Trans-epithelial transport of extracellular vesicles produced by probiotic bacteria Bacillus subtilis 168 across intestinal epithelial cells

DOMINGUEZA RUBIO, AP; PIURI, M; MARTÍNEZ, JH; PÉREZ, OE; PALAVECINO, M

Buenos Aires

Simposio; 3rd Frontiers in Physical Sciences; 2018

MERCOSUR-FOCEM

Bacillus subtilis 168 is a regular resident of the mammalian gastrointestinal (GI) microbiome. Extracellular vesicles (EVs) have been proposed to be involved in signaling between probiotic bacteria and their mammalian hosts. B. subtilis 168 produces EVs (50?300 nm). We hypothesize that transcytosis of EVs across intestinal epithelial cells is a crucial step in the host-probiotic communication. To test this, the ability of EVs produced by the probiotic strain B. subtilis 168 to cross intestinal epithelial cell barrier was investigated in an in vitro model of human Caco-2 cells. B. subtilis 168 was grown in BHI medium at 37°C under agitation for 18 h. Cells were removed from the culture by ultracentrifugation (Domínguez Rubio et al., 2017). Human colon carcinoma Caco-2 cells were differentiated for 14 days (100% confluence). Isolated EVs were stained with CFSE. EVs uptake was analyzed as the number of EVs labeled inside the cell by Confocal Laser Scanning Microscopy (CLSM). Transcytosis was studied as the fluorescence measured in the collected medium from the transwell lower chamber and EVs were analyzed. Intact EVs uptake in Caco-2 cells was linear for up to 30 min: y=1.02x-1.25 and R²=0.97 (p0.05).EVs produced by the probiotic strain B. subtilis 168 crossed intestinal epithelial cell barrier of human Caco-2 cells. This evidence suggests that EVs could play a key role in signaling between GI bacteria and mammalian hosts. The expression and further encapsulation of proteins into EVs of GRAS bacteria could represent a scientific novelty, with applications in food and clinical therapies.