Figure 2

Monocyte fusion is stimulated by PR3. Monocytes stimulated with PR3 were first imaged by Giemsa staining. Distinct cellular aggregates formed in the presence of PR3 (A). At higher magnification (B), a larger cellular aggregate is present showing dense nuclear staining and cytoplasmic fusion (scale bars 100 µm). Three-dimensional volume rendering of an aggregate, including all acquired slices by confocal microscopy, was undertaken using cell membrane (red), cytoskeletal (green) and nuclear staining (blue) (C, D). Midslice (D) confirmed membrane fusion, cytoskeletal rearrangement and a single cellular structure with fusion of three monocytes, defining an MGC. Scanning electron microscopy highlighted monocyte fusion, with representative images showing an MGC size of 48.8 µm (E) and 34.6 µm (F). Membrane fusion was measured by profile plot of the fluorescence intensity of the three channels over the x/y diameter (G). There was prominent membrane staining encasing cytoskeleton and three nuclei. MGC, multinucleated giant cells; PR3, proteinase 3.