The Mechanism And Significance Of PD98059 On Cerebral Glutathione Content After Resuscitation In Rats

Background and objective: Cerebral ischemia reperfusion injury(IRI)is a vital factor which affects the prognosis and survival status for those suffering from cardiac arrest/cardiopulmonary resuscitation(CA/CPR).Meanwhile,several studies has demonstrated that inhibiting excessive oxidative stress contributed to alleviating cerebral IRI.Meanwhile,as an endogenous antioxidant,glutathione(GSH)is the major substance whose function is maintaining redox balance in vivo.Moreover,during the period of cerebral insults,GSH promote cell survive by down-regulating oxidative stress via reducing reactive oxidative species(ROS).Previous studies from our group have found that PD98059(PD),at the dose of 0.3mg/Kg,reduced ROS level by inhibiting the activity of Extracellular signal regulated kinas(ERK),which eventually attenuated cerebral IRI.However,it still remains obscure whether PD plays its neuroprotective effect through alleviating oxidative stress via increasing cerebral GSH content.Hence,this study aims to explore the effect of ERK specific inhibitor PD98059 on cerebral glutathione content after resuscitation and to investigate its possible mechanism in rat CA/CPR model.Methods: Adult male Sprague-Dawley(SD)rats were selected to establish CA/CPR model by esophageal electrical stimulation,and conventional CPR was engaged after 6min of CA.When restoration of spontaneous circulation was establish,all rats were divided into PD98059 group(PD group,n=20)and model group(NS group,n=20)randomly.0.3mg/Kg PD98059 or the same volume of saline was injected respectively.Another 20 rats were chosen as Sham group,which were only given tissue separation,trachea and femoral arteriovenous intubation but without inducing CA/CPR by esophageal electrical stimulation.24 hours later,after assessing neurological defect score(NDS),all rats were euthanized,Nissl staining and Nissl body counting was performed to assess cerebral damage,the glutathione content and glutathione reductase activity were detected by chemiluminescence method;the expression of ERK1/2,p-ERK1/2,P62,system Xc-(SLC7A11)glutathione reduce(GR)and glutamate-cysteine ligase catalytic subunit(GCLC)were detected by immunoblotting.Results: After 24 hours later,neurological defect score in Sham group was full(80 points).Nissl staining showed that the morphology of neuron in Sham group was intact,good and clear with obvious nuclear,and blue granular Nissl bodies were distributed around the nuclear.Compared with Sham group,both the neurological defect score and the number of Nissl body were significantly reduced in NS group with nuclear morphology incomplete deep staining and obscure,the distribution of Nissl bodies around the nuclear was less than that in Sham group(P < 0.05).Compared with NS group,neurological defect score was significantly increased in PD group,Nissl staining showed that both the morphology and the number of neurons were improved with a clear nuclear and the number of Nissl body was increased in PD group.In addition,although there was no significant difference in total glutathione content among all groups,oxidized glutathione content,system Xc-,GCLC expression and p-ERK/ERK ratio in NS group were significantly higher than those in Sham group,while reduced glutathione content,glutathione reductase activity,glutathione reductase and P62 expression were significantly decreased in NS group.Compared with NS group,cerebral oxidized glutathione content,system Xc-,GCLC expression and p-ERK/ERK protein ratio in PD group were significantly decreased,while glutathione content,glutathione reductase activity,glutathione reductase and P62 expression were significantly increased in PD group.There was no significant difference in the data above between PD and Sham group(P > 0.05).Conclusion: As an inhibitor of ERK signal pathway,PD98059 alleviated cerebral IRI by increasing reduced cerebral reduced glutathione content and maintaining cerebral redox homeostasis through reviving both the expression and activity of cerebral glutathione reductase after CA/CPR.Results from this study provide a new idea for exploring and rescuing cerebral insult after CA/CPR.