The Expression Of TRAF6, ERK And P-ERK In Cerebral Ischemia In Rats And The Neuroprotective Effect Of Sophoridine

Objective: Acute ischemic stroke is a mainly cause of death and the mostfrequent cause of permanent disability in adult worldwide. Brain tissueinjuries secondary to ischemia aggravated the illness and inhibited therecovery of patients, its main including cell inflammation, oxidative stress,Ca2+steady-state disorders, cytotoxicity, active oxygen and free radicals etc,this is a complex pathological cascade, inflammatory reaction as human mostdisease pathological mechanism, its role in cerebrovascular disease has beenpaid more and more attention, and at present for excessive inflammation ofbrain damage to further study, how to reduce inflammation in the treatment ofacute cerebral infarction become an important method.In recent years,in the research of inflammation and immune responseprocess tumor necrosis factor receptor related factor6(TRAF6) plays a valuedrole, TRAF6is a member of the TRAF family, which plays physiological rolesdistinct from other TRAFs. TRAF6participates in signal transduction of TNFreceptor (TNFR) superfamily and the interleukin-1receptor (IL-1R)/Toll-likereceptor (TLR) superfamily, leading to the activation of transcription factorssuch as extracellular signal-regulated kinase (ERK) and nuclear factor-kappaB(NF-κB). Many studies had indicated that TRAF6partake in acute traumaticbrain injury and neurodegenerative disease pathogenesis.Sophora flavescens, which contains quinolizidine alkaloidsas its bioactiveconstituents, has long been used in the history in traditional Chinese medicine.Its root is used to treat tumors, fever, throat inflammation and other diseases.Sophoridine (SOP) is a major quinolizidine alkaloid from the root of sophoraflavescens Ait (kushen). It improves cardiac function and protects myocardialcells from reperfusion injury. It has also been shown to antitumor,anti-inflammation and sedation hypnosis action. Our experiment observed the expressions of TRAF6, ERK, p-ERK in rat model of focal cerebral ischemia;investigated the potential neuroprotective effect of sophoridine and underlyingmechansms after cerebral ischemia induced in male adult Sprague-Dawly ratsby permanent middle cerebral artery occlusion(MCAO).Methods: Male, Sprague-Dawley rats were randomly assigned to fivegroups: Vehicle (pMCAO+saline), High dose (pMCAO+SOP10mg/kg),Middle dose (pMCAO+SOP5mg/kg), Low dose (pMCAO+SOP2.5mg/kg)and Sham operated group. Permanent middle cerebral artery occlusion(pMCAO) model was used and SOP administered intraperitoneallyimmediately after cerebral ischemia and once daily on the following days.Neurological deficit was evaluated using a modified six point scale; brainwater content and infarct volume were measured. The expression of TRAF6and ERK1/2were measured by immunohistochemistry and western blotting.Results:1Compared with Vehicle group, water content markedly reduced (Highvs. Vehicle:81.11±2.04%vs.83.87%±0.93%P <0.05) in High dose group,but no statistical significance was observed in Middle and Low dose group(Middle vs. Vehicle:82.24±1.15%vs.83.87±0.93%Low vs. Vehicle:81.99±1.97%vs.83.87±0.93%P>0.05). Moreover, there was a no significantdifference for the effect of SOP on water content between Low dose group,Middle dose group and High dose group.2Compared with Vehicle group, Low dose SOP treatment significantlyreduced the%HLV (Low vs. Vehicle:27.15±5.26vs.43.32±5.70P <0.05)at24hours after MCAO. There was no significant difference in the infarctvolume between Vehicle group and Middle dose and High dose group (Highvs. Vehicle:37.82±6.24vs.43.32±5.70P>0.05).3Compared with Vehicle group, at the dose of Middle and Low dosegroup the neurological function deficit scores were not lowed (P>0.05).Although the deficit scores in High dose group were reduced, there were stillno significant differences in neurological deficit scores between High dosegroup and Vehicle group (P=0.05). 424h after the injury, the number of cells labeled with TRAF6and theprotein level of TRAF6was increased, but phospho-ERK1/2decreased in theischemic cortex. In SOP group, the number of cells labeled with TRAF6in thecytoplasm and cytomembrane was decreased (P <0.01), the difference wasnot significant among different SOP dose groups. The increased protein levelof TRAF6in Vehicle group was decreased by intraperitoneal injection of SOPsignificantly (P <0.01).72h after the injury, in Low dose group, the numberof cells labeled with phospho-ERK1/2was increased compared with Vehiclegroup (P <0.05). The decreased protein level of phospho-ERK1/2in Vehiclegroup was increased in Low dose group (P <0.05).Conclusions: Our study showed that the expressions of TRAF6up-regulated and the p-ERK down-regulated after cerebral ischemia. Systemicadministration of SOP is effective which can decrease neurologic impairmentand tissue injury under cerebral ischemic conditions, and this effect may bethrough down-regulation of TRAF6activation pathway, up-regulation ofp-ERK expression.