Article Figures & Data
Figures
- Figure 1.
Induction of HNSCC cell death by proteasome inhibitors. UM-22A, UM-22B, and 1483 cells were left untreated or were treated for 48 h with 0.1% DMSO or varying concentrations of MG-132 (A) or bortezomib (B). Following treatment, MTS assays were done. Points, average of triplicate wells; bars, SD. The experiments were done twice, with similar results each time. C, UM-22A, UM-22B, and 1483 cells were left untreated or were treated for 24 or 48 h with concentrations of bortezomib corresponding to the IC50 of each cell line (20 nmol/L for UM-22A and UM-22B and 80 nmol/L for 1483). As a control, cells were treated with 0.1% DMSO for 24 h. Whole-cell lysates (40 μg/lane) were subjected to immunoblotting with anti-PARP or anti-caspase-3. The processed, active forms of caspase-3 are depicted. Blots were stripped and reprobed with anti-β-actin to show equal protein loading.
- Figure 2.
Bortezomib regulates the expression of proapoptotic and antiapoptotic Bcl-2 family members in HNSCC cells. UM-22A and 1483 cells were left untreated or were treated for 24 or 48 h with bortezomib (UM-22A: 20 nmol/L; 1483: 80 nmol/L). Control cells were treated for 24 h with 0.1% DMSO. Whole-cell lysates (40 μg/lane) were subjected to immunoblotting for the indicated proteins. Probing with anti-β-actin was used to show equal protein loading. Similar results were obtained in three independent experiments.
- Figure 3.
Bik and Bim mediate bortezomib-induced HNSCC cell death. A, UM-22A cells were transfected with Bik siRNA or a nonspecific siRNA as described in Materials and Methods. After 24 h, the transfected cells were either left untreated or were treated for 48 h with 20 nmol/L bortezomib. Cell viabilities were determined by trypan blue exclusion assays, and Bik expression levels were determined by immunoblotting. Columns, average of means obtained in three independent experiments; bars, SE. *, P = 0.029, compared with bortezomib-treated cells transfected with nonspecific siRNA versus Bik siRNA. B, UM-22A cells were transfected with Bim siRNA or a nonspecific siRNA, then treated, and analyzed as in A. Columns, average of means from three independent experiments; bars, SE. *, P = 0.01, compared with bortezomib-treated cells transfected with nonspecific siRNA versus Bim siRNA.
- Figure 4.
Inhibition of Mcl-1L up-regulation enhances bortezomib-induced death of HNSCC cells. UM-22A cells were transfected with Mcl-1 siRNA or nonspecific siRNA followed by 48 h of treatment with 20 nmol/L bortezomib. Trypan blue exclusion assays were used to determine cell viabilities, and immunoblotting was used to assess siRNA-mediated inhibition of Mcl-1L up-regulation. Columns, average of means from three independent experiments; bars, SE. *, P = 0.004, compared with bortezomib-treated cells transfected with nonspecific siRNA versus Mcl-1 siRNA.
- Figure 5.
Bortezomib synergizes with cisplatin in the killing of HNSCC cells. UM-22A cells (A) and 1483 cells (B) were treated for 24 h with bortezomib alone, cisplatin alone, or a constant ratio of bortezomib plus cisplatin. Following treatment, MTS assays were done. Points, average of triplicate wells; bars, SD. CI was determined as described in Materials and Methods. Similar results were obtained in three independent experiments. C, UM-22A and 1483 cells were left untreated or were treated for 24 h with 0.1% DMSO alone, 40 nmol/L bortezomib alone, 8 μmol/L cisplatin alone, or the combination of 40 nmol/L bortezomib plus 8 μmol/L cisplatin. Whole-cell lysates were analyzed by immunoblotting for cleavage of PARP protein and processing/activation of caspase-3. The blot was stripped and reprobed with anti-β-actin to show equal protein loading. The experiment was done twice, with similar results each time.
Additional Files
Supplementary Material, Li et al
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Volume 7, Issue 6
