Change of Autophagic Activity of U266 Cells after Bortezomib -Resistance and Its Mechanisms

Abstract

To explore the changes of autophagic activity after resistance of U266 cells to bortezomib (Bor) and its mechanisms. The proliferation inhibition rate, 50% inhibitory concentration (IC50), drug-resistance coefficient, drug-resistance reversed multiple by 3-methyladenine (3-MA) in U266 and U266/Bor cells treated with Bor were detected and calculated by using MTT method, then the proliferation inhibition curve was drawed. The Western blot was used to detect the expression of LC3-I, IC3-II, p-mTOR, Beclin-1, ATG5 and ATG7 proteins. The Bor showed the its proliferation inhibition effect on U266 cells and U266/Bor cells, IC50 of Bor on U266 and U266/Bor cells on 24 hours were 35.7 nmol/L and 526.5 nmol/L respectively; the drug-resistance coefficient was 14.7; the drug-resistance reversed multiple by 3-MA was 2.7. The expression of LC3-II, Beclin11, ATG5 and ATG7 in U266/Bor cells was higher than that in U266 cells; after the treatment with Bor for 24 h, the expression levels of LC3-II, Beclin-1, ATG5 and ATG7 in U266 cells all decreased, as compared with levels before treatment; while the expression levels of LC3-II, Beclin-1, ATG5 and ATG7 in U266/Bor cells were higher than those before treatment. There were no significant difference of p-mTOR expression among U266, U266+Bor, U266/Bor, U266/Bor+Bor cells. The increase of autophagy closely relates with resistance of U266 cells to bortezomib, moreover with up-regulation of Beclin-1, ATG5 and ATG7 expression.