| bjective: Intracerebral hemorrhage is a kind of acute cerebrovascular disease,neurology multiple diseases,and because of its high mortality and morbidity,which cause serious damage to the patient’s quality of life and health.The hematoma after intracerebral hemorrhage placeholder effect,hematoma stimulates the release of the surrounding tissue of vascular active substances and hematoma breakdown products,thrombin cascade amplification reaction and the activation of the complement system appears,these factors interact directly or indirectly caused by or in the brain tissue damage,induce inflammatory reaction and apoptosis,cause nerve function defect.In view of the situation of high morbidity and mortality,the treatment of brain hemorrhage in our country at present,there is no breakthrough.Current treatments focused primarily on intracraninal pressure,removal of oxygen free radicals,trophic nerve,control blood pressure,blood sugar,maintaining water and salt electrolyte balance or surgical treatment,however there is lack of specific and effective treatmens.Neural synaptic plasticity is closely related to neural functional recovery after ICH,a longer period of time can be adjust,is advantageous to the damaged neural function recovery.Erythropoietin(EPO)is a kind of secreted by the kidney relative molecular weight of 34000 glycoprotein,ever think that erythropoietin(EPO)in the body surface of redblood cell precursors and is the main purpose of the specificity of erythropoietin receptor(EPOR),thus stimulating the value-added differentiation,in its clinical application is limited to the correction of chronic anemia.In recent years a large number of studies have shown that the brain of erythropoietin(EPO),and the expression of erythropoietin receptor(EPOR),also found that erythropoietin(EPO)generation is closely related to brain tissue oxygen and blood supply situation,when cerebral ischemic anoxia erythropoietin(EPO)generation multiplied,EPO play an important role in protecting neurons,and by reducing the sensitivity of the neurons of ischemia hypoxia,enhance the capacity of the survival of the neurons,to promote the increment of neural progenitor cells and inhibiting apoptosis.But the research about that even if erythropoietin(EPO)can possibly help synapse remodeling is few,so the research on erythropoietin(EPO)on the influence of synapse after cerebral hemorrhage is necessary.This experiment by using recombinant human erythropoietin(rh EPO)after the intervention to observe synaptic around the hematoma after ICH,Synaptophysin(SYP)and postsynaptic dense-95(Post synaptic density,PSD-95)protein changes in the situation,to explore erythropoietin(EPO)on experimental cerebral hemorrhage rat neural synaptic plasticity effect.Methods :1.In front of the experiment according to Morris water maze prepare rat video tracking analysis system for training,testing,4days after continuous training to choose escape latency in rats between 10 and40 seconds into the building;2.using stereotactic apparatus,Autologous bloodwas injected into the caudate nucleus(fontanelle before the 0.2 mm,near the center line on the right side 3.0 mm)of rat that produced ICH model.after two hours animals awake use Zea-longa score grading standard score to slect score between 1 to 3 points of SD rats as model rats.3.Grouping and processing:By Morris water maze of 105 healthy male SD rats were randomly divided into three groups: control group(Sham group),cerebral hemorrhage group(ICH group),cerebral hemorrhage+erythropoietin treatment group(EPO group);then they are according to the postoperative time points are divided into 6h,24 h,48h,72 h,7d and 14 d and 21 d seven groups,each group of five rats.Experimental animals for each category in the following treatment(1)Sham group With micro syringe in basal ganglia region on the right caudate nucleus in locating and slowly Pierce,not injection of autologous blood,the remaining steps in line with cerebral hemorrhage group.(2)ICH group With micro syringe in basal ganglia region on the right caudate nucleus in injection not anticoagulation 50μl autologous blood,at the same time,abdominal cavity injection with EPO group the same amount of saline solution.(3)EPO group With micro syringe in basal ganglia region on the right caudate nucleus in injection not anticoagulation 50 μl autologous blood,at the same time,according to3000U/kg dose to the abdominal cavity injection of EPO.4.Specimen extraction and detection Will building successful rats in postoperative 6h,24 h,48h,72 h,7d,14 d and 21 d the corresponding time point with Garcia first nerve function score method to evaluate nerve function defect,then finish the Morriswater maze test,after chloral hydrate anesthesia,after anesthesia immediately beheaded brain,brain tissue specimens,with dry wet weight method to test the hematoma moisture content changes around brain tissue,with HE staining observation of brain tissue around hematoma basic morphology and structure;using immunohistochemical method to detect the time SYP and PSD-95 protein content around the hematoma in the five groups at different times around hematoma changed by Immunohistochemical and analyzed its average integral optical density.Results:1.The brain water content: at 6h,24 h,48h,72 h,7d point time the hematoma moisture around brain tissue in ICH group and EPO group were significantly higher