Fig. 7.

LPI protects against Shiga toxin by reducing toxin binding to cells. a HEp-2 cells were treated with 10 µM LPI or 0.05 % (v/v) ethanol (control) for 30 min before increasing concentrations of Shiga toxin were added. Protein synthesis was measured 3 h later by quantifying the incorporation of [³H]leucine. One representative experiment out of four is shown. b Shiga toxin binding to HEp-2 cells at 37 °C was determined by quantification of total cell-associated ¹²⁵I-labeled Stx1-mut. The cells were pretreated with 10 µM LPI or 0.05 % (v/v) ethanol for 30 min at 37 °C, and then incubated with 10 ng/ml toxin in the presence of LPI or ethanol for 20 min before removal of the medium, three times wash in PBS, and measurement of toxin associated with the cells. The figure shows the mean and deviation of two experiments. c HEp-2 cells were pretreated with LPI or ethanol (control) as described above, before continuous incubation with Stx1-mut in the presence of LPI or ethanol at 37 °C for 45 min. Cells were fixed, stained with an anti-Shiga toxin antibody (red) and appropriate secondary antibodies, mounted in the presence of DAPI (blue), and analyzed by immunofluorescence microscopy. d Total cell-associated Shiga toxin was determined by quantifying the immunofluorescence signal in control cells and LPI-treated cells, and is expressed as % of control. The figure shows the quantification for approximately 1,500 cells of each condition