Cytotoxic quantification of maitotoxin-like activity from the dinoflagellate Gambierdiscus toxicus.
Gambierdiscus toxicus is a marine dinoflagellate involved in the food-borne disease ciguatera. Its toxicity is mainly due to maitotoxin, a hydrophilic toxin, the chemical structure of which has recently been described. This toxin increases internal Ca(2+) concentration and triggers phosphoinositide breakdown. Three cytotoxic tests were set for the detection and quantification of toxicity in G. toxicus extracts, further purified by HPLC: the neutral red uptake assay (NRU), observation of morphological alterations on baby hamster kidney fibroblasts (BHK 21/C13) and the measurement of internal Ca(2+) concentration on human lymphocytes (Jurkat D). In order to evaluate cytotoxicity tests, results were compared with the mouse bioassay. A positive correlation was observed between the minimum lethal dose (MLD), estimated at 24 hr by the mouse bioassay, and the IC(50), estimated by the NRU assay, for extracts of increasing degree of purity. By the use of the NRU assay, a precise detection and quantification of toxicity in multiple extracts was possible after 8 hr. Cell blebbing was generally highest for extracts that strongly inhibited NRU. The analysis of the correlation between the two cytological tests facilitates monitoring of the progression of purification. An indication of the presence of maitotoxin activity was supported by the increase of internal Ca(2+) concentration in Jurkat D lymphocytes, which was maximal in cells exposed to the purest extracts.
Diogène
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Unité INSERM 303, BP 3, Villefranche-sur-Mer, F-06230, France.
Dubreuil
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Breittmayer
J P
JP
Puiseux-Dao
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