The Effect Of Bevacizumab On The Tolerance Of Glioblastoma To Chemo Drugs Through Autophagy

Objective:to discuss the inhibition effect of bevacizumab to glioblastoma cells and the mechanism of induced tolerance of the cells to bevacizumabMethods:Glioblastoma cells(U87-MG)were cultured in vitro.The glioblastoma cells were processed with different concentration of bevacizumab for 24 hours or 48 hours,the survival rate of cells were tested using MTT kit to testify the direct inhibition effect of bevacizumab to the glioblastoma cells.The apoptosis of the glioblastoma cells with different bevacizumab concentration treatment were tested using Annexin V-FITC/PI kit and flow cytometry.The expression level of autophagy marker(LC3B-I?LC3B-? ? SQSTM1(p62))were tested using Western Blot after 48 hours since different concentration bevacizumab treating.The expression level of AKt-mTOR signal related protein was tested to find the mechanism of autophagy induced by bevacizumab.When the glioblastoma cells were cultured by bavacizumab combined with chloroquine,the expression level of autophagy marker(LC3B-I?LC3B-II? SQSTM1(p62))were tested to testify the inhibition of autophagy.Using Annexin V/PI dye and flow cytometry to analyze the apoptosis of glioblastoma cellstreated with bevacizumab alone or combined with chloroquine.Results:The proliferation of glioblastoma cells were inhibited directly by bevacizumab and the inhibition was depended on dosage and timing.The apoptosis of glioblastoma cells were triggered with high level by bevacizumab and the tolerance was induced under the high concentration of bevacizumab.The up regulation of LC3B-? expression and down regulation of SQSTM1(p62)expression were demonstrated after treatment with bevacizumab,which mean autophagy was induced effectively.The change of expression level of Akt-mTOR signal related protein were exist and the autophagy was obviously suppressed by over expression of LC3-II?SQSTMl(p62).The higher level of apoptosis was observed when glioblastoma cells were treated with bevacizumab combined with chloroquine.Conclusion:Bevacizumab directly inhibits the proliferation of glioblastoma cells in vitro,and promotes the apoptosis of the cells.The high concentration of bevacizumab results in drug tolerance of glioma cells.The drug tolerance is implement through suppression of Akt-mTOR signal pathway.The autophagy inhibitor may reduce the drug tolerance of glioblastoma cells to bevacizumab.