| AimStroke is the common disease among elder people, which severely impact human being's health condition, and is fatal to people in the above the middle age. Recently with the increased senior population and the diagnosis of high blood pressure, the prevalence of cerebrovascular disease has been increased. The incidence of ischemic disease is three times more than cerebral hemorrhage, bringing in financial burden to the society and the family, therefore emphasizing treatment and rehabilitation is urgent. Traditional Chinese medicine has been widely used in treating cerevrovascular disease, particularly in effectively regaining functions of impaired nerves, especially, xuesaitong is main consists of PNS, which has been very helpful in the clinical practice. This project is to utlize PNS, through the observation of the group of animals, immune tissue chemical technology and real-time fluorescence PCR, to explore different timing in 7 days, 14 days,28 days regarding different models of focal ischemic in mousse. Through the expressing of cerebral cortexinhibitor Nogo-A, to reveal that traditional Chinese medicine has the effect in re-constructing the mechanism of the brain; this project also further explore the interactions between the dosage and the effectiveness of xuesaitong, in order to further application of traditional Chinese medicine and provide scientific evidence.MethodsNormal male SD mice, weight 300±20g, randomly selected to be experimental group and the non-experiemntal group (control group). The experimental will use improved Longa method to re-construct MCAO, maintain the temperature. After mice waking up for 2 hours, implementing EZLonga evaluation until after the 72 hours, based on the evaluation and weight, randomly assigned into xuesaitong big dosage, and xuesaitong small dosage, nimodipine group,model group, they are administrated on the same day as the operation.After the operation randomly select 3 group models to take TTC dyeing coloring, to evaluate if the model is successful. Selecting 6-8 animals on the 7th, 14th, and 28th day from each group, bring out the brain,at optic decussation of brain cut it into two parts,put the front part into 10% formalin,after 48h,wash the front part with PBS. lay tissiue embedded in paraffin,slice thickness of about 4um, dyeing color with HE,to observe morphologicalchanges of nerve cells dyeing color with immunohistochemistry to obseve different time points expression of nogo-a; in addition the back bring out infarction brain tissue weigh with Electronic Scale about 120mg, rapidly into liquid nitrogen using real time quantitative pcr method to observe the nogo-a mRNA expression change at different time points.Results1 preparation MCAO model and the evaluation of the function of nerves1-3points was included in the standard points, animals are selected for TTC dyeing, infarction area become gray compared to the normal areas which is red, indicating the model is successful.5 hours after the operation, there is no statistically significant finding, showing that animals in each group with the different initial stage status,24 hours,48 hours,72 hours after the operation, according to the nerve function evaluation, with the time going by, the points decreased,24h,48h after operation nimodipine,xuesaitong big dose-xuesaitong small dose are all below the model group, without any statistically significance,there is statistically significant finding in 72 hours after surgery xuesaitong big dose,xuesaitong small dose nimodipine and model group.2 observation of the overall animal state2.1 observation of general statusAfter 24 hours, in all groups animals present less activity, fluffy hair, vertical, unresponsive, huddled, limited drinking and diet; 7 days see improvement in each group, which xuesaitong group and the nimodipine group recover faster, apparently better than the model group. After 14 days,28 days, animals gradually recovered, animals in each group are close to the normal condition, daily activity, mental health, eating normally, and bowels regular urine normal.2.2 observation of the weightControl group animals showed positive growth.other operationg group within 7 days, showed negative growth. Weight loss in xuesaitong group and nimodipine group was smaller than other group. After 28 days, weight gaining found in nimodipine group and model group, p<0.01, statistically significant finding was found in weight gaining in xuesaitong large group and model group, p<0.05.2.3 observation in animal mortalityNormal mortality rate of zero, experimental group, in short, within 24 hours, mortality rate is 0.48 hours, the highest rate of mortality happens; 7 days is the 2nd highest rate of mortality,14 days and 28 days mortality rate remain the same, there was no animal dead basically. Compared to three different timing, model group has the highest mortality rate, nimodipine has the lowest, and xuesaitong with bigger or smaller dosage both lower than the control group.3 xuesaitong MCAO rats on pathological examination of brain tissue infarctionNormal neurons, nucleolus clearly seen, nerve cells arranged in clear rows, the model group infarction area presents large, loose reticulate structure, neuron swelling, increased volume, with a fan shape or polygonal, neurons cytoskeleton disappeared, the cytoplasm stain lighter, condensed nuclei, deep color, blood vessels visible, inflammatory cell infiltration. Xuesaitong group compared to the control group, showing small, light impairment, with more residual cells, better cell status, with prolonged time, pathological of each drugis better the same time model group.4 effect of xuesaitong on the Nogo-A expression at different recovery time pointsAverage optical density analysis of infarction area, borderline area, the normal region showed that infarct zone with the time prolonged, Nogo-A expression gradually decreased, within each time point, the group of xuesaitong and nimodipine both higher than the model group. In the infarct borderline area, In the 7th day, the model group expressed the high Nogo-A trend, and reached the high rate in 14th day, in the 28th day, although slightly lower than the 14th day, still maintains at a high level. At the same time, xuesaitong group presents a significant different between the bigger dosage group and the control group P <0.05, in the 7th day, 14th day,28th day, significant differences are found in xuesaitong big dose and model group significant p<0.05. the opposite side of Infarction area in each treated group expressed a lower Nogo-A score than the control group, and there is no statistically significant differences between groupsConclusion1 xuesaitong can improve the overall condition of animals, reduce the neurological deficit.2 xuesaitong can change and protect nerve cells in infarction area and its peripheral reas3 xuesaitong can reduce the expression of infarction peripheral area Nogo-A at different time.4xuesaitong is more effective with larger dosage, representing a good dose-effect relationship5 xuesaitong can reduce the expression of Nogo-AmRNA at different time6 xuesaitong may protect the brain and reconstruct brain function by reducing the expression of the Nogo-A... |
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