Figure 2.

ATM and Chk1/2 inhibition blocks MAP kinase activation, uric acid production, and MICA/B expression. HeLa cells grown in 6-well plates were incubated for 24 hr in the absence or presence of 5-FU (10 μM) or gemcitabine (2 μM) minus or plus KU-55933 (5 μM) or DBH (10 μM). Cell lysates were prepared and analyzed for TAK1 and ERK phosphorylation (a) by Western blot and uric acid levels by using a uric acid assay kit (b). Relative phosphorylation levels were analyzed by quantifying the density of phosphorylated TAK1 and ERK bands normalized by the density of their corresponding total protein bands with NIH Image-J software and presented as bar graphs. Data in (a) and (b) are the mean ± SD of three experiments. **p < .01, compared to the untreated control. ^#p < .05, ^##p < .01; ns, not significant, compared to their corresponding no-inhibitor controls. For assaying the levels of MICA/B, HeLa cells were treated as above and incubated for 24 hr, single-cell suspensions were stained with a PE-conjugated anti-MICA/B antibody (c). The levels of MICA/B expression were analyzed by flow cytometry. Green line, isotype control; Red line, MICA/B. The data represent one of three independent experiments with similar results.