| BackgroundsIschemic cerebrovascular disease(ICVD)has a high morbidity,high mortality and high disability.The rate of disability caused by cerebrovascular accident is the highest among many diseases in China.Cerebral ischemia is the main cause of cerebrovascular disease.It is difficult to recover after the onset of ischemic cerebrovascular disease.It often leads to impairment of neurological function and obstacles to physical activity,and brings serious pain to patients and their families.Social burden.At present,studies on cerebral ischemia have shown that most of the neuronal cells in the cerebral ischemic area after cerebral ischemia undergo structural destruction,dysfunction,and necrosis with cerebral ischemia.Around the ischemic ischemic area,there are often scattered neurons that are not completely damaged.Neurons have plasticity.The recovery of brain function after brain injury is closely related to the plasticity of neurons.Synapse is a key part of information exchange between nerve cells.The complete synapse structure and function are the physiological basis of neuronal cell repair and regeneration.Synapses also have plasticity,synaptophysin(SYN),microtubule-associated protein-2(MAP-2),and postsynaptic density-95(PSD-95).Participate in the formation of synapses and the information transfer process between synapses.It also plays an important role in the process of synaptic remodeling of damaged neurons,and promotes the enhancement of synaptic remodeling for the repair of neuronal structures and neurons.The restoration of function is of great significance in improving the neurological dysfunction after cerebral ischemia.The MiJian ChangPu is a prescription for the treatment of impaired hemiplegia and impaired memory after the stroke,which has been proved to beeffective in improving the recovery of neurological dysfunction after cerebral ischemia.The mechanism of action is not yet clear.Objectives1.To observe the brain protection and nerve function recovery of MiJian ChangPu prescription in rats with cerebral ischemia reperfusion injury by detecting neurological function scores,the pathological changes in brain tissue.2.To study the neurological function recovery of MiJian ChangPu prescription in rats with cerebral ischemia reperfusion injury by detecting changes in the expression of synaptic remodeling markers SYN,MAP-2 and PSD-95 in hippocampal neurons,and discuss the possible mechanism of the prescription.Method1.SD male rats,3~4 months,250,weight about 300±50 g,were randomly divided into normal group,control group,model group,nimodipine group,Tianzhi granule group,MJCPF low dose group,MJCPF middle dose group and MJCPF high dose group.Among them,10 rats were in normal group,and the remaining groups were divided into 7d,14 d and 28 d groups according to 3 samples,and 10 rats in each group.2.Focal cerebral ischemia reperfusion model in rats inserted with a linear plug to prepare middle cerebral artery occlusion(MCAO)model.2 hours after the operation,the line embolus was pulled out for ischemia reperfusion.3.The normal,sham-operated and model groups were intragastrically administered with distilled water,and the drug-treated groups were intragastrically administered with the corresponding drugs.The amount of drug administered to the rat was calculated according to the equivalent dose conversion relationship according to the crude drug content.The volume of intragastric administration was l0m1/kg,and the dose of injection was calculated according to the equivalent dose of body surface area of rats and humans.Nimodipine and Tianzhi granules were used.20mg/kg/d,low dose of Mi Jian ChangPu prescription was 4.3g/kg/d,middle dose of MiJian ChangPu prescription was8.6g/kg/d,and high dose of Mi Jian ChangPu prescription was 17.2g/kg/d.30 minutes afterintragastric administration,2 times per day after modeling.4.Rats were scored for neurological function at 2h and 48 h after surgery using the modified Longa8 score system.Rats were scored for neurological function 2h before harvest using Garcia18 scoring method.HE(hematoxylin-eosin,HE)staining was used to observe the pathological morphology of neurons in brain tissue;Immunohistochemistry was used to detect the changes of expression of SYN,MAP-2 and PSD-95 after focal cerebral ischemia-reperfusion injury in rats.Results1.Structural changes and neurological recovery of neurons after cerebral ischemia-reperfusion in rats.1.1 Pathological changes in brain tissue: In the sham-operated group,the hippocampal neurons were uniform in cell size,complete in morphology,clear in cell body and cell membrane,and distinct in nucleus and nucleoli.In the model group,the gaps between the hippocampus and cortex tissue in the ischemic area were widened,the endothelial cells were swollen,the central cell structure of the lesions was loose,and the hippocampal neuron processes were broken.The number of degenerate and necrotic neurons was seen,and some nerve cells became smaller and darker in concentration.The nucleolus disappeared,and most cells had degenerative changes.The nerve cell structure of the drug administration group improved to varying degrees.The pathological changes of hippocampal neurons in MJCPF groups were significantly improved at the 14 th and 28 th days at the two points of time compared with the nimodipine and Tianzhi granule groups.1.2 Changes in neurological scores in rats(2h,48h): Compared with the normal and the sham operations,the neurological function scores of the model group at 2h and 48 h after surgery were significantly higher(P<0.01).Compared with the model group,the neurological scores of the rats in the administration group decreased significantly after 48 hours(P<0.05).Compared