Effects of E. coli O157:H7 on human colonic Epithelial cells lines and translocation of Shiga Toxin.

GARIMANO, NICOLAS; ALBANESE, ADRIANA; AMARAL, MARIA M; IBARRA, CRISTINA

Mar del Plata

Congreso; LXI Reunión Científica de la Sociedad Argentina de Investigación Clínica (SAIC). LXIV Reunión Anual de la Sociedad Argentina de Inmunología (SAI). XLVIII Reunión Anual de la Sociedad Argentina de Farmacología Experimental (SAFE). VII Reunión Anual de la S; 2016

SAIC-SAFIS.

Shiga toxin (Stx) producing Escherichia coli STEC strains are responsible of bloody diarrhea hemorrhagic colitis and hemolytic uremic syndrome (HUS). STEC is by far, the most prevalent serotype associated with HUS and Stx2 is the major virulence factor associated for the more severe symptoms of the infection. After passage through the acidic barrier, STEC colonizes the human colon promoting production and absorption of Stx2. However, the mechanisms involved in the pathogenesis of diarrhea mediated by Stx2 are not well known yet. Our aim as to study the cytotoxic effects of STECH on human colonic epithelial cells in order to better understand the means by which Stx2 induces diarrhea and translocate the intestinal barrier. In this study, the examined HCT-8 and Caco viability after incubation􀁙with purified Stx2, STEC O157:H7 strain 125/99(125/99wt) a mutant of strain lacking stx2 gene (125/99Δstx2) and the filtered 125/99wt supernatants. Cells were growing in 96-well culture plate and viability as measured by neutral red uptake after a incubation period under growth arrested condition. We have also evaluated the translocation of purified Stx2 across HCT-8 cultured as monolayers on Millicell cell culture inserts in the presence of (125/99Δstx2).Transepithelial electric resistance was monitored daily during the development of cell culture until confluence 􀁙as achieved, and after Stx2 treatment to ensure monolayer integrity. Cytotoxicity of collected basal media on Vero cells assays asenhanced by bacterial presence on the luminal side. Furthermore, the cytotoxic effects induced by strains on HCT-8 and Caco cell lines were significantly higher than those observed with purified Stx2 and filtered 125/99wt tsupernatants. These results indicate the importance of bacterial cells in the interaction and translocation of Stx through the intestinal barrier to cause HUS.