than Sham group(P<0.05);at6h point time the brain water content surrounding hematoma between EPO group with ICH group has no difference(P>0.05),but at 24 h,48h,72 h,7d point time brain tissue water content around the hematoma in EPO group decreased significantly compares with ICH group(P<0.05).2.Each groups HE staining around the hematoma showed:(1)the Sham group,without brain hematoma,brain tissue structure at each time point is complete;(2)ICH group72 h brain tissue around hematoma is a large number of blood cells,brain structure disorder,a large number of inflammatory cell infiltration,edema,severe nerve cells degeneration necrosis,even when the 7d,14 d visible blood cells around the hematoma was gradually reduced,compared with 72 h the number of inflammatory cells gradually reduce,continuously reduce edema degree and the nerve cells,to 21 d around the hematoma blood cells less evendisappear almost disappeared,inflammatory cells,a large number of fibrous tissue and glial cell hyperplasia,organization structure restoration of damaged.EPO group,each point in time the degree of inflammatory cell infiltration,edema and nerve cell degeneration necrosis is mitigated,damaged tissue repair is better than ICH grounp.3.The nerve dysfunction score:The experimental group rats in postoperative 6h,24 h,48h,72 h,7d,14 d and 21 d each time point to nerve function score before death,(1)Sham group without neurologic deficits(P>0.05);while ICH group,EPO group has a different degree of neurologic deficits and lower than Sham group with statistically significant(P<0.05).there is also 14 d of the score was lower than those of 21d(P<0.05);(2)EPO group at each time point nerve function score were higher than in ICH group,with statistical difference(P<0.05).4.Water maze test results:Before cerebral hemorrhage doing Morris water maze training four days in a row,4times a day,5 days test escape latency,cross platform,platform quadrant time percentage percentage and platform distance,between groups had no significant difference(P>0.05);After cerebral hemorrhage doing water maze test again,recording the above indexes,Sham group were no significant change with earlier in each index(P>0.05),in ICH group and EPO group the escape latency,cross platform,platform quadrant time percentage and platform distance percentage these many kinds of learning and memory ability of indicators,were all decreased with previous,and longer duration of ICH each index rising gradually,and the rising degree and the speed of EPO group is superior to ICHgroup(P<0.05).5.The brain tissue around hematoma SYP protein content:(1)Sham group,only a small amount of SYP protein expression were positive at each point time,there were no significant difference(P>0.05).(2)at 6h,24 h,48h,72 h point,EPO group only a small amount of SYP protein expression were positive,compared with the Sham group no significant difference(P>0.05);7d,14 d and 21 d point ICH group,EPO group SYP positive protein expression significantly more than the Sham group(P<0.05);(3)7d,14 d and 21 d point,SYP positive protein expression EPO group was obviously higher than that of ICH group,with statistical difference(P<0.05);(4)ICH group,EPO group SYP positive protein began to increase after 72 h,14 d to spike(P<0.05),and then express decreased,but there were still more expression when 21 d around the hematoma.6.The immunohistochemical display PSD-95 protein around hematoma(1)Sham group,only a small amount of PSD-95 protein expression were positive in each point time there were no significant difference(P>0.05).(2)at 6h,24 h,48h,72 h point,EPO group only a small amount of PSD-95 protein expression were positive,compared with the Sham group no significant difference(P>0.05);7 d,14 d and 21 d point ICH group,EPO group PSD-95 positive protein expression significantly more than the Sham group(P<0.05);(3)7d,14 d and 21 d point,EPO group PSD-95 positive protein expression was obviously higher than that of ICH group,with statistical difference(P<0.05).(4)ICH group,EPO group since 72 h after the PSD-95 positive protein increased,14 d to spike(P<0.05),and then express decreased,but there were still more expression when 21 d around the hematoma.7.Linear correlation about SYP and PSD-95 protein expression and nerve dysfunction analysis :(1)at 14 d point,ICH group,EPO group of SYP protein content and the corresponding experimental rats nerve function score was positively related to(r=0.940,P<0.001);(2)at 14 d point,ICH group,EPO group of PSD-95 protein content and the corresponding experimental rats nerve function score was positively related to(r=0.848,P<0.848).Conclusion: 1.Using no anticoagulant autologous blood into the legal system should be taken as hemorrhage rats model method is simple,easy to operate,and is similar to the pathologic changes of the human body spontaneous cerebral hemorrhage clinical process,which is the study of the ideal animal model of cerebral hemorrhage;2.synaptic plasticity protein SYP,PSD-95 content increasing in the peripheral nerve around the hematoma after intracerebral hemorrhage,conducive to neural functional recovery;3.Erythropoietin(EPO)can increase SYP,PSD-95 protein expression around the hematoma promote synapse remodeling after ICH;4.Erythropoietin(EPO)can promote the ICH rats the recovery of limb function and the ability of learning and memory. |
PDF Full Text Request