RÉSUMÉ
Objective To investigate the effect of endoplasmic reticulum stress on sensitivity of glioma cell line U87 to temozolomide (TMZ) and underlying mechanism.Methods (1) Glioma U87 cells were routinely cultured for 24 h in vitro;different concentrations of TMZ (0,12.5,25,50,100 μmol/L) were added to intervene the cell proliferation of U87 cells for 24 h;MTT method was used to detect the cellular proliferation inhibition rate,and half maximal inhibitory concentration (IC50) of TMZ was calculated;tunicamycin (TM) treatment (0,2,4,8 μmol/L) was given to the cells for 6 h,and then,50 μmol/L TMZ was given for 24 h,and cellular proliferation inhibition rate of U87 was detect by MTT method.(2) U87 cells were randomly divided into control group,TM group,TMZ group and TM+TMZ group;pretreatment of TM for 6 h was given to cells from TM group and TM+TMZ group;and 50 μmol/L TMZ was given to cells from TMZ group and TM+TMZ group;same amount of medium was given to cells from control group;24 h after treatment,the apoptotic rate was examined by flow cytometry;Westem blotting was used to detect the protein expressions of caspase-3,B-cell lymphoma 2 associated X protein (Bax),O-6-methlguanine-DNA methyltransferase (MGMT),glucose-regulated protein 78 (GRP78),and inositol-requiring enzyme 1 (IRE-l).Results (1) MTT showed that IC50 of TMZ was 50 μmol/L.As compared with that of 0 μmol/L TM+50 μmol/L TMZ group or 2 μmol/L TM+50 μmol/L TMZ group,the cellular proliferation inhibition rate of 4 μmol/L TM+50 μmol/L TMZ group and 8 μmol/L TM+50 μmol/L TMZ group was significantly decreased (P＜0.05).(2) As compared with TMZ group,TM+TMZ group had significantly decreased apoptotic rate (46.98％±4.79％ vs.35.74％ ±4.09％),significantly decreased caspase-3 and Bax protein expressions,and significantly increased MGMT,GRP78 and IRE-1 protein expressions (P＜0.05).Conclusion Endoplasmic reticulum stress can increase the resistance gene MGMT expression,decrease the chemotherapy sensitivity to TMZ,and induce chemoresistance ofglioma cell line U87.
RÉSUMÉ
Objective To investigate the effect of Eicosapentaenoic scid (EPA) on the sensitivity of glioma cell line U87 to temozolomide (TMZ) and the mechanism behind this effect.Methods U87 cells were randomly divided into four groups:control group,TMZ group,EPA+TMZ group and endoplasmic reticulum stress (ERS) activation tunicamycin group (EPA+TMZ+TM group).MTT method was used to evaluate inhibition ratio of cell proliferation.The apoptotic ratio was examined by flow eytometry.Western blot was used to detect the protein expressions of apoptosis cytokines (caspase-3 and Bax) and ERS cytokines [glucose-regulated protein 78 (GRP78) and inositol-requiring enzyme 1 (IRE-1)].Results EPA causes concentration-dependent and time-dependent inhibition of the cell proliferation (all P=0.00).EPA significantly enhanced the sensitivity of glioma cell line U87 to temozolomide.Compared to TMZ treatment alone,the inhibition ratio [(56.27+6.15)％ vs.(42.32±4.12)％,P=0.03] and apoptotic ratio [(49.78±5.94)％ vs.(37.74± 4.24)％,P=0.04] of U87 cells were enhanced by EPA+TMZ treatment.Western blot showed that the expression of apoptotic factor caspase-3 and Bax proteins were increased by EPA+TMZ treatment,while the protein expressions of ERS-related factors (GRP78 and IRE-1) were significantly inhibited (P=0.01).However,the salutary effects of EPA were reversed by ERS activation tunicamycin.Conclusion EPA enhances the sensitivity of glioma cell line U87 to temozolomide,the mechanism of which may be the suppression of ERS response.