| Background and purpose:The aging of the society makes older diseases an important medical and social problem.Cognitive dysfunction is almost the most prevalent and most common in senile diseases.It is also the most conceiving in cerebrovascular disease.Many studies have shown that cognitive dysfunction is caused by cervical arterial disease.With the improvement of social living standards,as well as the proportion of the elderly population is getting higher and higher,carotid artery atherosclerosis,the gradual formation of plaques,resulting in stenosis or occlusion of blood vessels,the incidence is getting higher and higher,the population healthy The harm is also growing.Therefore,the mechanism of carotid artery stenosis(CAS)leading to vascular cognitive impairment(VCI)and how to prevent and treat it has become a hot research topic for many researchers.Epidemiological survey and more studies have suggested that patients with carotid artery stenosis are often prone to cognitive impairment.And some of the heavier patients,even a high risk of dementia.The formation of atherosclerotic plaques is the most important risk factor for carotid artery stenosis and occlusion.Carotid plaques,especially in the neck,external carotid artery at the bifurcation of the plaque,if caused by arterial stenosis,will directly lead to the occurrence of cerebral ischemic disease.And treatment of plaque-induced stenosis in the treatment of cognitive dysfunction in the brain has a very critical significance.According to the different symptoms of patients,carotid artery stenosis in clinical can be divided into two categories:symptomatic carotid artery stenosis and asymptomatic carotid artery stenosis.Symptomatic carotid stenosis includes:ccarotid artery stenosis with cerebral ischemia,previous transient ischemic attack or ischemic stroke and other medical history.And symptomatic carotid stenosis can lead to vascular cognitive dysfunction is already clinically recognized results,and often cause more serious consequences.Asymptomatic patients with carotid artery stenosis,almost no clinical symptoms and signs of the nervous system.At present,most of the research and investigation,the patients with asymptomatic carotid artery stenosis,whether the complications of vascular cognitive dysfunction less.However,the research institutions and scholars agreed that:there have been severe carotid artery stenosis or occlusion,or the occurrence of plaque ulcers,is caused by vascular cognitive dysfunction risk factors.Therefore,in recent years,researchers have also paid more and more attention to the study of asymptomatic carotid artery stenosis.The current series of studies suggest that cognitive dysfunction if it is caused by carotid artery stenosis,then its mechanism may be:1.Insufficient cerebral blood flow.Carotid artery stenosis can lead to insufficient blood supply to the brain,the amount of input decreased.And chronic blood perfusion caused by cerebral ischemia caused by pathological changes,nerve cell metabolic disorders:the use of glucose cells to reduce the use of various enzymes and protein synthesis and decomposition of the disorder and so on.Resulting in long-term perfusion of cerebral blood flow in the brain frontal lobe,hippocampus cholinergic system dysfunction,leading to reduced neurotransmitter synthesis,choline receptor deficiency,white matter lesions and neuronal damage.And these changes may be the pathophysiology of patients with vascular cognitive impairment.2.Carotid atherosclerotic plaque due to blood flow will break down and collapse.These tiny plaques,which run with the bloodstream,become small emboli,blocking blood vessels from the bloodstream,causing multiple lacunar infarction in the brain.And multiple small lacunar infarction,as an intermediate medium,in the carotid artery stenosis leading to cognitive function damage in the process played a role.Possible reasons are:these emboli caused by the ground stalk injury in the frontal lobe and basal ganglia,internal capsule forelimb/knee fiber connection between,resulting in impaired frontal lobe function,cognitive dysfunction.3.White matter lesions.It has become one of the factors that cause cognitive impairment agreed by organizations and scholars.The number and volume of carotid plaques are positively related to the range and extent of white matter lesions.Brain MRI findings in patients with carotid stenosis associated with cerebral ischemia revealed lesions in the periventricular and deep white matter.The extent and type of cognitive impairment are strongly related to these lesions.4.Plaque occurred in the carotid sinus,especially in the carotid sinus chemoreceptor or baroreceptor location of the plaque,most likely to cause these two receptors dysfunction.Receptors appear in the plaque,may cause the wrong hypoxia and blood pressure reduction signal,the two receptors cause the wrong stimulus.So that the sympathetic nervous system appears a series of morbid regulation:neurological hyperactivity,cerebrovascular regulation dysfunction.And even cause coagulation enhancement,secondary inflammatory response and a series of lesions,leading to patients with VCI.At present,carotid artery stenosis,as well as stenosis caused