Figure 5.

TcdB does not directly intoxicate thymocytes. Thymocytes and splenocytes (1x10⁶) isolated from three-month-old male C57BL/6 J mice and Vero cells (1x10⁴) were incubated with dilutions of recombinant C. difficile TcdB ranging from 80 ng/ml to 0.128 ng/ml, using media alone (negative), or 10% DMSO (positive) or 10% oligomycin (positive) as controls for thymocyte death. a) Thymocyte viability after TcdB incubation. b) Vero cell detachment as a measure of cell rounding after TcdB intoxication. c) Representative images of thymocytes and Vero cells following incubation with TcdB. Scale bar = 50 µm. Percentage of live thymic (d) CD4 T cells and (e) CD8 T cells and splenic (f) CD4 T cells and (g) CD8 T cells following incubation with TcdB; as measured by exclusion of both LIVE/DEAD™ Fixable Aqua (Invitrogen) and Propidium Iodide (BD Biosciences) stains. n = 4. Error bars indicate S.E.M. Statistical analysis by Mann Whitney U test. * = p ≤ .05.