| BackgroundCerebrovascular disease is common and frequently occurring serious hazards to human health, has become the first disability and the third death in the world, the incidence of ischemic cerebrovascular disease accounting for75%of cerebrovascular disease. The early treatment in the window period for thrombolytic to restore the blood supply to ischemic and the surrounding areas, and re-supply of the oxygen and reversibility of certain neurons damage in a timely manner to obtain the functional recovery of ischemic brain tissue. Reflow blood not only failed to be effectively restored, but more emphasis on the destruction of the ischemic core and the surrounding cells of the blood brain barrier. Caused by cerebral edema and neuronal apoptosis and necrosis, leading to severe brain dysfunction, this is called cerebral ischemia and reperfusion injury CIRI (Cerebral Ischemia-Reperfusion Injury). In recent years, studies have shown that thrombin neurotoxin injury in the CIRI evolution process occupy a very important position, especially as the central nervous system of the discovery of the thrombin receptor,but the thrombin damage mechanisms and application of thrombin inhibition agent protection mechanisms of CIRI has not been fully elucidated. This experiment starting from the"Hot toxic genomics"perspective of Chinese medicine, the use of intraluminal filament in rats middle cerebral artery ischemia-reperfusion model to explore the mechanism of the CIRI thrombin neurotoxicity and application of baicalin intervention, and provide a new ideas and methods to the clinical treatment of CIRI for Chinese medicine treatment of cerebrovascular disease.ObjectiveBy the intraluminal filament middle cerebral artery after focal ischemia and reperfusion model. To observe the different dose of baicalin in rat focal cerebral ischemia reperfusion injury of rat brain function of different time points:Assess the defect score^pathological changes/ischemic and PAR-1protein levels of brain tissue of> NF-KBp65and Caspase-3positive cells in the level of detection. Explore the possible mechanism of baicalin on the CIRI thrombin neurotoxicity.Methods1. Preparation of rat focal the CIRI model:Young male Wistar rats of SPF grade, weighing280-320g, adaptive feeding for one week, then randomly divided rats into:sham operation group, model group, baicalin(Low-dose, Medium, High-dose group)(25、50、100,mgkg), control group, n=15of the team, prepare Arterial cerebral ischemia and reperfusion model in the instraluminal filament in rat brain after ischemia and3h reperfusion, the peritoneal cavity immediately of the treatment group and control group after surgery, Algarrobas injection saline group, once a day.2. Assess nerve function score after surgery3h、1、3、7day, take the ischemic brain tissue in fixed and frozen to preparations on1、3、7day.3. By conventional HE staining of brain tissue pathological change.4. Western blot was used to observe the ischemic region of PAR-1protein expression.5. Immunohistochemical method to observe the expression of NF-κBp65and Caspase-3positive cell number.Results1. Observed each group of rat brain function of5-point score:Neurological score compared with the sham operation group, model group was significantly increased, different doses of the treatment group at different time points with model than model group, the neurological score decreased significantly. And the7days of high dose group, the neurological score was significantly better than3h, compared with control group almost significant difference at all.2. Use Microscope staining brain path morphology structural change after HE:the sham group structure of the hippocampus is normal, neuronal cells pigmented uniform. The model group, hippocampus、striatum appear small focal ischemic area, part of the transformation of neurons showed the characteristics of apoptosis, chromatin condensation. The treatment group, can see the nervous derangement, but the extent is significantly reduced, some neurons to restore basic morphology, and the most obvious improvement of baicalin high dose groups of7days.3. Western blot were observed PAR-1protein levels in the ischemia brain tissue of rats: sham operation group PAR-1protein expression was maintained at a low level and relatively constant. Compared with the sham group, PAR-1continued to increase with model group. Treatment group, the PAR-1protein expression was lower levels, and although increase was significantly reduced7days dose of the high dose expression close to the sham group, no significant difference compared with the control group.4. Immunohistochemical method to observe the NF-κBp65number of positive cells: the sham group showed obvious positive cells in the cytoplasm and nucleus have different levels of expression of the model group after reperfusion, the apparent distribution in the ischemic core and penumbra, shrinkage and swelling, distorted, distortion and other forms of distribution, the treatment group, the infarct border, showing morphology close to normal neurons, especially in the treatment of high-dose7days groups of neurons have been restored.5. Immunohistochemical method to observe the caspase-3positive cells:positive expression of sham-operated group remained at a low level, the model group have a large number of densely distributed in the cortex and striatum of apoptotic cells and apoptotic bodies, cytoplasm, cell nuclear coloring is relatively deep compared with model group, treatment group coloring lighter cytoplasm, reduced number of apoptotic cells, and Caspase-3in the strongest, with the control group compared to7days dose group no significant difference。ConclusionEffect of baicalin in neuron protective on focal cerebral ischemia-reperfusion injury, the mechanism is by inhibiting the expression of PAR-1, and thus further inhibit thrombin neurotoxin damage, reduce the activation of PAR-1receptor inflammatory cascade reaction and apoptosis, cerebral edema, and protect neurons at three aspects. |
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