The Study On Protective Effect And The Mechanism Of Erythropoietin On Oxygen-glucose Deprivation Neurons Via PI3K/Akt And Erk1/2 Pathway

Objective: To investigate the protective effects of erythropoietin(Epo)on oxygen-glucose deprivation(OGD)neurons as well as the relevant signal pathways and molecular mechanism of action.Metheds : Cortical neurons in suckling mice were isolated and cultivated.Identified neurons oxygen-glucose deprivation models were constructed.The pre-experiment of different concentrations of Epo intervention,using flow cytometry(Annexin V/PI staining),the group apoptosis rate were measured and calculated the most suitable concentration of Epo as the drug concentration in the follow-up experiment.Experimental grouping:(1)control group(normal neurons group);(2)model group(oxygen-glucose deprivation neurons);(3)Epo group(intervention of Epo group);(4)LY294002 group(intervention of Epo combined LY294002 group);(5)U0126 group(intervention of Epo combined U0126group).The group apoptosis rate were measured by flow cytometry.The transcriptional level of Bax and Bcl-2 was detected by RT-PCR,while the expression of Akt,p-Akt,Erk1/2,p-Erk1/2,Bax,Bcl-2 and caspase-3 was detected by Western blot.Results : 1.Epo could reduce oxygen-glucose deprivation induced neuronal apoptosis(P<0.05).In addition,it showed the strongest protective effects at the concentration of 12.5U/ml(P<0.05).2.Compared with the control group,model group showed associated with increased apoptosis rate(P<0.05);up-regulated the transcriptional level of Bax and Bcl-2(P<0.05);Akt?Erk1/2 and Bcl-2 was no difference(P>0.05),Akt phosphorylation was inhibited and Erk1/2 phosphorylation was elevated,thus inducing up-regulationed of downstream Bax and caspase-3(P<0.05).3.Compared with the model group,Epo group showed down-regulated the transcriptional level of Bax while up-regulated that of Bcl-2(P<0.05);Akt and Erk1/2 was no difference(P>0.05),Akt phosphorylation was elevated,while Erk1/2 phosphorylation was reduced,thus down-regulating the expression of downstream Bax and caspase-3 while up-regulating that of Bcl-2(P<0.05).4.Compared with the Epo group,LY294002 group showed associated with increased apoptosis rate(P<0.05);up-regulated the transcriptional level of Bax while down-regulated that of Bcl-2(P<0.05);Akt was no difference(P>0.05),Akt phosphorylation was inhibited,thus inducing up-regulationed of downstream Bax and caspase-3 while down-regulationed of Bcl-2(P<0.05).5.Compared with the Epo group,U0126 group showed reduced apoptosis rate(P<0.05);down-regulated the transcriptional level of Bax(P<0.05)while the transcriptional of Bcl-2 was no difference(P>0.05);Erk1/2was no difference(P>0.05),suppressed Erk1/2 phosphorylation,thus resulting in down-regulation of downstream Bax and caspase-3 and up-regulation of Bcl-2(P<0.05).Conclusion: Epo inhibits oxygen-glucose deprivation induced neuronal apoptosis,which is achieved through regulating Akt and Erk phosphorylation.