| 1 PurposePanax notoginseng is widely used in clinic and has good pharmacological activity in stroke,myocardial ischemia and other cardiovascular and cerebrovascular diseases.At present,there are few studies on the protective effect and mechanism of different processed products of Panax Notoginseng on cerebral ischemia-reperfusion injury.In this study,through comparing the pharmacodynamic difference of ethanol extract of raw and steamed Panax Notoginseng on cerebral ischemia-reperfusion injury,we determined that the pharmacological activity of raw Panax notoginseng was strong,and further explored the protective effect and mechanism of Panax Notoginseng on blood-brain barrier injury after cerebral ischemia-reperfusion from both in vivo and in vitro,so as to provide reference for clinical application of different processed products of Panax notoginseng in the prevention and treatment of ischemic stroke.2 Methods2.1 Pharmacodynamic difference of ethanol extract of raw and steamed Panax Notoginseng on cerebral ischemia-reperfusion injury in miceC57BL/6 mice were divided into six groups: sham operation group,model group,ethanol extract of raw Panax notoginseng group(200mg/kg),ethanol extract of raw Panax notoginseng group(400 mg/kg),ethanol extract of steamed Panax notoginseng group(200 mg/kg),ethanol extract of steamed Panax notoginseng group(400 mg/kg).The middle cerebral artery occlusion model of mice was established by thread embolism method.The neurological function score,cerebral infarction volume,MDA and MPO contents were used as pharmacodynamic indexes.The pharmacodynamic differences of ethanol extract of raw and steamed Panax Notoginseng on cerebral ischemia-reperfusion injury in mice were compared by observing the pathological sections of brain tissue.2.2 Protective effect and mechanism of ethanol extract of raw Panax Notoginseng on cerebral ischemia-reperfusion injury in miceC57BL/6 mice were divided into four groups: sham operation group,model group,ethanol extract of raw Panax notoginseng group(200mg/kg),ethanol extract of raw Panax notoginseng group(400 mg/kg).The middle cerebral artery occlusion model of mice was established by thread embolism method.The neurological function score,cerebral infarction volume,Evans blue content,MDA and SOD content were used as pharmacodynamic indexes to observe the pathological section of brain tissue and further study the pharmacodynamic effect of Panax Notoginseng on cerebral ischemia-reperfusion The expression of ZO-1,occludin,MMP-2,MMP-9 and MAPK signaling pathway related proteins in brain tissue were detected by western blot and immunohistochemistry to further study the mechanism of Panax notoginseng improving blood-brain barrier injury after cerebral ischemia-reperfusion.2.3 protective effect and mechanism of ethanol extract of raw Panax Notoginseng on mouse brain microvascular endothelial cells injury induced by OGD/RbEnd.3 cells as the research object,They were divided into five groups: normal group,model group,ethanol extract of raw Panax notoginseng group(50 μg/ml),ethanol extract of raw Panax notoginseng group(100 μg/ml)and ethanol extract of raw Panax notoginseng group(200 μg/ml).The model of oxygen glucose deprivation/reoxygenation was established in three gas incubator.The cell viability was determined by MMT method,and the cell morphology was observed to determine the protective effect of ethanol extract of raw Panax Notoginseng on bEnd.3cells after OGD/R treatment The tight junction proteins ZO-1,occludin,MMP-2,MMP-9 and MAPK signaling pathway related proteins were detected by western blot,and the mechanism of action was further studied at the cellular level.3 Results3.1 pharmacodynamic difference of ethanol extract of raw and steamed Panax Notoginseng on cerebral ischemia-reperfusion injury in miceCompared with the model group,the ethanol extract of raw and steamed Panax notoginseng could reduce the neurological function score and cerebellar infarction volume,but the effect of the ethanol extract of raw Panax notoginseng was significantly better than that of the ethanol extract of steamed Panax notoginseng;The results of MDA and MPO content showed that the content of MDA and MPO in each treatment group were significantly decreased,but the effect of alcohol extract of raw Panax notoginseng was significantly better than that of alcohol extract of steamed Panax notoginseng;he staining results showed that the pathological state of brain tissue in each treatment group was improved,but the effect of high-dose alcohol extract of raw Panax notoginseng was the best,which indicated that alcohol extract of raw Panax notoginseng had the best effect The effect was better than that of alcohol extract of steamed Panax notoginseng.In conclusion,the effect of ethanol extract of raw Panax Notoginseng on cerebral ischemia-reperfusion injury was significantly better than that of ethanol extract of steamed Panax notoginseng.3.2 protective effect and mechanism of ethanol extract of raw Panax Notoginseng on cerebral ischemia-reperfusion injury in miceCompared with the model group,the neurological function score and infarct volume of the ethanol extract group were significantly decreased In addition,the expressions of ZO-1 and occludin were significantly increased,the expressions of MMP-2 and MMP-9 were significantly decreased,and the phosphorylation levels of p38,ERK and JNK were significantly decreased.3.3 protective effect and mechanism of ethanol extract of raw Panax Notoginseng on mouse brain microvascular endothelial cells injury induced by OGD/RCompared with the model group,the cell viability and cell state were significantly improved in the ethanol extract group;the expressions of ZO-1 and occludin were significantly increased,the expressions of MMP-2 and MMP-9 were significantly decreased,and the phosphorylation levels of p38,ERK and JNK were significantly decreased in the ethanol extract group.4 ConclusionAfter confirming that the ethanol extract of raw Panax notoginseng is better than the ethanol extract of steamed Panax notoginseng in anti cerebral ischemia-reperfusion injury,we further explored the protective effect and mechanism of ethanol extract of raw Panax Notoginseng on cerebral ischemia-reperfusion injury.The results of animal experiments show that the ethanol extract of raw Panax notoginseng can improve the neurological function injury after cerebral ischemia-reperfusion,reduce oxidative stress injury,and play a protective role in maintaining the permeability of blood-brain barrier,so as to play a protective role in cerebral ischemia-reperfusion injury.The mechanism may be related to the inhibition of the activation of MAPK signaling pathway,the decrease of MMP-2 and MMP-9 protein expression and the up regulation of tight junction related proteins. |
