Figure 4

CycT effectively inhibits the levels of proteins and enzymes involved in heme uptake, synthesis, and transport. (A) Representative IHC images of H1299 NSCLC tumor tissue sections and graph showing the levels of HRG1 in control and treated tumors. Shown are montages and 10X images of control, CycT-treated, and Bev-treated tumor tissue sections stained with DAPI or antibodies against the indicated protein HRG1. The light blue lines in DAPI images outline the tumors in the lung. The white rectangles in DAPI images denote the regions shown in 10X images. The lines and rectangles are not placed on IHC fluorescent images to avoid the obstruction of tumor images. The heart was often stained and is marked with “H”. Scale bar: montage, 1 mm; 10X, 20 μm. Protein levels were quantified with cellSens dimension software (Olympus), as described in Methods. Data are plotted as mean ± SEM. For statistical analysis, the levels in treated tumors were compared to the levels in control tumors with a Welch 2-sample t-test. **p-value < 0.005. (B) The effects of CycT and Bev on the levels of the rate-limiting heme biosynthetic enzyme ALAS1 in orthotopic tumor xenografts. (C) The effects of CycT and Bev on the levels of the heme chaperone and sensor protein PGRMC1 in orthotopic tumor xenografts. IHC images are representative of 3 independent experiments.