Figure 7. Inhibition of NF-03BA;B in DLBCL cells leads to cell growth inhibition, G0/G1 cell cycle arrest, and apoptosis.

(A) Representative ABC- and GCB-DLBCL cell lines were treated with bortezomib (BZ) or BAY-11 for 48 hours and cell proliferation was measured using 3H-thymidine incorporation assays. The percentages of growth inhibition of treated cells relative to untreated (control cells) were plotted. The data shown are the means and ranges of triplicate cultures from three independent experiments. (B) DLBCL-MS cells were cultured in the absence or presence of bortezomib (BZ) or BAY-11 and subjected to a 20S proteasome assay. Purified 20S proteasome was used as a positive control. Abbreviations: RLU, relative light unit; 20S pro, 20S proteasome, Neg Cont., negative control. (C) DLBCL-MS cells were cultured in the absence or presence of BZ (50 nM) or BAY-11 (1 μM) and analyzed for cell cycle profile. The percentages of cells in G0/G1, S, and G2M phases are shown. (D) DLBCL-MS cells were cultured in the absence or presence of BZ (50 nM) or BAY-11 (1 μM) for the indicated time points and then analyzed for apoptosis using annexin V assays. (E) DLBCL-MS cells were cultured in the presence of BZ (50 nM) or BAY-11 (1 μM) and in some cases with the caspase 3 inhibitor DEVD or the caspse 1 inhibitor VAD. Caspase 3 activity was measured after 24 hours of treatments. Caspase 3 activity was observed after 12 hours of treatment. Abbreviations: RLU, relative light units. (F) DLBCL-MS cells were cultured in the presence of BZ (50 nM) or BAY-11 (1 μM) for the indicated time points and cell extracts were subjected to Western blotting for a known caspase substrate, poly-(ADP-ribose) polymerase (PARP) cleavage.