Clostridium difficile toxin B inhibits the secretory response of human mast cell line-1 (HMC-1) cells stimulated with high free-Ca²⁺ and GTPγS.
Clostridium difficile toxins A and B (TcdA and TcdB) belong to the class of large clostridial cytotoxins and inactivate by glucosylation some low molecular mass GTPases of the Rho-family (predominantly Rho, Rac and Cdc42), known as regulators of the actin cytoskeleton. TcdA and B also represent the main virulence factors of the anaerobic gram-positive bacterium that is the causal agent of pseudomembranous colitis. In our study, TcdB was chosen instead of TcdA for the well-known higher cytotoxic potency. Inactivation of Rho-family GTPases by this toxin in our experimental conditions induced morphological changes and reduction of electron-dense mast cell-specific granules in human mast cell line-1 (HMC-1) cells, but not cell death or permeabilisation of plasma-membranes. Previously reported patch-clamp dialysis experiments revealed that high intracellular free-Ca(2+) and GTPγS concentrations are capable of inducing exocytosis as indicated by significant membrane capacitance (Cm) increases in HMC-1 cells. In this study, we investigated the direct effects of TcdB upon HMC-1 cell "stimulated" Cm increase, as well as on "constitutive" secretion of hexosaminidase and interleukin-16 (IL-16). Compared to untreated control cells, HMC-1 cells incubated with TcdB for 3-24h exhibited a significant reduction of the mean absolute and relative Cm increase in response to free-Ca(2+) and GTPγS suggesting an inhibition of secretory processes by TcdB. In conclusion, the HMC-1 cell line represents a suitable model for the study of direct effects of C. difficile toxins on human mast cell secretory activity.
['Balletta|Andrea|A|', 'Lorenz|Dorothea|D|', 'Rummel|Andreas|A|', 'Gerhard|Ralf|R|', 'Bigalke|Hans|H|', 'Wegner|Florian|F|']
D001426:Bacterial Proteins D001427:Bacterial Toxins D002118:Calcium D002460:Cell Line D003956:Dialysis D025802:Electric Capacitance D005089:Exocytosis D016244:Guanosine 5'-O-(3-Thiotriphosphate) D006596:Hexosaminidases D006801:Humans D019410:Interleukin-16 D008407:Mast Cells D008564:Membrane Potentials D018408:Patch-Clamp Techniques D013997:Time Factors
Human mast cell line-1 (HMC-1) GTPγS Membrane capacitance Hexosaminidase Clostridium difficile toxin B (TcdB) Whole-cell patch-clamp