The effect of Epo/EpoR pathway on Sunitinib in 786-0 cells.

(A) 786-0 cells stably expressing EpoR siRNA, 786-0 cells stably expressing negative siRNA, and parental 786-0 cells were incubated in the medium containing various concentrations of Sunitinib for 24 hours, and the viable cells were measured by modified MTT method. Error bar represents mean ± SD from 6 wells for each time point and triplicate experiments for each sample. 786-0 cells stably expressing EpoR siRNA were slightly more sensitive to Sunitinib than the two controls. (B) 786-0 cells stably expressing EpoR siRNA were cultured in medium containing 5 µg/ml Sunitinib for 24 hours, then subjected to assess cell apoptosis by flow cytometry. Suntinib treatment caused more apoptotic cells(Annexin V-PE⁺/7-AAD^+/−)in 786-0 cells stably expressing EpoR siRNA than in 786-0 cells stably expressing negative siRNA (★★★: P<0.001). (C) 786-0 cells were exposed to various concentrations of Sunitinib with or without 10 U/ml Epo for 24 hours and assessed relative viable cell number by modified MTT assay. Error bar represents mean ± SD from 6 wells for each time point and triplicate experiments for each sample. Epo slightly antagonized the effect of Sunitinib. (D) 786-0 cells were cultured in medium containing 1 µg/ml Sunitinib with or without 10 U/ml Epo for 24 hours, and apoptotic cells (Annexin V-FITC⁺/7-AAD^+/−)were measured by flow cytometry. Epo attenuated the effect of Sunitinib (P<0.01, from 4 repeated experiments).