EFFECT OF CLOSTRIDIUM-DIFFICILE TOXIN-A ON HUMAN COLONIC LAMINA PROPRIA CELLS - EARLY LOSS OF MACROPHAGES FOLLOWED BY T-CELL APOPTOSIS

We have previously shown that Clostridium difficile toxin A induces de tachment of human colonic epithelial cells from the basement membrane and subsequent cell death by apoptosis. Because these cells require ad hesion-dependent signalling from the extracellular matrix for survival , their detachment from the basement membrane by other means also indu ces apoptosis. The role of toxin A in the induction of apoptosis there fore remains to be determined. In addition, sensitivities to C. diffic ile toxin A of lamina propria lymphocytes, macrophages, and eosinophil s, which lie below the surface epithelium, are not known. In contrast to epithelial cells, these lamina propria cells do not require adhesio n-dependent signalling from the extracellular matrix for survival, and this may allow the mechanisms of toxin A-induced cell death to be fur ther investigated. The aim of this study was to investigate the effect of purified C. difficile toxin A on human colonic lamina propria T ce lls, macrophages, and eosinophils. We show that C. difficile toxin A i nduces loss of viability in isolated colonic lamina propria cell prepa rations containing the three different cell types in a dose- and time- dependent fashion. Exposure to high concentrations of the toxin led to loss of macrophages within 72 h. T-lymphocyte and eosinophil cell dea th was prominent at later time points and occurred by apoptosis. Expos ure to toxin A also induced the production of tumor necrosis factor al pha by the isolated colonic lamina propria cells. However, the presenc e of neutralizing antibodies to this cytokine did not influence C. dif ficile toxin A-induced T-cell apoptosis. Moreover, Purified T cells al so underwent apoptosis following exposure to toxin A, implying that ap optosis occurred as a consequence of a direct interaction between T ce lls and the toxin. Our studies suggest that C. difficile toxin A is ca pable of suppressing human colonic mucosal immune responses by inducin g early loss of macrophages followed by T-cell apoptosis.