with the postoperative 2 hours,the neurological scores of the administration group decreased significantly after 48 hours(P<0.05).1.3 Changes in rat neurological scores(7,14,28d): Compared with the normal and the sham operations,the neurological function scores at 7d,14 d,and 28 d in the model group were significantly lower(P<0.01).Especially in 7d group,the rats had the most obvious reduction.Compared with the model group,the scores of neurological function in the rats of the administration group increased(P<0.05).Compared with the nimodipine group,the neurological scores of the Tianzhi granule,MJCPF middle and dose decreased significantly(P<0.05).Compared with the Tianzhi granule group,the neurological scores of the MJCPF high dose group increased significantly(P<0.05).Compared with the MJCPF middle dose group,the neurological score of the high dose group was significantly higher(P<0.05).Compared with 7 and 14 d groups,the neurological function scores were significantly higher in the 14 and 28 d groups(P<0.05,P<0.01)with the prolonged administration time.2.Changes of synaptic remodeling markers in hippocampal neurons after cerebral ischemia-reperfusion injury in rats and effects of MiJian ChangPu prescription on them.2.1 The chandges of SYN expresstion: Compared with the normal and the sham operations group,the number of SYN positive cells in the model group decreased significantly at each time point(P<0.01),and the positive average optical density(OD)values decreased at 7 and 14 days(P<0.01,P<0.05).28 d increased significantly(P<0.01).Compared with the model group,the number of SYN positive cells in the treated group(excluding MJCPF low dose group)increased(P<0.05),and the expression of SYN positive OD in the treated group increased(P<0.01,P<0.05).Compared with nimodipine and Tianzhi granules group,the number of SYN positive cells increased in the MJCPF middle and high dose groups at 14 and 28 days(P<0.05),and the positive OD value increased significantly(P<0.01,P<0.05).Compared with MJCPF middle dose group,the expresstion of SYN decreased in the MJCPF low dase group(P<0.01,P<0.05).Compared with 7 and 14 d groups,the positive expresstion of SYN increased significantly in the 14 and 28 d groups(P<0.05,P<0.01)withthe prolonged administration time.2.2 The chandges of MAP-2 expresstion: Compared with the normal and the sham operations,the number of map-2 positive cells in the model group was significantly decreased at each time point(P<0.01),and the positive OD was significantly reduced at 7d(P<0.01).Compared with the model group,the number of map-2 positive cells in the drug group increased(P<0.01,P<0.05),and the expression of positive OD in nimodipine,MJCPF middle and high dose groups was significantly increased(P<0.01,P<0.05).Compared with nimodipine and Tianzhi granule groups,the number of map-2 positive cells in the MJCPF middle and high dose groups increased significantly(P<0.01,P<0.05),and the expression of positive OD increased(P<0.05).Compared with MJCPF middle dose group,the expresstion of SYN decreased in the MJCPF low dase group(P<0.01,P<0.05).Compared with 7 and 14 d groups,the positive expresstion of MAP-2 increased significantly in the 14 and 28 d groups(P<0.05,P<0.01)with the prolonged administration time.2.3 The chandges of PSD-95 expresstion: Compared with the normal group and the sham operations,the number of PSD-95 positive cells in the model group decreased significantly at each time point(P<0.01),and the positive OD values decreased at 7days(P<0.01).It increased at 28 days(P<0.01).Compared with the model group,the number of PSD-95 positive cells in the nimodipine,MJCPF middle and high dose groups increased significantly at 14 and 28 days(P<0.01).The expression of positive OD value was increased(P<0.01,P<0.05)at each time point.Compared with nimodipine group,the number of PSD-95 positive cells and the positive OD value decreased in the Tianzhi granule and MJCPF low dose groups at 14 and 28 days(P<0.05,P<0.01).The number of PSD-95 positive cells increased significantly in MJCPF middle and high dose groups at 14 and 28days(P<0.01).Compared with Tianzhi granule group,the number of PSD-95 positive cells increased in the MJCPF middle and high dose groups at 14 and 28 days(P<0.05),and the positive OD value increased significantly(P<0.01).Compared with MJCPF middle dosegroup,the number of PSD-95 positive cells in MJCPF high dose group was higher at each time(P<0.05),and the positive OD value was significantly higher at 28d(P<0.05).The number of PSD-95 positive cells in MJCPF low dose group was lower at 14 and 28days(P<0.01),and the positive OD value was significantly lower at each time(P<0.05,P<0.01).Compared with the 7 and 14 d groups,the positive expression of PSD-95 increased significantly in the 14 and 28 d groups(P<0.01)with the prolonged administration time.Conclusion1.MiJian ChangPu prescription has obvious protective effect on nerve cells of cerebral ischemia reperfusion injury in rats,and promotes the recovery of nerve function.The brain protective effect increases with the prolonged administration time,and the effect is obvious on the 14 th and 28 th days after reperfusion.2.The expression of synaptic remodeling markers in hippocampal neurons decreased after cerebral ischemia-reperfusion injury in rats,and the recovery of neurological function was associated with increased expression of SYN,MAP-2 and PSD-95.MiJian ChangPu prescription could increase the expression of SYN and MAP-2,PSD-95 in hippocampal neurons and its mechanism to improve the recovery of neural function in rats may be related to the promotion of synaptic remodeling in hippocampal neurons. |
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