Figure 5.
BCL-XL, IKAROS, and CK2 regulate sensitivity to doxorubicin in B-ALL cells. (A) B-ALL cells, retrovirally transduced with BCL2L1 (MIG-BCL2L1) or control vector (MIG-CTL) underwent fluorescence-activated cell sorting (FACS) with the same gate setting of GFP expression, and treated for 3 days with indicated doses of doxorubicin. Cell proliferation was assessed by WST-1 assay. (B) B-ALL cells, retrovirally transduced with IKZF1 or a control vector, were stained with 7-AAD and Annexin V for flow cytometry to determine apoptosis 3 days following retroviral transductions. Gated GFP+ cells were analyzed for apoptosis by flow cytometry. The percentage of cells in the lower right quadrant and upper right quadrant of each flow chart represents the percentage of early apoptotic or late apoptotic cells, respectively, in samples treated with the indicated drugs. (C) B-ALL cells, retrovirally transduced with IKZF1 or a control vector, were FACS-sorted with the same gate setting of GFP expression and treated for 3 days with indicated doses of doxorubicin and assayed using WST-1 cell proliferation assay. (D) B-ALL cells were retroviral transduced with IKZF1 or with control vector and treated with 10 nM doxorubicin (DOX) then evaluated for apoptosis as in panel B. (E) B-ALL cells, transduced with lentiviral IKZF1 shRNA (shIK) or scramble shRNA control (shCTL), underwent FACS with the same gate setting of GFP expression and treated for 3 days with indicated doses of doxorubicin. B-ALL cells with (F) retroviral CK2α overexpression (MIG-CK2α) or vector only control (MIG-CTL); or (G) lentiviral CK2α shRNA or scramble shRNA control (shCTL), underwent FACS based on GFP expression as described previously. FACS cells were treated with indicated doses of doxorubicin for 3 days then evaluated by WST-1 proliferation assay. (H) B-ALL cells were treated with 10 μM CX-4945 for 3 days and stained with 7-AAD and Annexin V for flow cytometry to determine apoptosis. (B, D, H) Flow cytometry plots depict a representative experiment and percentages are the mean ± SD of 3 independent experiments. (A, C, E, F, G) Mean ± SD of triplicates representative of 1 of 3 independent experiments. *P < .05, **P < .01, ***P < .001, ****P < .0001.

BCL-XL, IKAROS, and CK2 regulate sensitivity to doxorubicin in B-ALL cells. (A) B-ALL cells, retrovirally transduced with BCL2L1 (MIG-BCL2L1) or control vector (MIG-CTL) underwent fluorescence-activated cell sorting (FACS) with the same gate setting of GFP expression, and treated for 3 days with indicated doses of doxorubicin. Cell proliferation was assessed by WST-1 assay. (B) B-ALL cells, retrovirally transduced with IKZF1 or a control vector, were stained with 7-AAD and Annexin V for flow cytometry to determine apoptosis 3 days following retroviral transductions. Gated GFP+ cells were analyzed for apoptosis by flow cytometry. The percentage of cells in the lower right quadrant and upper right quadrant of each flow chart represents the percentage of early apoptotic or late apoptotic cells, respectively, in samples treated with the indicated drugs. (C) B-ALL cells, retrovirally transduced with IKZF1 or a control vector, were FACS-sorted with the same gate setting of GFP expression and treated for 3 days with indicated doses of doxorubicin and assayed using WST-1 cell proliferation assay. (D) B-ALL cells were retroviral transduced with IKZF1 or with control vector and treated with 10 nM doxorubicin (DOX) then evaluated for apoptosis as in panel B. (E) B-ALL cells, transduced with lentiviral IKZF1 shRNA (shIK) or scramble shRNA control (shCTL), underwent FACS with the same gate setting of GFP expression and treated for 3 days with indicated doses of doxorubicin. B-ALL cells with (F) retroviral CK2α overexpression (MIG-CK2α) or vector only control (MIG-CTL); or (G) lentiviral CK2α shRNA or scramble shRNA control (shCTL), underwent FACS based on GFP expression as described previously. FACS cells were treated with indicated doses of doxorubicin for 3 days then evaluated by WST-1 proliferation assay. (H) B-ALL cells were treated with 10 μM CX-4945 for 3 days and stained with 7-AAD and Annexin V for flow cytometry to determine apoptosis. (B, D, H) Flow cytometry plots depict a representative experiment and percentages are the mean ± SD of 3 independent experiments. (A, C, E, F, G) Mean ± SD of triplicates representative of 1 of 3 independent experiments. *P < .05, **P < .01, ***P < .001, ****P < .0001.

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