| Objective:Stroke,a Chinese medicine called stroke,is the second leading cause of death after myocardial infarction,accounting for 9% of the global death toll,and the third leading cause of adult disability worldwide.The most effective treatment for ischemic stroke in modern medicine is thrombolysis.By eliminating the abnormal state of brain tissue ischemia and hypoxia,the development of ischemic injury and further restoration of damaged tissues and organs are controlled.However,this reperfusion promotes secondary damage to the severe blood vessels and brain tissue in the core region.In this study,the rat model of middle cerebral artery ischemia-reperfusion was used to simulate acute ischemic stroke,and the protective effect of aromatic sputum medicine combined with Huoxue and Lishui compound(hereinafter referred to as comprehensive group)on cerebral edema in acute stroke was studied.Through transcriptomics analysis,the protective mechanism and potential targets of drugs against ischemia-reperfusion injury were explored,which provided a basis for the development and utilization of traditional Chinese medicine.Method: 1.After establishing a rat MCAO/R model and setting model and differentdrug-administered groups,neurological deficits were scored before and beforecerebral ischemia-reperfusion in rats.The mortality of rats was counted.TTCstaining was used to determine the extent of cerebral infarction in rats withischemia-reperfusion injury.2.After establishing a rat MCAO/R model and setting model and differentdrug-administered groups,the dry and wet body weights of the rats were weighedseparately,the brain water content was calculated,and the changes of cerebralblood flow were detected by laser speckle blood flow imager.3.After establishing a rat MCAO/R model and setting model and differentdrug-administered groups,the expressions of AQP4 and NKCC1 m RNA in thepenumbra of rat brain were detected by real-time PCR.The expressions of AQP4,Claudin-5,Occludin and ZO-1 in the penumbra of rat brain were detected Western Blot.4.After establishing a rat MCAO/R model and setting model and differentdrug-administered groups,brain tissue penumbra transcription genes are studiedby high-throughput chips,and differential genes are screened to analyze relatedpathways.5.After establishing a rat MCAO/R model and setting model and differentdrug-administered groups,the differentially expressed genes in the penumbra ofrat brain tissue were verified by real-time PCR.Results: 1.After cerebral ischemia-reperfusion injury,the neurological deficit of the rat isobvious,the cerebral infarction in the middle cerebral artery blood supply area isobvious,and the brain swelling is also obvious,while the Lishui compound group(referred to as Lishui group)and the comprehensive group medium dose(Referred to as comprehensive-M group,it can improve the neurological damageof rats,reduce the volume of cerebral infarction and the percentage of brainswelling,thereby reducing the mortality of cerebral ischemia-reperfusion injuryin rats.2.After cerebral ischemia-reperfusion injury,Lishui group and Comprehensive-Mgroup can effectively reduce brain water content in rats and improve blood flowin cerebral ischemia.3.After cerebral ischemia-reperfusion injury,both Lishui group andcomprehensive-M groups can decrease the expression of AQP4 and increase theexpression of Claudin-5,improve the formation of brain edema and the highpermeability of blood-brain barrier after acute cerebral ischemia-reperfusion.4.According to the chip data,compared with the ischemia-reperfusion model group,there were 9099 differentially expressed genes in the sham operation group,accounting for 34.94% of the total number of detected genes,of which 4188 wereup-regulated and 4911 were down-regulated;differential expression of the Lishuigroup There were 417 genes,accounting for 1.59% of the total number ofdetected genes,of which 251 were up-regulated and 166 were down-regulated;423 genes were differentially expressed in the comprehensive-M group,accounting for 1.61% of the total number of detected genes,of which 215 wereup-regulated and 208 were down-regulated.By analyzing GO and KEGGsignaling pathways for differential gene difference,ischemia-reperfusion injurymainly interacts with Toll-like receptor signaling pathway,TNF signalingpathway,neuroligand receptor,chemokine signaling pathway,and thyroidhormone signaling pathway,PI3K-Akt signaling pathway,MAPK signalinpathway,apoptosis and other pathways.The protective effect of Lishui group andComprehensive-M on cerebral ischemia-reperfusion injury may be related tothyroid hormone signaling pathway.5.After cerebral ischemia-reperfusion injury,the expression level of DIO3 wassignificantly up-regulated in the ischemic penumbra of rats,and the expressionlevel of SLC22a6 was significantly decreased.However,after administration toLishui group and comprehensive-M group,thyroxine was significantly inhibited.The up-regulation of the 5-deiodinase-encoding gene DIO3 m RNA and thedecrease in the organic anion transporter SLC22a6 m RNA.The above resultshave been consistent with the chip results.The STRING analysis byprotein-protein interaction analysis tool showed that AQP4,Claudin-5 and DIO3were in the common interaction network,which further confirmed the reliabilityof the chip results and the biological function of cerebral ischemia-reperfusion.Conclusion:In the rat middle cerebral artery ischemia-reperfusion model,we found thatLishui group and Comprehensive-M group can reduce the mortality of rats,reducethe volume of cerebral infarction caused by acute cerebral ischemia-reperfusionand the percentage of hemispheric swelling,and improve neurological function.Interms of mechanism,the effect of Lishui group and Comprehensive-M group onreducing brain water content and maintaining blood-brain barrier homeostasismay be related to down-regulation of AQP4 expression and up-regulation ofClaudin-5 and ZO-1 expression.The protective effects of Lishui group andComprehensive-M on cerebral ischemia-reperfusion injury may also includethyroid hormone signaling pathway.In conclusion,the Lishui group andComprehensive-M groups may inhibit the expression of DIO3,thereby inhibitingAQP4 and promoting the expression of Claudin-5,and thus protecting the brainedema in acute stroke. |
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