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Typhimurium can reach the lamina propria by transcytosis, i.e., invasion at the apical side of epithelial cells, migration to the basolateral side, and exocytosis into the interstitial space of the lamina propria. During intravital microscopy of S. Typhimurium-infected pvilcre �� dsRedfloxSTOP or pvilcre �� tdRFPfloxSTOP mice, we observed infected and noninfected epithelial cells releasing RFP+ material at the basolateral side into the lamina propria ( Figure?6A). These dsRed+ or tdRFP+ ��epithelial bodies�� grew over minutes to hours, reached sizes of up to several micrometers in diameter, and detached from the overlying epithelium. Immunofluorescence microscopy using an epithelium-specific ��-EpCAM (CD326) antibody verified their epithelial Sotrastaurin price origin ( Figure?6B). It seemed plausible that such epithelial bodies might serve the presentation of enterocyte material to the gut immune system, as earlier work had described such a mechanism in the rat intestine ( Huang et?al., 2000). Due to the potential importance of such a mechanism, we decided to follow up on this initial observation and investigated potential links to the mucosal S. Typhimurium infection. First, we performed high-resolution fluorescence microscopy of fixed tissue sections from S. Typhimurium (pssaG-GFP)-infected pvilcre �� dsRedfloxSTOP mice ( Figure?6C; 12?hr p.i.). Staining of the lamina propria cell surface marker ICAM-1 allowed colocalization analysis of S. Typhimurium (pssaG-GFP; green) and RFP+ epithelial bodies within cells of the lamina propria. Strikingly, numerous Salmonella-containing Mdm2 lamina propria cells harbored at the same time RFP+ epithelial bodies ( Figures 6C�C6E). To control for background Cyclopamine fluorescence, we took advantage of the mosaic-like expression pattern of the dsRed in pvilcre �� dsRedfloxSTOP mice. No RFP+ epithelial bodies were detected in infected (and noninfected) lamina propria cells?underneath RFP? areas of the epithelium ( Figure?6D). Furthermore, quantitative colocalization analysis of green (S.?Typhimurium pssaG-GFP) and red (epithelial-derived dsRed) fluorescence verified that approximately half of the bacteria in the lamina propria were clearly colocalized with epithelial cell material (p?
