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Assessment of aspects of the toxicity of Clostridium perfringens E-toxin using the MDCK cell line

Protection and Life Sciences Division, CBD Porton Down, Salisbury, Wiltshire, SP4 OJQ, UK

1 The epithelial Madin Darby Canine Kidney (MDCK) cell line was used to study the toxicity of -toxin from Clostridium perfringens. The epithelial MDCK cell line is known to be sensitive to -toxin of Clostridium perfringens and to investigate its mechanism of action, the neutral red assay has been used to determine the viability of cultures of this cell line.

2 Comparison of the LC ₅₀s obtained at 34°C and 0°C showed that the lethality of -toxin was reduced by 18- fold at the lower temperature. The effect of tempera ture on -toxin lethality is unlikely to be due to reductions in membrane fluidity for the addition of Ca²⁺ or Mg²⁺ (2 mM) to buffer containing toxin was without effect. Varying the pH of the toxin-containing buffer from 6.9 to 8.7 did not increase the lethality of the toxin, though the most acidic pH used (5.8) was found to potentiate its action on MDCK cells.

3 The effect of inhibiting endocytosis on the lethality ofe toxin was also investigated by incubating cultures of MDCK cells with and without sodium azide over a range of concentrations of toxin. The co-administra tion of sodium azide did not reduce the toxicity of toxin, suggesting that energy-dependent uptake pro cesses such as endocytosis were unlikely to be involved in its mechanism of action. The results are, however, consistent with known receptor-based me chanisms of uptake and with other mechanisms of internalisation across the plasma membrane. -toxin thus interacts with cell surfaces by a temperature sensitive mechanism potentiated by low pH.

Key Words: -toxin • MDCK cell line • Clostridium perfringens • cell membrane effects

Human & Experimental Toxicology, Vol. 15, No. 11, 904-908 (1996)
DOI: 10.1177/096032719601501107


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