Figure 2

Purified gingipains cleave CD14 from the surface of macrophages and NØ. (a) Human primary NØ or THP-1 cell-derived MØ were treated with the indicated purified gingipains for 1 h at 37 °C prior to indirect immunofluorescence staining for receptors/ligands implicated in apoptotic cell clearance. Fluorescence levels are shown as percentage of cells positive for antigen (left) and mean fluorescence intensity (right). (b) THP-1 cell-derived MØ were treated with Rgp for up to 12 h at 37 °C prior to indirect immunofluorescence staining for CD14 at the indicated times. Fluorescence levels are shown as percentage of cells positive for antigen and mean fluorescence intensity. (c) MØ were treated with Kgp for 1 h in the presence or absence of the inhibitor TLCK prior to assessment of CD14 expression via flow cytometry. (d) Representative flow cytometric analyses of THP-1-derived MØ or NØ treated with RgpB or Kgp for 1 h at 37 °C and stained with annexin V and propidium iodide to reveal cell viability. Flow data shown are collected from at least 5000 events. Data presented are mean±S.E.M. of at least three independent experiments. Statistical analysis was performed using ANOVA followed by a Bonferroni post hoc-test. **P<0.01; ***P<0.0001 compared with untreated cells