Figure 6.

Human NK-LGL and T-LGL leukemias with reduced Bid expression are susceptible to bortezomib-induced cell death. A, PBMCs from one healthy, four NK-LGL, and six T-LGL leukemia donors were lysed in Triton X-100. SDS-PAGE fractionated protein extracts were analyzed by Western blot using a rabbit polyclonal anti-Bid antibody. Blots were reprobed with goat anti-actin. B, PBMCs from three healthy, one NK-LGL, and four T-LGL leukemia donors were isolated and treated with the indicated bortezomib concentrations for 16 or 24 h. Protein extracts were analyzed by Western blot as described above. C, PBMCs from healthy and LGL leukemia donors were treated with bortezomib at the indicated concentrations and times. Apoptosis was evaluated by Annexin V-FITC and 7AAD staining (left). Samples assays were in triplicate. P < 0.05, statistical significance determined using the Student's t test. Leukemia cells from T-LGL donors 9 and 10 were transduced with Bid expression constructs, pD4-GFP-Bid, TA-GFP-Bid, and a GFP-only construct, pd4EGFP (right). Apoptosis was analyzed 24 h later by Annexin V-PE and 7AAD staining. All samples were assayed in triplicate.