Fig. 4. Ibrutinib switches MDSCs to mature dendritic cells.

Single-cell suspensions were prepared from both the spleens and tumours of both groups of mice and analysed for mature DCs (Ly6C^low MHCII^hi) and immature DCs (Ly6C^hi MHCII^low) cells. a Representative gating strategy followed to analyse the mature and immature DCs from total CD11C gated live splenocytes. Bar graphs represent the percentage of b mature DCs and c immature DCs from total gated CD11C+ live cells from the spleens and tumours of vehicle- and ibrutinib-treated mice. d, e Total MDSCs were sorted from the spleens of WT tumour-implanted mice and treated with DMSO or ibrutinib (1 µM) for 1 h. Cells were washed, stained with CD11C and MHCII, and analysed for DC maturation by flow cytometry. d Gating strategy followed to identify the mature DCs (CD11C+MHCII+) from the total DMSO/ibrutinib-treated MDSCs. e The bar graph represents the percentage of matured DCs from total DMSO/ibrutinib-treated MDSCs. Data are representative of 1 of the 3 independent experiments, with 10–12 mice/group. Error bars represent means ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.0001, by unpaired t test.