Hri and Atf4 dependent induction of antioxidant genes. (A) Hri-dependent Ho-1 protein expression. Hri^+/+ and Hri^−/− FL cells were treated for 5 hours with increasing concentrations of arsenite as indicated. (B) Atf4-dependent Ho-1 protein expression. Atf4 +/+, +/−, and −/− FL cells were treated with 5μM arsenite for 3 hours. (C-D) Relative expression of mRNA of antioxidant genes. Hri^+/+, Hri^−/−, Atf4^+/+, and Atf4^−/− FL cells were treated with 5μM arsenite for 3 hours. mRNA levels of treated and untreated cells were analyzed by qPCR. Data are presented as mean ± SD (n = 3). P values denote the comparison between arsenite-treated Hri^+/+ and Hri^−/− samples; P < .01 for Ho-1 and Nqo1, P < .05 for Gstμ, and P = .06 for Sod2; or between the comparison of arsenite-treated Atf4^+/+ and Atf4^−/− samples; P < .005 for Chop, P < .05 for Ho-1, P < .001 for Nqo1. (E) Elevated ROS levels in Hri^−/− and Atf4^−/− RBCs upon H₂O₂ treatment. Blood samples were treated with 50μM H₂O₂. (F) ROS levels in RBCs of Hri^+/+ (wt) and Hri^−/− (Ko) mice in iron deficiency.