Effect Of NPPB In The Cell Injury Of Glial Cell Line Induced By DDP

Objective:To explore the function of chloride channel blockers NPPB in the cell injury of malignant C6 and U251 glioma cells induced by DDP.Methods:The viability of C6 and U251cells was measured by MTT assay,Bcl-2,Bax and ClC-3 mRNA level was determined by RT-PCR and Bcl-2,Bax,Cyt-C protein level was analyzed by Western Blotting.Results:1.The results of MTT assay shows:Compared with the control group,there is no obvious change of the viability of C6 and U251 cells,the viability ratio of DDP treated group[74.95%(p＜0.05)]decreased obviously.But compared with DDP group, the cell viability ratio of DDP and NPPB treated group increased obviously.2.The result of RT-PCR shows:Compared with the control group,the expression of ClC-3, Bax,Bcl-2 mRNA in NPPB group did not change obviously;the expression of ClC-3, Bcl-2 mRNA in DDP group decreased obviously,but the expression of Bax mRNA increased obviously and the ratio of Bax/Bcl-2 increased.Moreover,compared with DDP and NPPB treated group,the expression of ClC-3mRNA increased,the ratio of Bax/Bcl-2 decreased.3.The result of western blotting shows:Compared with the control group,the ratio of Bax,Bcl-2 and Cyt-C protein level with NPPB treated group did not change obviously;the ratio of Bax/Bcl-2 and Cyt-C protein level with DDP treated group increased obviously.But the ratio of Bax/Bcl-2 and Cyt-C protein level with DDP and NPPB treated group decreased obviously.Conclusion:1.DDP can decrease the viability of C6 and U251cells obviously. 2.DDP can decrease the expression of ClC-3 obviously and increase the ratio of Bax/Bcl-2 and Cyt-C protein level as the same time;however as the decreasing of the expression of Bcl-2 and the increasing of the expression of Bax,DDP perhaps can induce the apoptosis of the cells,even the death.3.NPPB can reverse the decreasing of the expression of ClC-3 obviously by inducing of DDP;decrease the ratio of Bax/Bcl-2 and Cyt-C protein level.From the above,it is proved that there is a closed relationship between the apoptosis of the cells and the control of C6 and U251 cells' reproduction by the control of DDP.4.Chloride channel blockers NPPB can reverse the inhibition of DDP on the proliferation of C6 and U251cells by modulating the expression of Bax, Bcl-2 and the release of Cyt-C.