The Role Of AMPA-GluR2 In The Apoptosis Of OPCs After Spinal Cord Injury

Objective:To investigate the expression of glutamate receptor(AMPA-GluR2)in oligodendrocyte precursor cells(OPC)before and after spinal cord injury,and to clarify the role of AMPA-GluR2 in the apoptosis OPCs after spinal cord injury.Method:1.OPCs were obtained after dissecting,shredding,digesting,separating and purifying from the brain tissues of specific pathogen free female and male,1-2 days old SD rats.Immunofluorescence was used to mark oligodendrocyte precursor cells with antibody A2B5,and the nucleus was labeled with DAPI,and the cell purity was observed under the microscope.2.The spinal cord injury models were established using specific pathogen free female SD rats with a weight of 200-250 g aging from 6 to 8 weeks.Allen’s attack method is used to make spinal cord injury(T9)model in SD rats.10 g weight,landing height 30 mm,causing T9 spinal cord injury with Spinal NYU IMPACTOR MODEL-II cord percussion apparatus.Injured spinal cord of spinal cord injury without glutamate receptor antagonist group was quickly removed after 12 h under the aseptic condition.In the control group,the spinal cord was removed 12 hours after exposure to the corresponding segments of the spinal cord.Using Transwell Co-culture Technology,culture of OPC cells in culture plate,placement of spinal cord tissue in Transwell small room.They were divided into control group,spinal cord injury group,glutamate excitant group,antagonist group NBQX,antagonist group JSTX.The apoptosis rate in each group by flow cytometry3.Western blot and immunohistochemical method were used to detect the expression of AMPA-GluR2 in the control group and the spinal cord injury group.The role of AMPA-GluR2 in the apoptosis of OPC was discussed according to difference of apoptosis rate in each group.Result:1.OPCs purity identification: after purification,oligodendrocyte precursor cells were photographed with inverted phase contrast microscope after immunofluorescence staining.The apoptosis rate of different groups was detected by flow cytometry,which coThe purity of OPC turned out to be greater than 95% which met the experiment requirements.2.Results of flow cytometry assay showed that compared with the control group,apoptosis rate of spinal cord injury without glutamate receptor antagonist group was significantly elevated(P < 0.001),while apoptosis rate was significantly upregulated when glutamate receptor antagonist was added((P < 0.001)3.Immunohistochemistry showed that AMPA-GluR2 expression was significantly higher in the control group than in spinal cord injury group.Western blot results showed that compared to the control group,expression of AMPA-GluR2 in oligodendrocyte precursor cells was significantly lower in spinal cord injury group both at 24 h and 48 h(P<0.05).In spinal cord injury group,AMPA-GluR2 expression at 48 h group was significantly lower than that at 24 h and no significant difference was found between 24 h and 48 h in the control group(P>0.05).Conclusion:1.The The injured damaged spinal cord can induce OPC apoptosis,and the apoptosis mechanism may be significantly decreased with the expression of AMPA-GluR2.CP-AMPAR can lead to the increase of intracellular calcium flow and induce the apoptosis of OPC.2.NBQX and JSTx,as CP-AMPA receptor antagonists,can block CP-AMPAR to a certain extent,terminate the intracellular rectifying of calcium and promote the survival of cells.