Inhibition of xCT with sulfasalazine blocks cystine uptake and ROS detoxification by metaplastic cells. (A) Relative mRNA expression of Cd44v9 and xCT in ImChief and ImSPEM cells determined by reverse-transcription quantitative PCR (P = .0002∗∗∗ and .0149∗, respectively). (B) ImChief and ImSPEM cells were immunostained for CD44v9 (red) with nuclear counterstain 4′,6-diamidino-2-phenylindole (DAPI) (blue). Scale bar: 100 μm. (C) Percentage of cells per 20× field that are CD44v9-positive (P < .0001∗∗∗∗). (D) Diagram of ROS indicators CellROX green reagent and DHE. (E) Representative fluorescence images of cystine-FITC (green), glutathione (green), CellROX (green), and oxidized DHE (DHE-ox) (red) in ImSPEM cells ± sulfasalazine with nuclear counterstain Hoechst (blue). Scale bar: 100 μm. (F) Relative fluorescence intensity of cystine-FITC (P = .007∗∗), glutathione (P = .001∗∗), CellROX (P = .0019∗∗), and DHE-ox (P = .0003∗∗∗) in ImSPEM cells ± sulfasalazine. Statistical significance was determined by unpaired Student t test (n = 4 per condition).