Figure 4

Paracellular polymorphonuclear neutrophils (PMN) traversal across human choroid plexus papilloma cells (HIBCPP). HIBCPPs were stained for the tight junction (TJ) protein zonula occludens (ZO)-1 (red), actin (phalloidin, yellow), cell nuclei (4‘-6-diamidino-2-phenylindole dihydrochloride (DAPI), blue), and additionally we stained for the PMN marker (CD66b-fluorescein isothiocyanate (FITC), green). (A-E) Three-dimensional immunofluorescence images of HIBCPP and transmigrating PMNs reconstructed from 0.3 μm Apotome® optical sections, using Zeiss software Inside 4D. The views from above and the side views show paracellular transmigration, where PMNs are migrating through the intercellular spaces. Images show the transmigration of PMNs through HIBCPP after previous stimulation with strain MC58 (A), strain MC58ΔsiaD (B), strain α14 (C), TNFα (D), and control conditions (E). The figures show representative examples of three independent experiments that all gave similar results. Scale bar as indicated.