| Spinal cord injury (SCI) may cause by trauma, hemorrhage, tumor compression. As a common neurosurgery disease. SCI can affect the quality of life of patients and their families seriously. In recent years, more and more traffic accidents and natural disasters happen and the severity of spinal cord injury may result in permanent disability. The treatment of spinal cord injury includes surgery, cell transplantation, medication, gene engineering. Hormone shock treatment is widely used in drug treatment, but because of its side effects, using of the hormone is restricted.It is confirmed that early application of large dose of hormone therapy has neurological rehabilitation effect in spinal cord injury. The most widely used drugs for early treatment of spinal cord injury are dexamethasone and methylprednisolone, which are advocated by the vast number of clinical workers. Hormone can relieve cellular edema, improve blood circulation, inhibit inflammation and reduce the release of toxic substances, etc. But large doses of the hormone application may lead to serious complications such as blood glucose, water electrolyte disorders, stress ulcer and so on, so the shock therapy can only be performed for a short time in clinic. Clinical doctors are putting more efforts in looking for other alternative medicine for dexamethasone and methylprednisolone treatment of spinal cord injury (sci). In recent years, it was reported that estrogen and progesterone could improve neural function in rat spinal cord injury, but the mechanism is not clear. Chinese traditional medicine treatment of spinal cord injury has certain curative effects, especially in promoting blood circulation to remove blood stasis drugs, however, some medicines are lack of verification by evidence based medicine, and evaluation of their side effects is not thorough and complete.2-methoxyestradiol (2me2) is the natural metabolites of hormone estradiol, existing in human blood. The present study shows that it has certain anti-tumor and nerve protection effects. Because of the small toxic effects, more and more experts and scholars pay attention to it. In recent years, it is widely recognized that 2ME2 can reduce the nerve cell apoptosis, relieve edema and has other nerve cells therapeutic effects on ischemia anoxic encephalopathy. Spinal cord injuries can also cause nerve cell degeneration and necrosis. Damage after ischemia hypoxia, internal environment change and intravascular subsequent accumulation of harmful substances can cause spinal cord damage. There are few reports about whether 2me2 has certain therapeutic effects on spinal cord injury. This experiment intends to test whether 2me2 has therapeutic effects on acute spinal cord injury and explore its possible mechanism.Tau protein is a protein associated with microtubule, whose main function is to combine with tubulin, maintain the stability of microtubules and induce the formulation of microtubules. It induces and promotes the polymerization of tubulin into microtubules, combines with newly polymerized microtubules, keeps the distance between microtubules, influences the attachment point of protein kinase of neurons and plays an important role in neuronal plasticity. Currently, the level of Tau protein among neurons in AD patients is evidently more than normal, but with the opposite result in functional ones. Recent study has found that hypoxic ischemic encephalopathy has a close relationship with the level of tau protein and phosphorylation in nerve cells, as increased tau protein, which is accompanied by excessive phosphorylation, may cause destruction in microtubule structure of neurons and damage of normal axonal transport, thus resulting in impairment of neuronal function. Also, there is damage in microtubules in the process of spinal cord injury, but only few reaches regarding its relationship with tau protein and phosphorylation level have been conducted, therefore, we hope to discuss the relationship between spinal cord injury and tau protein by testing the level of tau protein and phosphorylation after spinal cord injury.Hung KS found that tau protein phosphorylation level of rats increased following spinal cord injury but was dropped after treatment, and raised that there is positive correlation between tau protein phosphorylation level and spinal cord injury. In our experiment, we found that 2me2 has therapeutic effect on acute spinal cord injury, and can improve the motor function of spinal cord injury in rats and reduce the nerve cell apoptosis; Therefore, we hope to find the possible mechanism for the treatment of spinal cord injury through testing the change of tau protein phosphorylation level before and after 2me2 treatment.Acute cerebral infarction, with high incidence and mortality rates in China, is one of serious health-threatening disease types. Acute cerebral infarction paroxysm will lead to obstruction of cerebral artery and impairment of brain blood supply, resulting in necrosis or softening in local brain tissues due to hydroxy-ischemia. However, functionally reversible damage may be presented in the tissues on reversible ischemia penumbra around the infarction lesions. This part of tissue cells still has normal organization, but with flaws in the physiological function. And they