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. Author manuscript; available in PMC: 2019 Apr 1.
Published in final edited form as: Leukemia. 2017 Oct 3;32(4):960–970. doi: 10.1038/leu.2017.304

Figure 4. IL-10 production in CLL cells is reduced after ibrutinib therapy.

Figure 4

Figure 4

Figure 4

(a) PBMCs from CLL vs. healthy donors were stimulated with CpG (4 μg/ml) plus CD40L (100 ng/ml) for 12 hours followed by PMA, ionomycin and brefeldin A for the last 6 hours of culture and then stained for CD19 and intracellular IL-10 production. Frequencies of IL-10+CD19+ B cells are greater in CLL patients compared to healthy donors (n=6 each) are shown. (b) PBMCs from patients given ibrutinib therapy were stimulated with CpG plus CD40L, followed by analysis of IL-10 secretion as described above (n=7). Representative FACS plots are shown. (c) PBMCs from treatment-naïve CLL patients were cultured with ibrutinib (1 μM) or cucurbitacin (0.05 μM) overnight, and their IL-10 response to stimulation with CpG and CD40L was measured by intracellular cytokine staining (n=6). (d) CLL cells from treatment-naïve patients were treated with a GFP lentiviral hSTAT3 sh-RNA vector or a mock GFP lentiviral vector as described in methods. Cells were then stimulated with 4 μM CpG and CD40L (100 ng/ml), followed by analysis for IL-10 production. IL-10 production was compared in gated CD19+GFP+ cells in transfected vs. mock-transfected CLL cells (n=3). Representative FACS plots are presented.