by cognitive dysfunction in the conventional treatment for surgery to relieve stenosis and conservative treatment of traditional drugs.Surgery and drug treatment may improve the symptoms of these patients.But whether it is surgery,or drug treatment,have their own limitations and risks.The risk of surgery operation itself,and the poor condition of patient's own physical:such as suffering from severe lung disease patients with intolerance;severe coronary heart disease patients with high mortality;surgery may occur nerve injury;postoperative lesions The posterior hyperplasia syndrome and ischemia-reperfusion caused permanent damage to neurological function;the postoperative brain function,the postoperative function of the brain,the posterior segment of the brain,Including the uncertainty of cognitive function improvement mechanism;the improvement of symptoms of patients is not significant;and even social medical security and patient economic conditions and other reasons,have restricted the operation of patients with carotid artery stenosis.The drug treatment,although there is a certain therapeutic effect,but the efficacy of common drugs is not particularly prominent.Especially in the presence of carotid artery stenosis after brain injury,resulting in cognitive dysfunction in patients,whether it is traditional surgery or drug treatment,the effect is unsatisfactory,we need more effective drugs or treatments to solve this medical problems.The study found that the French,like other European and American countries,have high risk of coronary heart disease,such as high-fat food consumption,blood triglyceride levels,high blood pressure and a lot of smoking.But whether the French have a rate of coronary heart disease,or his death rate,than other European countries and regions were significantly lower,and this situation is known as the "French paradox"(French Paradox).And the reason for this phenomenon,after a lot of research and investigation found:the French people like to drink red wine.Further in-depth studies have found that resveratrol,an important ingredient in red wine,plays a major role in reducing the incidence of coronary heart disease.In recent years,people pay more and more attention to the paradox of France,and the function of resveratrol is more clear.Resveratrol is found mainly in nature:grapes,Polygonum cuspidatum,peanuts,red skin,cassia seed and other plants.It has the highest content in fresh purple grape skin.Resveratrol has a very wide range of medicinal value:such as delaying the process of tumor,anti-oxidation,anti atherosclerosis,lipid-lowering and anti platelet,expansion of blood vessels,antibacterial,anti tussive,antiasthmatic effect.In recent years,studies have shown that resveratrol can increase the density of peripheral capillaries in ischemic myocardium,reduce the area of myocardial insufficiency;in myocardial and cerebral ischemia and reperfusion in the experiment,it can reduce the acute injury of heart and brain.Resveratrol can avoid the excessive exposure of collagen fibers under vascular endothelium,thus preventing the injury of vascular endothelial cells.Resveratrol can also reduce the release of procoagulant substances and prevent thrombosis.A number of studies have now found that resveratrol has a protective effect on brain neurons,and studies have also found that resveratrol acts as a therapeutic agent for certain central nervous system diseases.In particular,the protective effect of resveratrol on heart,cerebral vascular and nervous system has recently made resveratrol become the focus of research and hot spots.In vitro experiments have confirmed that resveratrol can reduce neuronal cell damage.Studies have found that resveratrol can protect neuronal cells,to avoid or reduce hypoxia,ischemia and toxins caused by damage.Resveratrol can significantly reduce the cultured hippocampal brain tissue cells in vitro,due to prolonged hypoxia,ischemia caused by injury.The reseratrol of the small trick can significantly reduce the necrosis of the neuronal cells caused by the oxygen-glucose deprivation injury model,which is a very classic ischemic and hypoxic cell model in vitro.The results suggest that resveratrol Brain tissue culture of dopamine neurons in the organization,have a certain protective effect.These in vitro experimental studies can better help us understand the protective effect of resveratrol on various nerve damage mechanisms.In some in vivo experiments,it has been shown that the resveratrol has neuroprotective effects;an experimental study of the animal model of stroke has found that pretreatment of the animal model prior to manufacture with resveratrol;or after the animal model is manufactured The treatment of resveratrol can significantly reduce the volume of cerebral infarction in the experimental animals and reduce the water content of the brain tissue,which indicates that the resveratrol has a neuroprotective effect on the animal model of stroke.In the study of rat animal model experiments,found that different ways of resveratrol administration,can significantly reduce the experimental animal brain damage.In addition,it has been confirmed that albinool can improve the cognitive ability of rat animal model;through