In vitro effects of Clostridium difficile toxins on hepatocytes

Clostridium difficile infections are associated with development of the systemic inflammatory response, including the production of hepatic acute phase proteins. Lipopolysaccharide (LPS) directly stimulates the production of at least one of these proteins, a 23-kDa acute phase protein (the LPS-induced protein, or LIP) by murine hepatocytes in vitro. The aim of the present study was to determine if C. difficile toxins also stimulated the synthesis of this protein in vitro.

Cultured murine hepatocytes were treated for 24 h with various concentrations of C. difficile culture extract or purified toxins A and B in the presence or absence of dexamethasone or interleukin-1 (IL-1) receptor antagonist (IL-1 RA). The cells were then metabolically radiolabeled with [35S]methionine. Secretory proteins were identified using electrophoresis and autoradiography, and their synthesis was quantitated by image analysis of the autoradiograms.

The C. difficile culture extract, at dilutions as low as 1:200,000, significantly stimulated LIP synthesis in vitro. Toxins A and B, at concentrations as low as 1.6 and 0.02 pg/ml, respectively, also induced production of this protein. Dexamethasone further augmented C. difficile toxin-stimulated synthesis of LIP, but IL-1 RA inhibited the effects of these toxins on the synthesis of this protein. Only minimal quantities of IL-1 were found in culture supernatants following treatment with the toxins.

C. difficile toxins A and B, at very low concentrations, stimulate hepatocyte acute phase protein synthesis. Even though IL-1 RA inhibits this process, it does not appear that local production of IL-1 mediates the action of these toxins.