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Figure 4 | Molecular Medicine

Figure 4

From: Inhibition of High-Mobility Group Box 1 Protein (HMGB1) Enhances Bacterial Clearance and Protects against Pseudomonas Aeruginosa Pneumonia in Cystic Fibrosis

Figure 4

HMGB1 is sufficient to impair macrophage phagocytotic function. C57BL/6 mice (A, B) were treated with 50 µg/mouse of either anti-HMGB1 mAb (αHMGB1) or isotype control mAb (control), 12 h before the inoculation with GFP-expressing P aeruginosa PAO1 and FITC-labeled latex beads. Four hours after inoculation, mice were sacrificed and BAL harvested. (A) PMN and monocytes/macrophages in BAL were quantified. (B) Alveolar macrophages/monocytes in BAL were plated on chamber slides and stained with Texas Red X-phalloidin. Phagocytic activity of macrophages was quantified by counting 50 cells/slide and no less than six slides from three independent experiments. Scale bar, 10 µm. (C, D) RAW 264.7 macrophages were treated for 24 h with either the indicated concentrations (C) or 10 ng/mL (D) of either recombinant HMGB1 (rHMGB1) or GST peptide tag control (control) or trypsinized rHMGB1 (trHMGB1) and then exposed for 1 h to either FITC-labeled latex beads (C) or heat-killed GFP-expressing PAO1 (D). Macrophages/monocytes in the fields were stained with phalloidin and internalized PAO1 or beads were counted and analyzed (B–D). Data represent mean ± SEM of at least three independent experiments. *P < 0.05 (t test) and #P < 0.001 (ANOVA), compared with the groups that were treated with either GST control or trypsinized rHMGB1. Mac, macrophages.

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