may degenerate and die in the unfavorable circumstances but restore to their normal function under the good environment. The current clinical studies were focused on the protection of nerve protection treatment in the peripheral areas of the infarction. It’s reported that 2me2 plays a protective role in cerebral ischemia hypoxia, including inhibition of nerve cell apoptosis, reduction of edema and other nerve cells damage and certain protective effect upon defective anoxic encephalopathy. This experiment studies the effect of 2me2 on acute cerebral infarction of rats to provide certain the theoretical basis for clinical treatment of acute cerebral infarction.When the cute cerebral infarction occurs, nerve cells are in a state of apoptosis due to ischemia hypoxia, edema compression after infarction, free radicals accumulation, cell metabolic disorders and other complex factors. Neurons apoptosis is the process of programmed cell death under gene control. The genes that control cell apoptosis can be divided into promotive and inhibitory apoptotic genes. The activity of these two genes determines the apoptosis of nerve cells. Caspase 3, the BCL-2 family and Fas belong to promotive apoptotic genes, among which Caspase3 plays an important role in the progress of nerve cell apoptosis. When studying the model of acute cerebral infarction of rats, Adam Hafeezet and other experts founded that ischemia hypoxia can induce the apoptosis of nerve cells and cause neuronal loss. And the expression of caspase-3 is detected to be increased in peripheral tissues of cerebral infarction of rats. Hence, in experimenting we verify the effect of 2me2 on the apoptosis of acute cerebral infarction through the detection of apoptosis and caspase-3 via TUNEL dye.Chapter I The effects of 2-methoxyestradiol on the neural function and apoptosis of rats with spinal cord injuryObjective:To investigate the effects of 2-methoxyestradiol on the function and apoptosis of rats with spinal cord injury.Methods:A total of 72 adult male Sprague-Dawley rats were randomly divided into 3 groups:sham group (n=24), spinal cord injury group (SCI,n=24) and spinal cord injury+2ME2 treated group (SCI+2ME2, n=24),and the rats of SCI group and SCI+2ME2 group were sacrificed after perfusion. Models of SCI were created by modified Allen’s method,and Basso, Beatti, Bresnahan(BBB) score after modeling at 1,3,7,14,21,28 days,the expression changes of caspase-3 protein after SCI were observed by immunofluorescence staining, and comparative analyses were also carried.Results:BBB score after operation showed that in SCI group and SCI+2ME2 group the scores were lower obviously compared with sham group(P<0.05). BBB score in SCI+2ME2 group were higher obviously compared with SCI group(P<0.05)on the14,21,28days(P<0.05).2ME2 can reduce the expression of caspase-3 and reduce the nerve cell apoptosis.Conclusions:2ME2 offers protection in rats with spinal cord injury and it may inhibit the expression of caspase-3, and finally reduce apoptosis.Chapter Ⅱ The effect of 2ME2 on the phosphorylation of Tau protein after spinal cord injury in ratsObjective:To investigate the effects of 2ME2 on the phosphorylation of Tau protein of rats with spinal cord injury.Methods:A total of 72 adult male Sprague-Dawley rats were randomly divided into 3 groups:sham group (n=24), spinal cord injury group (SCI,n=24) and spinal cord injury+2ME2 treated group (SCI+2ME2, n=24) Rats in treatment group were treated with 2ME2.The rats of SCI group and SCI+2ME2 group were sacrificed after perfusion. The expression changes of p-Tau (Ser 262) protein after SCI were observed by immunofluorescence method and Western blot, and comparative analyses were also carried out.Results.-The p-Tau (Ser 262) protein expression of SCI+2ME2 group were lower obviously compared with SCI group(P<0.05).Conclusions:2ME2 offers protection in rats with spinal cord injury and it maybe reduce the lever of tau phosphorylation.Chapter III Neuroprotective effects of 2ME2 in rats with acuteObjective:To investigate the neuroprotective effects of 2ME2 in rats with acute cerebral infarction.Methods:72 rats were randomly divided into sham group (n=24), model group (n=24) and treatment group (n=24). Rats in treatment group were treated with 2ME2. After middle cerebral artery infarction model were constructed at 7d, rats were sacrificed, caspase-3 protein of rat brain were analyzed and apoptosis index, infarct size and neural function defect of brain cell were analyzed.Results:Caspase-3 protein levels in model group significantly increased than that of sham group (P<0.05), while Caspase-3 protein levels in treatment group were significantly lower than that of model group (P<0.05) and were significantly higher than that of sham group (P<0.05). Apoptosis index of brain tissues in model group and treatment group were significantly enhanced than that of sham group (P<0.05). Apoptosis index range of brain tissues in treatment group were significantly decreased than that of model group (P<0.05).Conclusion:2ME2 can reduce the level of apoptosis in rat brain nerve cells and have a protective effect on nerve.It provides a frame of reference for the clinical treatment of acute cerebral infarction. |
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