| Objective: Ischemic cerebrovascular disease is the major threat to thelife and health of residents in our country and has a characteristic of highmorbidity and mortality. It has becomes a hot topic and difficult problem tostudy the pathophysiologic mechanism of cerebral ischemia and search for anefficient way to cure it. The pathological mechanism for cerebral ischemia wasa case of complex cascade reaction, including inflammation, oxidative stress,calcium overload and cell apoptosis and so on, among which inflammationand oxidative stress are considered a major cause in the pathogenesis of braininjury secondary to ischemia, existing in necrosis and ischemic region.Therefore, how to reduce inflammation and oxidative stress have becomeimportant purpose of curing brain injury secondary to cerebral ischemia. Akt,which is also known as protein kinase B, is the main upstream target of theGSK3β. Ample evidences have demonstrated that Akt/GSK signaling pathwayplays a central role in physical and pathological conditions to regulateinflammation factors as well as boost survival. Activated Akt phosphorylatesseveral molecules of apoptosis effect such as the serine-threonine kinaseglycogen synthase kinase3-β (GSK3β)(Ser9), BAD, etc. In order to promotecell survival. So it could resistance to cerebral infarction, and improve theability of brain tissue against brain injury secondary to cerebral ischemia. Thepresent study found that GSK3β regulated the inflammatory response bydifferentially affecting the nuclear amounts of transcription factors NF-κBsubunit p65. Therefore, inhibiting the activity of GSK3β can be used as atarget for the treatment of secondary brain injury after cerebral ischemia. A lotof traditional Chinese medicine composition have the effect of brain protectionvia regulating the activity of Akt. Evodiamine (Evo), an alkaloidal component extracted from the fruit of Evodiae-fructus (Evodia rutaecarpa Benth.,Rutaceae) has vast pharmacological activities. Including antiinflammatory,antitumor, retarding development of atherosclerosis and antinociceptiveeffects. Furthermore, it not only exerts a protective effect onischemia/reperfusion damage in heart and anti-inflammatory properties in thetreatment of Alzheimer’s disease and so on, but also has the effect ofanti-tumor. However, little is known regarding to the effect of Evodiamine inthe acute phase of cerebral ischemia.Methods: The study included two parts: Experiment1, detecting Evo’sneuroprotective effect in cerebral ischemia. Experiment2, evaluating theexpression of phospho-Akt, phospho-GSK3, NF-κB and claudin-5in thecerebral ischemia.Experiment1: Male ICR mice were randomly divided into five groups:sham operated group (0.9%NaCl), pMCAO group (0.9%NaCl), Vehiclegroup (0.9%NaCl+0.1%DMSO+0.5%Tween80), low dose group (50mg/kg)(Evo-L) and high dose group (100mg/kg)(Evo-H).permanent middlecerebral artery occlusion (pMCAO) model was induced by using intraluminalfilament technique in mice. The mice was on the administration for three daysbefore pMCAO, twice a day, and given again thirty minutes before pMCAO.neurological deficit was evaluated, brain water content was measured, andinfarct size was analyzed with2,3,5-triphenyltetrazolium chloride (TTC)24hafter cerebral ischemia.Experiment2: Male ICR mice were randomly divided into five groups:sham operated group (0.9%NaCl), pMCAO group (0.9%NaCl), Vehiclegroup (0.9%NaCl+0.1%DMSO+0.5%Tween80), low dose group (50mg/kg)(Evo-L) and high dose group (100mg/kg)(Evo-H). Western blottingwas used to analyse the expression of phospho-Akt, phospho-GSK3β, NF-κBand claudin-5, while QRT-PCR was used to analyse the expression of NF-κBand claudin-5.Results:1Neurological deficit was evaluated using a modified six point scale24 hours after pMCAO. Mice in pMCAO, Vehicle, low dose group (Evo-L) andhigh dose group (Evo-H) performed a left palsy to varying degrees.Neurological deficit score in evodiamine high dose group (P <0.01) andevodiamine low dose group (P <0.05) was decreased compared with Vehiclegroup.2Brain water content measurement: Sham group was78.68±0.59%.pMCAO group was84.5±0.76%, Vehicle group was84.25±0.29%.Compared with Vehicle group, Evo-L and Evo-H group showed an intensedecline in the percentage of brain water content (Vehicle group vs. Evo-Lgroup:84.25±0.29%vs.83.16±0.49%, P <0.05; Vehicle group vs. Evo-Hgroup:84.25±0.29%vs.82.23±0.31%, P <0.01)(Fig.2D)(n=6in eachgroup).3Compared with Vehicle group (%HLV:43.9±2.61%), the infarctvolume of evodiamine high dose group and evodiamine low dose group wassignificantly reduced (Evo-L%HLV:36.57±3.36%；Evo-H%HLV:32.78±3.78%)(P <0.05).4Evo’s role to pAkt, pGSK3β, NF-κB expression: The expression ofphospho-Akt (P <0.01), phospho-GSK3β (P <0.05) were upregulated atprotein level after systemic administration of Evo. At the same time, thecytosolic NF-B was significantly upregulated (P <0.01) and the nuclearNF-κB was significantly decreased (P <0.05).At mRNA level, the expression of NF-κB was downregulated in Evo-Lgroup (P <0.05) and Evo-H group (P <0.05).5Evodiamine ameliorated BBB permeability. the expression of claudin-5was decressed after cerebral ischemia, Western Blot and QRT-PCR showedthat evodiamine upregulated the expression of claudin-5in the cerebralischemia.Conclusions: Systemic administration of evodiamine after cerebralischemia is effective which can decrease the infarct size, improve the brainedema, and ameliorate the neurological deficit; At the same time, evodiaminecan upregulate the expression of pAkt, pGSK3β and downregulated NF-κB expression, which has a protective effect to the brain tissue after ischemia.Evodiamine also can upregulate the expression of claudin-5, which wasassociated with ameliorating BBB permeability. |
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