Salvianolic Acid B On Cerebral Ischemia And Reperfusion Of Neuronal Apoptosis And Its Mechanism In Mice

Cerebrovascular disease is a clinical common, frequently encountered disease in the gerontal patient. The rates of morbility, deformity and mortality are very high. It is one of the most dangerous diseases that lead significant harm to human health. Among them, the incidence of ischemic cerebrovascular disease is more accounted about 70﹪in the clinic. Following the accelerating step of population aging,cerebrovascular disease, which brought us deadweight both in economy and society, has the harmful effects on health status and quality of life. Therefore, to seek the medicine with high performance and harmfulless becomes the focus of empirical study.Cerebral ischemia belongs to the ischemic cerebrovascular disease, including cerebral thrombosis, cerebral infarction and cerebral vasospasm. Once brain ischemia persists over 6 min, as a result, the energy exhaustion, signal transduction occurs abnormally, which initiates mitochondrial dysfunction. Those changes cause progressive damage which is comprehensive and irreversible in the high level function of brain. When the blood flow restore after cerebral ischemia, the cell function, dysbolismus and disorganization of the tissue not recover but aggravate instead, that's called reperfusion injury .On the basis of preliminary work, we established the model of acute cerebral ischemia- reperfusion in mice to observe the effects of salvianolic acid B on free radical and apoptosis modulin of neuron and investigate the mechanism of action. It will provide scientific experimental evidence for the clinical application of ischemic cerebrovascular disease.The experiment includes two parts:Part One:Effects of SalB on free radical change after cerebral ischemia- reperfusion in miceNIH mice(Male mice of SPF grade, 25±3 g)were randomly divided into four groups: Sham-operated group, cerebral ischemia-reperfusion model group, SalB-treated group and Nim-control group.The cerebral ischemia-reperfusion model was established by ligating bilateral common carotid arteries in mice for 30 min. After 24 hour's reperfusion, the mice were killed. The activity of superoxide dismutase (SOD), the content change of malondialdehyde (MDA), the nitric oxide synthase (NOS)and the total antioxidative capacity(T-AOC )in cerebral cortex were measured to investigate the effects of SalB on free radical change after cerebral ischemia - reperfusion and analyze the mechanisms.Results:Effects of SalB on SOD,MDA,T-AOC and NOS in cerebral cortex Compared with the sham operated group, the content of MDA was increasing(P﹤0.05)and the actitive of SOD was significant drop in the model group(P﹤0.01). SalB and Nim can improve the activity of SOD and T-AOC, whereas reduce activity or content of MDA and NOS(P﹤0.05 ~ 0.01).Part Two:Effects of SalB on protein expression of Bcl-2 and Bax after cerebral ischemia- reperfusion in miceGroup and model establishment:same as Part OneAfter restored blood flow for 24 h,brain tissue was obtained to make into paraffin section. TO observe the morphological changes of brain tissue by HE staining and determin the protein expression of Bcl-2 and Bax both in hippocampi CA1 area and cerebral cortex by immunohistochemistry.Results:1. Morphology observation and cell population in hippocampi CA1 areaWe observed the pyramidal neuron of hippocamp area of mice by using were light microscop. The results showed that pyramidal neuron in sham operated group were line up in order, nuclear membrane and nucleolus were clear and regular,extracellular space was narrow. The pyramidal neuron of hippocamp area in model group was not only dysmorphosis and pyknosis but also karyorrhexis. The extracellular space increased and cell population grew downwards. The number of pyramidal neuron which had regular shape was increased both in SalB group and Nim group(P﹤0.05 ~ 0.01).2. Effects of SalB on Bcl-2 and Bax protein Expression in cortex and hippocampi CA1 area in ischemia-reperfusion mice:The experimental result show that the expression of Bax significantly increased and the expression of Bcl-2 reduced in model group at 24 h after cerebral ischemia- reperfusion. SalB, as a kind of prophylactic,increased the protein expression of Bcl-2 and reduced the protein expression of Bax.Conclusion:SalB has the following effects on free radical after cerebral ischemia-reperfusion: improving the activity of SOD and T-AOC, whereas reducing activity or content of NOS and MDA, increasing the protein expression of Bcl-2 and the ratio of Bcl-2/Bax and reducing the protein expression of Bax, then reducing the number of apoptotic cells, to alleviate cerebral injury and protect mice brain tissue after ischemia and reperfusion injury.