Fig. 6
Knockout of Gstz1 promotes DEN/CCl4-induced hepatocarcinogenesis in vivo via activation of the KEAP1/NRF2 pathway. a Schematic representations of the experimental design for WT and Gstz1−/− mice. b Gross appearances of murine livers. Red arrows indicate tumors. c Liver/body weight ratio (left) and number of tumors (right) in each group. d Representative H&E staining and immunohistochemistry staining of Ki67 in liver tumors. e-f Oxidative stress levels in liver tumors. e Representative fluorescence staining of ROS with CellROX Orange probe in hepatic tumors of WT and Gstz1/− mice (left). Intracellular ROS quantification (right). f 4-HNE modification is all proteins in tumor tissues, analyzed via Western blotting. g NRF2 transcriptional activities in liver tumors. Relative Nqo1 mRNA expression, determined via qRT-PCR (Left). NQO1 expression in total cell extracts, and NRF2 expression in cytoplasmic and nuclear extracts, analyzed via Western blotting (Right). h Representative immunohistochemistry images of GSTZ1 and NQO1 in hepatic tumors. For Western blotting, 30 μg protein was loaded per well. Values represent the mean ± SD (n = 3, performed in triplicate), *p < 0.05, **p < 0.01, Student’s t-test (two groups) or one-way ANOVA followed by Tukey tests (four groups). H&E, hematoxylin and eosin; DEN, diethylnitrosamine; HCC, hepatocellular carcinoma; ROS, reactive oxygen species; 4-HNE, 4-hydroxy-2-nonenal; qRT-PCR, quantitative reverse transcription polymerase chain reaction