Clostridium difficile toxin: Cytoskeletal changes and lactate dehydrogenase release in hepatocytes

Background. We have found that Clostridium difficile toxins can evoke hepat ocyte acute-phase protein synthesis, and that this effect is dependent on a functioning interleukin-1 (IL-1) receptor. The present study was undertake n to determine if C. difficile toxicity, as determined by actin rearrangeme nt and lactate dehydrogenase (LDH) release, also requires a functioning IL- 1 receptor. Methods. Primary hepatocyte cultures were prepared from normal mice, knocko ut mice deficient in the IL-1-converting enzyme (ICE), and knockout mice de ficient in the IL-1 p80 receptor. Hepatocytes were treated for 24 h with C difficile culture extract, purified C. difficile toxin A, or purified C. di fficile toxin B. The actin cytoskeleton was examined using confocal microsc opy, and LDH release was measured by spectrophotometric analysis. Results, C. difficile culture extract, toxin A, and toxin B induced collaps e of the actin cytoskeleton in hepatocytes from normal mice. Hepatocytes fr om both the ICE-deficient mice and the IL-1 p80 receptor-deficient mice dem onstrated similar responses to both toxins. These toxins also induced signi ficant LDH release in a concentration-dependent fashion in the normal hepat ocytes and the ICE-deficient hepatocytes. However, no significant increase in LDH release was observed in hepatocytes from IL-1 p80 receptor-deficient mice. Conclusions, C. difficile toxins induce actin cytoskeletal collapse indepen dent of IL-l or the IL-1 receptor, In contrast, toxin-stimulated LDH releas e was dependent on the presence of the IL-1 receptor. Thus, separate pathwa ys appear to mediate toxic effects as manifested by actin rearrangement and LDH release. (C) 2000 Academic Press.