Construction Of Animal Model Of Ischemic Collateral Movement Syndrome

BackgroundIn recent years,with the continuous development and progress of interventional diagnosis and treatment techniques,it has been widely used in the diagnosis of coronary heart disease.However,relevant studies have shown that nearly 40%of patients who undergo coronary angiography due to chest pain and discomfort do not see significant stenosis of blood vessels,which is considered to be related to coronary microcirculation disorders.Therefore,coronary microcirculation disorders have received more and more attention.The mechanisms affecting the occurrence of coronary microcirculation disorders include increased myocardial capillary resistance,collateral blood flow of coronary microcirculation,coronary recanalization,microthrombosis,etc.,in which coronary recanalization(including thrombolysis and PCI process)caused by in situ thrombosis or microemboli fall off and some patients with acute coronary syndrome or stable angina pectoris the focus of attention.Currently,the evaluation methods and treatment methods for CMD are still limited,and we need to further explore the clinical.Microcirculation disturbance caused by microthrombosis can be the core of the research.In previous reports,there are many methods for modeling with physical injury and chemical stimulation as the main means,but both have their own advantages and disadvantages.At present,there is no standardized experimental modeling method that can be widely used.Therefore,it is necessary to create and optimize a new type of animal model to provide better research basis for clinical research.The clinical manifestations of coronary microcirculatory disturbance are similar to those of the pathogenesis of "ischemic collateral wind internal motion" proposed by Professor Wang Xiang's team based on the theory of collateral wind internal motion.Therefore,we will start from the direction of qi deficiency and blood stasis to simulate its occurrence and development process,explore a reasonable and objective new composite model of coronary microembolism,and improve the establishment of collateral wind internal motion syndrome model.ObjectiveAccording to modern science and technology,to construct the animal model of ischemic collateral wind internal motion syndrome which is suitable for clinical,reasonable,feasible and widely applicable.MethodsHealthy SD male rats weighing 260 ± 20 g were randomly divided into sham-operated group(sham group)and model group(CME group),with 30 rats in the former group and 40 rats in the latter group.According to different observation time points,the sham group and CME group were subdivided into 5 subgroups(1 h after operation,3 h after operation,1 d after operation,3d after operation and 1w after operation),with 8 rats in each group.Before the operation,7-8 ml/rat of blood was collected from the abdominal main vein of 2 normal rats,allowed to stand and placed in an incubator at 37 ? overnight to form a blood clot,which was ground into a powder using a glass grinder and sieved through a 38 um sieve plate to make the size uniform and made into allogeneic thrombus particles for future use.0.2 ml pure water was injected into the aortic root of the sham-operated group after thoracotomy,and 0.2 ml of the mixture of microthrombus(2.5mg)and sodium laurate(0.75 mg/ml)was injected into the model group.Pathological examination and hematological examination were performed at corresponding time points after operation.HE pathological section staining method was used to observe the formation of myocardial microthrombosis and histopathological manifestation of myocardial microinfarction.ELISA method was used to detect the levels of vWF and cTnI in rat plasma.Results1.Experimental conditions and methods To explore the best particle preparation for allogeneic thrombus baked overnight at 37?,injection method is the thrombus microparticles and sodium laurate mixed and injected at the same time,the best mixing ratio is 2.5 mg thrombus microparticles + 0.75 mg/ml sodium laurate 0.2ml,injection site to close to the aortic root the nearest convenient operation,vertical needle about 2 mm.2.General Conditions of Experimental Rats There was no significant difference in the starting and ending body weights of experimental animals in each group,with an overall mortality of 20%and a molding rate of 80%.3.Histopathological examination:(1)The model group showed mainly microthrombosis at 1 hour after operation and 3 hours after operation,which was higher than that at 3 hours after operation.There was statistical difference between the two groups(p<0.05).No obvious thrombosis was observed in the remaining time period,and no microthrombosis was observed in the sham-operated group;(2)There was vascular endothelial injury and inflammatory cell infiltration at 3 hours after operation,and the corresponding performance was also observed in each time period;(3)The model group showed mainly multiple and focal microinfarction at 3 days after operation and 1 week after operation,in which the percentage of microinfarction in the total myocardial area in the visual field was greater at 3 days after operation than that at 1 week after operation,and there was no statistical difference between the two groups(p>0.05).No obvious infarction was observed at other time points and in the sham-operated group.4.Hematological examination:(1)Compared with the sham-operated group,the cTnI concentration in the model group at each time point was higher than that in the sham-operated group at the corresponding time point,with significant changes at 1 hour after operation and 1 day after operation,with statistical significance(p<0.05);compared with the model group at 1 hour after operation,the vWf concentration in the model group was significantly lower than that at 1 hour after operation,with statistical significance(p<0.01);compared with the sham-operated group,the vWf concentration in the model group was lower than that in the sham-operated group from 1 hour after operation to 3 days after operation,with statistical significance(p<0.01);compared with the sham-operated group,the vWf level in the model group was significantly lower at 1 hour after operation and 3 days after operation,with statistical significance(p>0.05).5.Evaluation of TCM syndromes(1)Deficiency of collaterals:The model simulates disharmony of qi and blood and loss of collaterals by damaging the endothelium.After the operation,the rats show slow activity,mental retardation,frequent squinting,pale fur and other symptoms of auricle,claw nail and tongue are all manifestations of deficiency of collaterals;(2)Blood stasis:the allogeneic thrombus particles can highly simulate the pathological products of "blood stasis",microthrombosis at 1 hour and 3 hours after the operation,multiple focal microinfarcts can be observed by HE staining of pathological sections at 3 days after the operation and at 1 week after the operation.The symptoms of dark red tongue accompanied by blood stasis in the model group are also manifestations of blood stasis;(3)Signs of wind movement:Intraoperative thoracotomy in rats resulted in rapid onset of disease and varied disease conditions,which could be fully reflected by the rapid formation of thrombus in small blood vessels,acute ST-T changes in ECG,elevated plasma cTnl,rapid reduction of vWf and other laboratory related examinations.Conclusion1.By injecting the mixture of allogenic thrombus microparticles and sodium laurate into the aortic root,the animal model of ischemic collateral wind internal motion syndrome can be preliminarily established;2.The optimal ratio of thrombus microparticles and sodium laurate solution is:2.5 mg thrombus microparticles dissolved in 0.75 mg/ml sodium laurate solution 0.2ml.