| Background and objectiveStroke is a common refractory disease, which serious harm human health and life safety,According to incomplete statistics, Chinese patients with stroke up to 2 million one year, and incidence rate up to 120/100 thousand, 7 million of survivors are now living. 4.5 million of these patients had life cannot take care of themselves, for different degrees of disability, Disability rate as high as 75%. Stroke is divided into two categories for ischemic stroke and hemorrhagic stroke, The commonest one was ischemic stroke. Lack of perfusion brain tissue would cause neural cell swelling, degeneration, necrosis, apoptosis, then appears energy metabolism disorder and a series of reactions, even loss of its function, led to a series of central nervous system symptoms. Stroke and relevant cerebrovascular diseases. cerebral hypoperfusion induced memory, cognitive and behavioral change,can cause severe cognitive dysfunction syndrome. Current clinical treatment on chronic cerebral ischemia is mainly to restore blood oxygen, inhibit the onset location inflammatory reaction and maintain neuron structure, protect nerve function. Recent studies show that energy metabolism disorder is an important initiating factor of cerebral ischemia, the main damage mechanism relates to acidosis secondary to energy metabolism disorder, excessive depolarization of cell membrane, ion disorder, oxidative stress injury, inflammatory reaction and apoptosis etc. Amp activated protein kinase, a cell energy receptor, has confirmed played an important role in myocardial ischemia, obesity, diabetes, ect energy metabolism disorder. Mammalian rapamycin target protein, a contrary energy sensor, confirmed to play a key role in the research field of tumor.Berberine, a common vision of quinoline alkaloids, The molecular formula is C20H18NO4, It exists in the Berberidaceae four families and ten genera. Its initial widely used in clinical application as heat clearing and detoxifying and antibacterial drugs. In recent years, with in-depth study of berberine and its derivatives found that, berberine may be a drug treatment of cardio-cerebrovascular diseases, diabetes, hyperlipidemia and chronic inflammatory diseases,which has attracted much attention V of the cardio-cerebrovascular and nervous system. But currently, the study of berberine more focused on the acute cerebrovascular disease and nervous system degenerative diseases, dementia has mainly concentrated in alzheimer’s disease research, less research of chronic cerebral ischemia, it remains to be seen whether berberine can improve cognitive function in chronic cerebral ischemia rats through amp activated protein kinase/mammals rapamycin target protein(AMPK/m TOR) signaling pathway. Establishing the model of 2VO rats with bilateral carotid artery ligation were used in the experiment, gives the berberine intraperitoneally(40mg·kg-1) for 14 days, observation of berberine on cognitive function in rats with chronic cerebral ischemia effect through AMPK/m TOR signal pathway application of Morris water maze, immunohistochemistry, Western blot method. Marerials and MethodsSelected 180 healthy male Wistar rats, about 24 weeks, weight about 250±30g, provided by experimental animal supply center in henan province, three groups of sham operation group, model group and intervention group were randomly divided, 60 rats in each group,selected of 2, 4, 8 weeks of the three time points after administration, 20 rats in each group. 2VO model of intervention group were given berberine(40mg?kg-1?d) for 14 days. Establish the model of chronic cerebral ischemia by application of permanent bilateral common carotid artery occlusion(2VO). When administered 2, 4, 8 weeks, cognitive function was evaluated through the Morris water maze test in rats of each group, the expression of rat hippocampal CA1 area AMPK and m TOR changes measured by Immunohistochemical detection, hippocampal nerve cell apoptosis studied by TUNEL test, the expression of rat hippocampal AMPK,p-AMPK,m TOR,p-m TOR were measured by using Western blot. Results1. Behavior observation: comparison of sham operation group rats at each time point, the escape latency had no statistical difference(P>0.05); compared to the sham operation group, the escape latency of the intervention group and model group were significantly prolonged(P<0.05); compared with model group, the incubation period for platform of intervention group was significantly shortened(P<0.05);2. Immunohistochemistry results: AMPK: Comparison in the sham group, no significant difference in the number of immunoreactive cells(P>0.05); compared with the sham operation group,the positive expression of immunoreactive cells in hippocampal CA1 region of model group decreased(P<0.05); compared with the model group, 2 weeks, 4 weeks of immune cells of intervention group was reduced significantly, while 8 week group increased(P<0.05).m TOR: Compared to the sham operation group at each time point, no significant difference in the number of immunoreactive cells(P>0.05); compared with the sham operation group, model and intervention groups immunoreactive positive cell number reduced(P<0.05);compared with model group,immune cells of intervention group significantly increased(P<0.05); with the prolongation of time,the positive cell number of model and intervention group increased(P<0.05).3. The result of TUNEL assay: compared with the sham operation group, the positive cell numbers increased significantly of model and intervention group, the difference was statistically significant(P<0.05); the positive cells of intervention group CA1 region decreased compared with the model group(P<0.05); with the prolongation of time,the positive cell number of model and intervention group significantly reduced(P<0.05).4. Western blot results: activity of AMPK, m TOR by p-m TOR/m TOR and p-AMPK/AMPK to illustrate in hippocampus of each group rats,three time points of AMPK activity, in the state of suppression basically, is very weak in Sham operation group(P>0.05), compared with sham operation group, p-AMPK/AMPK significantly increased of model and intervention groups(P<0.05), compared with model group, p-AMPK/AMPK ratio increased significantly of the intervention group(P<0.05), with the ischemia state continued, AMPK activity reduced gradually(P<0.05); three time points of m TOR activity had no obvious change of sham operation group, compared with sham operation group, p-m TOR/m TOR ratio was significantly reduced of model and intervention groups(P<0.05), compared with model group, p-m TOR/m TOR ratio decreased significantly of the intervention group(P<0.05), with the ischemia state sustained, the m TOR activity further decreased(P<0.05). Conclusion1 The expression of AMPK and m TOR in rats hippocampus could be influenced In the condition of chronic cerebral ischemia, and in a certain time range, with the extension of time, AMPK activity increased, meanwhile,m TOR activity reduced, Their expression was antagonistic at each time point.2 berberine could regulate the expression of AMPK / m TOR in rats hippocampus with chronic cerebral ischemia, in a certain range of time, with the extension of time, AMPK activity further increased, m TOR activity further decreased.3 The condition of chronic cerebral ischemia may lead to cognitive function of rats decreased, the application of berberine conld improve cognitive impairment in rats with chronic cerebral ischemia, its mechanism might be through the activation of AMPK and inhibition of m TOR expression.4 Berberine with AMPK/m TOR pathway might be provided a new direction for the treatment of cognitive disorder in chronic cerebral ischemia. |
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