Morphological and cytoskeletal changes caused by non-membrane damaging cytotoxin of Vibrio cholerae on Int 407 and HeLa cells

Vibrio cholerae produces a non-membrane damaging cytotoxin (NMDCY), also kn own as cell rounding factor, which causes rapid rounding of cultured cells like HeLa, CHO and Vero and reportedly elicits enterotoxic activity in the rabbit ileal loop assay. Pursuing the concept that NMDCY might be an access ory factor contributing to the diarrhea caused by V. cholerae, we investiga ted the effect of NMDCY on Int 407 (intestinal cell line) and HeLa (non-int estinal cell line) cells using light, fluorescent and electron microscopy t o gain insight into the cellular response evoked by NMDCY. Binding assays s howed that NMDCY has affinity for both Int 407 and HeLa cells. Changes in t he internal organelles and cytoskeletal structures of the cell lines were d ocumented indicating changes in the secretory and metabolic function of the toxin-treated cells. Toxin-treated cells visualized under the electron mic roscope revealed retraction of cell body, formation of blebs on cell surfac e, changes in mitochondria having dilated and rarefied matrix and an extens ively developed Golgi apparatus, endoplasmic reticulum and lysosomes compar ed to those in normal cells. Immunofluorescence study showed restructuring of microfilament network represented by actin, filamin and vinculin, as als o of the microtubular component, tubulin and the intermediate filament, vim entin, Immunogold study further revealed that the toxin is internalized eve n within the nucleus. Moreover, a rise in the intracellular calcium level o f the NMDCY-treated cells leads us to hypothesize that a cascade of events results in the final impairment of the cell machinery. (C) 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. A ll rights reserved.