Sox2 overexpression abolished the inhibitory effects of AP on DOX-induced stemness of osteosarcoma cells. (A) Flow cytometry analysis of the CD133+ subpopulation of KHOS and U2OS cells treated with DOX, AP+DOX, AP+DOX+vector, or AP+DOX+Sox2. (B) After transfecting with the empty vector or Sox2 lentiviral vector and culturing with DOX or DOX+AP for 7 days, KHOS and U2OS cells were subjected to a sphere-formation assay. (C) The percentage of the CD133+ subpopulation of KHOS and U2OS cells treated with DOX, AP+DOX, AP+DOX+vector, or AP+DOX+Sox2 was calculated. (D) The sphere numbers of the KHOS and U2OS cells were counted. (E and F) Western blot analysis of Oct4 and Nanog expression of KHOS and U2OS cells treated with DOX, AP+DOX, AP+DOX+vector, or AP+ DOX+Sox2. Data are represented as the mean ± SD. ***P < 0.001. AP = apatinib; DOX = doxorubicin; SSC = Side SCatter; FITC = fluoresceine isothiocyanate; SD = standard deviation.