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. 2012 Aug;32(15):3176–3186. doi: 10.1128/MCB.00086-12

Fig 1.

Fig 1

Abl family kinases are required for optimal phagocytosis. (A and B) Imatinib decreases C3bi- and IgG-opsonized bead uptake. BM Mϕs or RAW 264.7 cells were treated with 3.3 μM imatinib or DMSO for 2 h. Ten C3bi- or IgG-opsonized beads were incubated per Mϕ for 30 min at 37°C. Two-color IF distinguished between intracellular (green) and extracellular (orange) beads. Nuclei are labeled with DAPI. (A) Image of C3bi-opsonized bead uptake by BM Mϕs treated with DMSO (top) or imatinib (bottom). The left panels show representative fields; the right panels show enlarged areas (boxed). Left scale bar, 20 μm; right scale bar, 5 μm. (B) Imatinib inhibits phagocytosis. Bars show the mean phagocytic index (PI) ± standard error (SE) for RAW 264.7 cells treated with 3.3 μM imatinib normalized to the DMSO-treated PI (100%) for each experiment. **, P = 0.0063for C3bi-coated bead uptake by DMSO versus imatinib-treated cells, and P = 0.0093 for IgG-coated bead uptake by DMSO versus imatinib-treated cells, by one-sample t test (n = 3 experiments). (C) Imatinib does not affect adhesion. Bars show percentages of adhered beads per 100 imatinib-treated RAW 264.7 cells compared to DMSO-treated cells from the experiment shown in panel B. (D) The relative decrease in PI in imatinib-treated Mϕs does not change even after 3 h of incubation. Mϕs were treated with DMSO or 3.3 μM imatinib prior to incubation with zymosan particles for the time indicated. The PI is shown for each time point of imatinib-treated Mϕs, normalized to DMSO-treated Mϕs at the first time point. Shown is one representative experiment of two experiments. (E) Immunoblots of Arg and Abl expression in BM Mϕs isolated from WT, arg−/−, abl−/−, or arg−/− ablflox/flox/Tie2-Cre+ (dKO) mice. dKO Mϕs express low levels of a truncated, kinase-inactive form of Abl (indicated by arrow) that migrates 10 kDa faster than full-length Abl on immunoblots. Actin is a loading control. (F) Mϕs lacking Arg with decreased Abl levels (dKO) have phagocytic defects. The graph shows the normalized mean PI ± SE for dKO Mϕs compared to WTLM Mϕs (abbreviated “WT”). *, P = 0.035 for C3bi-coated bead uptake by WTLM versus dKO Mϕs, and P = 0.019 for IgG-coated bead uptake by WTLM versus dKO Mϕs, by one-sample t test.(n = 3 experiments)