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. Author manuscript; available in PMC: 2013 May 15.
Published in final edited form as: Biochem J. 2013 May 15;452(1):111–119. doi: 10.1042/BJ20130147

Figure 2. ABT-263 induced ceramide accumulation and CerS activity can be inhibited by MCL1 in U937 cells.

Figure 2

A, U937 cells stably expressing GFP and a portion of an irrelevant protein, a portion of ER (U937-ER) were treated with 1.6 μM of ABT-263, or vehicle (DMSO), for 8 and 24 hours and the total amount of cellular ceramides were determined by HPLC-MS/MS. B, U937 cells stably expressing GFP and BCLxL (U937-BCLxL) were treated with 1.6 μM of ABT-263, or vehicle (DMSO), for 8 and 24 hours and the total amount of cellular ceramides were determined by HPLC-MS/MS. C, U937 cells stably expressing GFP and MCL1 (U937-MCL1) were treated with 1.6 μM of ABT-263, or vehicle (DMSO), for 8 and 24 hours and the total amount of cellular ceramides were determined by HPLC-MS/MS. D. U937-MIG ER cells (U937 cells stably infected with MIG-ER virus) were incubated with 1.6 μM of ABT-263, or vehicle (DMSO), for 1 hour and 45 minutes. 1μM C17-sphingosine was then added to the media for an additional 15 minutes. Cells were immediately lysed and the amount of C17-ceramide in the cells were quantitated by HPLC-MS/MS. E, U937-BCLxL cells were incubated with 1.6 μM of ABT-263, or vehicle (DMSO), for 1 hour and 45 minutes. 1μM of C17-sphingosine was then added to the media for an additional 15 minutes. Cells were immediately lysed and the amount of C17-ceramide in the cells were quantitated by HPLC-MS/MS. F, U937-MCL1 cells were incubated with 1.6 μM of ABT-263, or vehicle (DMSO), for 1 hour and 45 minutes. 1μM of C17-sphingosine was then added to the media for an additional 15 minutes. Cells were immediately lysed and the amount of C17-ceramide in the cells were quantitated by HPLC-MS/MS. Lipids were normalized to total lipid phosphate and data expressed as a fold-change of the untreated control. All data points represent the compilation of at least biological triplicates. ***, p=<0.0005; **, p=<0.005; *, p=<0.05.