the behavioral,histological and cognitive function of different levels of experimental studies,suggesting that resveratrol in vivo,has a protective effect to the nervous system.Not only for ischemic encephalopathy,resveratrol on hemorrhagic cerebrovascular disease,also have the same neuroprotective effect;and for the time of administration study found that:whether adult or neonatal rats in the nerve injury at the same time or Is treated with resveratrol after injury,have a neuroprotective effect;only the longer the time of administration,the lower the degree of neuroprotective effect.The cognitive function of the brain is closely related to the central cholinergic system.Acetylcholine(ACh)is the closest neurotransmitter in the cholinergic system and the cognitive function.In the central cholinergic system,acetylcholine Distribution is very extensive.The study of the causes of memory loss in patients with Alzheimer's disease(AD)has found that the reduction of cholinergic neurons in the brain is a very important causative factor.Choline acetyltransferase(ChAT)in the cytoplasm of neuronal cells catalyzes the synthesis of acetylcholine by choline and acetyl-CoA,and releases it into extracellular vesicles.In the depolarization process,acetylcholine through extracellular secretion to reach the synaptic space,and its receptors(including muscarinic and nicotinic receptors)combined to play a role.A variety of causes of acetylcholine and its receptor levels decreased,leading to central cholinergic system dysfunction,and further lead to cognitive impairment.Therefore,the treatment of cholinergic system has been one of the important goals to improve brain cognitive impairment.In this experiment,we proposed animal experiments,animal models,and the implementation of Morris water maze experiments in experimental animals;observation of the use of resveratrol treatment can improve the cognitive ability of experimental rats,to confirm the white quinoa Whether the alcohol has a neuroprotective effect.(CHAT),acetylcholinesterase(AChE)and vesicular acetylcholine transporter(Vacht)in the hippocampal cholinergic system of carotid stenosis in rats with carotid artery stenosis.And whether the level of expression of these three enzymes in the brain changes,to explore whether resveratrol is through the cholinergic system to play a neuroprotective function,and to make clear its mechanism of action.Method:A total of 30 male SD rats were included in the experiment.These 30 rats were randomly selected and divided into 3 groups,each with 10 rats,as follows:sham operation group,Model group and treatment group.Rats in Sham group were subjected to sham surgery.In the Model group(Model),the rats underwent carotid artery stenosis modeling surgery;and the rats in Tau group underwent carotid artery stenosis modeling surgery and were treated with resveratrol.The rat model of severe carotid artery stenosis established according to a previously described procedure.animals were fixed after chloral hydrate anesthesia and neck midline incision was performed.In the Model and Tau groups,the common carotid artery was dissociated and ligated using#0 surgical suture.In the sham-operated groups the bilateral common carortid artery was exposed without the actual ligation.Antibiotic penicillin was applied to the surgery area to avoid infection.Rats were allowed to access food and water freely after the surgery.After the modeling finished,the rats in Tau group were given 100 mg/Kg resveratrol into the stomach,once a day in a 4-week time frame.As control,equal volume of Stroke-physiological saline solution was injected into the rats in Model and Sham groups.4-weeks later Morris water maze test was performed on each group.The cognitive ability of the rats was evaluated.Use of computer tracking system in the experiment records:? how many times the rat through the platform;? rats in the vicinity of the platform to maintain how much time;? rats reach the platform distanceAfter morris water maze tests,the rats were euthanized by an overdose of intraperitoneal pentobarbital.The brain of each rat was immediately removed,and the hippocampus was isolated.All brain tissues were stored at-80? for biochemical analysis.The mRNA and protein expression of ChAT,AChE and Vacht were examined in the rurrent study.After Morris water maze experiment,rats were anesthetized with pentobarbital sodium and immediately the rats were decapitated,the brain group was isolated from the hippocampus.After weighing,put it in a glass uniform tube,prepare a homogeneous solution,centrifuge and take the supernatant into the Eppendorf tube and place it in a-20? refrigerator.All other brain tissue specimens were stored at 80? for further biochemical analysis.Determination of AChE activity.Principle:The activity of acetylcholinesterase was determined by Ellman's method.The activity of AChE was measured by measuring the rate of increase of absorbance of 5-mercapto-nitrobenzoic acid at 412 nm.Determination of ChAT activity:ACh After ChAT action,one of the substrates is acetyl-CoA,which binds the developer to determine the content by absorbance at 324 nm.The expression of AChE,ChAT and Vacht mRNA in hippocampus of rats was detected by