Extruding epithelial cells show redistribution of some tight junction proteins but lack markers of apoptosis.
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(A–C) Immunofluorescence micrographs of small intestinal sections from mock and infected rabbits stained for (A) ZO-1, (B) occludin-1 and (C) claudin-1. Note altered localization of ZO-1 and occludin-1 but not claudin-1 in extruding epithelial cells with adjacent V. parahaemolyticus (arrowheads). Actin cytoskeleton was stained with phalloidin-Alexa 568 (yellow), bacteria constitutively expressed green fluorescent protein (green) and nuclei were stained with DAPI (blue). (D) Tissue sections from infected rabbits were stained for apoptotic cells using the TUNEL assay. TUNEL-positive cells were detected using a fluorescein-based detection system thus both bacteria and apoptotic host cells appear green, but can be distinguished based on their relative size and shape. Examples of TUNEL-positive (arrowhead) and TUNEL-negative (arrows) host cells are indicated. Sections were counterstained with phalloidin-Alexa 568 (yellow) and DAPI (blue) as described above.