The Effects Of SAS And Glutamate On Glioma Cells CLIC4 And The Apoptosis Related Genes

Objective:To explore the changes and the possible functions of mtCLIC/CLIC4 (mitochondrial chloride intracellular channel 4) proteins in malignant C6 glioma cells treated with glutamate and SAS.Methods:The viability of C6 cells was measured by MTT assay,LDH release rate and the content of reduced glutathione in cell were detected by ultraviolet spectrophotom-etry,mRNA levels of EAAT3,mGlu4,mGlu6,Bcl-2,Bax,CLIC4 were determined by RT-PCR,EAAT-3,CLIC4,Bcl2,Bax and p38 protein levels were detected by Western Blotting.Results:1.The proliferation of C6 glioma was significantly inhibited by SAS and glutamate in a dose-dependent manner.But the viability of C6 glioma in SAS combined with glutamate group was significantly lower than each group.2.The LDH release rate in SAS group was increased,but in glutamate group was not changed. 3.The content of GSH in SAS group and glutamate group all decreased,while in SAS combined with glutamate group decreased more remarkable.4.Compared with the control group,the Bcl-2mRNA level in SAS(1.5mmol/L) group was lower,Bax was constant;The mRNA level of CLIC4 in glutamate(30mmol/L) group and SAS combined with glutamate group was lower;the mRNA expression of EAAT-3 in SAS group deceased;The mRNA expression of mGlu4 and mGlu6 in glutamate(30mmol/L) group decreased;5.The protein expression of CLIC4,Bax and p38 in SAS group was higher than control group,but Bcl-2 was lower,the other groups was constant;The protein expression of EAAT-3 in SAS group decreased.Conclusion:1.The SAS and glutamate all could induce glial cell injury;2.Exhaust of GSH induced by oxidative stress could be the main mechanism of glial cell injury induced by SAS and glutamate;3.The heightened LDH,decreased expression of Bcl-2 and increased expression of Bax in glial cell deal with SAS suggested that SAS induced the apoptosis of cell,glutamate induced the necrosis of cell because of the indexes above didn't change in the process of working cell injury;4.SAS heightened the expression of CLIC4 which might concern with death of cell,the heightened expression of p38 suggested the signal passageway of p38MAPK might participate in the transmit passageway which SAS induced cell injury and necrosis;5.The decreased expression of EAAT-3 induced by SAS and mGlu4 and mGlu6 resulted by glutamate might participate in the cell injury and intercession progress above,the SAS and glutamate decreased the protect action of their transporter and metabotropic glutamate receptor(mGluR).But,the mechanism expected to be investigated further.