qRT-PCR.Rusults:In the water maze test,the rats in the sham operation group showed a shortened escape latency,indicating that the learning ability of the mice in this group was not impaired.In the control group,escape latency did not show a sustained shortening pattern,it was found much higher than that of the sham operated group.In the treatment group,after treatment with resveratrol,the escape latency continued to decline as training continued.It suggested that resveratrol can restore part of the learning ability.Resveratrol increased the expression levels of AChE,ChAT and Vacht in the hippocampus of rats.In this study,the expression levels of ACNE,ChAT and Vacht in the hippocampus of rats were measured.In the model group,all of these enzyme levels were significantly reduced,indicating that carotid artery stenosis caused by brain damage reduced the production of these enzymes.After treatment with resveratrol,the expression levels of all three enzymes were higher than those in the sham operation group,but were higher than those in the model group.AChE and ChAT activity in the central cholinergic system were measured:AChE activity in the sham operation group was 0.68±0.04U·mg-1pro,and the AChE activity in the model group was reduced to 0.36 ± 0.05 U·mg-1pro.After treatment with resveratrol,the AChE activity in the treatment group increased to 0.49±0.02 U·mg-1pro.The activity of ChAT was similar to that of sham operation group,which was 165.53±1.95U·mg-1pro,which was significantly higher than that of model group 105.74±2.64U·mg-1pro.Resveratrol treatment increased ChAT activity to 136.82 ±2.45U·mg-1pro.The mRNA expression levels of AChE,ChAT and Vacht were detected by RT-PCR.The mRNA levels of all three enzymes in model group were significantly lower than those in sham operation group.The treatment group was treated with resveratrol,the mRNA of these three enzymes increased,higher than the model group.Conclusion:1.Resveratrol shortens the escape latency of rats with cognitive impairment in carotid artery stenosis.The escape latency of the treatment group continued to decline as the training progressed.Compared with the model group,the escape latency of the treatment group was still higher than that of the sham operation group,but the latency was significantly decreased,indicating that resveratrol could restore some learning ability.2.Resveratrol improves cognitive memory in cognitive impairment rats.The model group showed consistent results in three measures,indicating a decrease in cognitive impairment and learning ability under carotid artery stenosis conditions,which reduced the number of platforms across the platform and remained near the platform and increased the reach of the platform Of the escape distance.In the treatment group treated with resveratrol,the rats showed a certain improvement in cognitive learning ability,and the results were between the sham operation group and the model group.3.Resveratrol improves AChE and ChAT activity.In this study,AChE and ChAT key enzyme activities in the central cholinergic system were measured:AChE activity in the sham operation group was 0.68±0.04 U·mg-1 pro,and the AChE activity in the model group was reduced to 0.36±0.05 U·mg-1pro After treatment with resveratrol,the AChE activity in the treatment group increased by 0.49±0.02 U·mg-1pro.The activity of ChAT was similar to that of sham operation group,which was 165.53±1.95U·mg-1pro,which was significantly higher than that of model group 105.74±2.64U·mg-1pro.Resveratrol treatment increased ChAT activity to 136.82±2.45 U·mg-1pro.4.Resveratrol increased the expression levels of AChE,ChAT and Vacht in the hippocampus of rats.The expression levels of AChE,ChAT and Vacht in the hippocampus of rats were measured.In the model group,the expression level of all these enzymes was significantly reduced,indicating that carotid artery stenosis leading to brain damage reduced the production of these enzymes.The expression levels of all three enzymes in the hippocampus of the treatment group were higher than those in the sham operation group,but were higher than those in the model group.These results suggest that resveratrol can affect the expression of these enzymes,and resveratrol is also able to play a role in cognitive function protection.5.Resveratrol up-regulates mRNA expression of AChE,ChAT and Vacht in hippocampus of rats.We also studied the mRNA expression levels of AChE,ChAT and Vacht by RT-PCR.Compared with the sham group,all three mRNA levels in the model group were significantly decreased.The treatment group was treated with resveratrol and the mRNA expression of the three genes was enhanced.Significance:This study shows that the treatment of resveratrol can improve the cognitive impairment of carotid artery stenosis in rats.The therapeutic effect of resveratrol may be to alter the expression of three key enzymes in the rat hippocampal cholinergic system to play a role.These results suggest that the neuroprotective function of resveratrol may be through the cholinergic system and that resveratrol may be a potential drug for the treatment of cognitive dysfunction in patients with carotid artery stenosis. |
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