burden for the family and society.So,the primary prevention of cerebral ischemia is particularly important,which interventions and nursing interferences are given when there are no strokes or risk factors,reducing the risk of stroke,improving the body’s ability to resist,to avoiding or reducing the severity of stroke.Chronic intermittent hypobaric hypoxia(CIHH)preconditioning has many beneficial effects on the body.It has been used for the prevention and treatment of some diseases for a long time,However,whether it can be used to prevent the occurrence of stroke need further study.In recent years,mitochondria have become a sub cellular target of ischemic injury,and its role in brain protection has been paid more and more attention.It was found that cerebral ischemic preconditioning was related to mitochondrial membrane ATP sensitive potassium channel(mito KATP),mito KATP channel blocker 5-hydroxydeenoate(5-HD)can be used to block cerebral ischemic tolerance,the results suggest that mito KATP may also be involved in the formation of cerebral ischemic tolerance.However,the role of mito KATP in the cerebral ischemic tolerance induced by CIHH is unknown.The purpose of this study was to use functional,morphological and molecular biological methods,from different levels of the whole animal,in vitro tissue and cell,dynamic observing the induction of cerebral ischemic tolerance by CIHH preconditioning,to investigate the role of mito KATP channel in ATP and its possible mechanism.The results of this study will provide an important theoretical basis for the further study of CIHH cerebral protection and prevention and nursing treatment of cerebral ischemic diseases.Objective: To establish the model of cerebral ischemia in rats,and confirm that CIHH has protective effect on cerebral ischemia,and to study the effect of mito KATP in it and its possible mechanism.Methods: Healthy male Wistar rats were randomly divided into the seven groups:(1)control group,without surgery and treatment;(2)Sham group,subjected to permanent bilateral occlusion of vertebral arteries 1 dafter CIHH and exposed to common carotid arteries without occlusion;(3)CIHH group,submitted to CIHH with simulating 5000 m high-altitude,6 h per day,once daily for 28 days;(4)ischemic insult(ISH)group,bilateral occlusion of the common carotid arteries was induced 2 d after the vertebral arteries occlusion;(5)CIHH + ISH group,ischemia was elicited 3 d after CIHH,and the remaining procedures were similar to the ISH group;(6)5-HD + CIHH + ISH group,5-HD(antagonist of mito KATP,40 mg/kg)was given intraperitoneally 30 min before ischemia;(7)5-HD + Sham group,5-HD was given to sham-treated rats.After 30 days of the cerebral ishemia,Morris water maze test(MWM)was used to observe the learning and memory behavior of the experimental animals.After that,neuropathological examination was detected by the investigation of delayed neuronal death(DND)in hippocampal CA1 region.Rats from the control and CIHH groups(six time points of 0 h,6 h,1 d,3 d,7 d and 10 d post-CIHH)were used for examining the dynamic expression of SUR1 and Kir6.2,two subunits of mito KATP,by Western blot analysis.Then the comparations the expression of SUR1 and Kir6.2 in the CA1 hippocampus among groups were examined at 2 d or 7d post-ischemia.analyzed the content of Cytochrome c,changes of ΔΨm and ultrastructure in the CA1 hippocampus.Results:1 In the Morris water maze,no significant change was observed between the Sham and CIHH groups.However,the latency in the ISH group was longer than that of the Sham group(P<0.05).In the CIHH + ISH group,the latency was shorter than that of the ISH group(P<0.05).The latency in the 5-HD + CIHH + ISH group was longer than in the CIHH + ISH group(P<0.05).During the spatial probe test,no significant difference was observed between the Sham and CIHH groups.Rats in the CIHH + ISH group showed a significant preference for searching in the target quadrant compared with the ISH group(P<0.05).In the 5-HD + CIHH + ISH group,the percentage of time in the target quadrant approached that of the ISH group(P<0.05).2 The histological characteristics of the CA1 hippocampus in the CIHH group were similar to those in the Sham group.However,in the ISH group,almost all neurons died.When animals were pretreated with CIHH before ischemia,the above noted lesions were prevented.In the 5-HD + CIHH + ISH group,is similar to ISH group.3 Compared with control group,the protein expression of Kir6.2,SUR1 was significantly higher in 0h,6 h,1d,3d,7d groups(P<0.05),and there was no significant difference between 10 d after CIHH and control group.4 After CIHH treatment 2 d or 7 d,compared with the Sham group,the Kir6.2,SUR1 protein expression of CIHH group increased significantly(P<0.05),in ISH+ CIHH group the protein expression of Kir6.2,SUR1 increased significantly than ISH group(P<0.05).5 Compared with Sham group,the Cytochrome c in the cytoplasm of hippocampus decreased significantly in CIHH group(P<0.05),compared with ISH group,the Cytochrome c in the cytoplasm increased significantly in CIHH+ISH group(P<0.05)5-HD could reverse the above changes,changes in mitochondria and cytoplasm Cytochrome c in the opposite.6 Compared with the Sham group,the ΔΨm levels was no significant changes in CIHH,ISH group decreased significantly(P<0.05),compared with the ISH group,CIHH ISH group could inhibit the decrease in the level of ΔΨm(P<0.05),5-HD can reverse these changes.7 Compared with the Sham group,CIHH group had no obvious change,mitochondrial ultrastructure in ISH group changed significantly and showed extensive vacuolization,cristae with severe membrane damage;while CIHH+ISH can partly improve the structure changes of mitochondria,in 5-HD+CIHH+ISH group and ISH group,the mitochondrial ultrastructure is similar.Conclusion: CIHH pretreatment has protective effect on cerebral ischemia,reduced learning and memory damage caused by ischemia.The mitochondrial membrane ATP sensitive potassium channel can regulate the mitochondrial subunits Kir6.2and SUR1 in hippocampal CA1,the expression of Cytochrome c protein,mitochondrial membrane potential and mitochondrial ultrastructure of mitochondria,which involved in the mechanism of cerebral ischemic tolerance induced by CIHH. |
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