Toxicity and glutathione implication in the effects observed by exposure of the liver fish cell line PLHC-1 to pure cylindrospermopsin

Cylindrospermopsin (CYN), a cyanotoxin produced by several freshwater cyanobacteria species, has been reported to cause human and animal intoxications. CYN is a potent inhibitor of protein and glutathione synthesis. In order to study these effects, various in vitro models have been used, which are representative of the organs targeted by the toxin. However, studies concerning CYN toxicity to fish species, both in vivo and in vitro, are still very scarce.To our knowledge, this is the first work dealing with the effects of CYN in a fish cell line. In the present work, we tried to test the hypothesis that CYN could be hepatotoxic to fish causing cell damage and oxidative stress, which may lead to pathogenicity. To deal this purpose, PLCH-1 cells, derived from fish liver, were exposed to concentrations that ranged from 0.3 to 40 μg/mL CYN during 24 and 48 h for the cytotoxicity study, and 2, 4 and 8 μg/mL CYN for the oxidative stress assays. The basal cytotoxicity endpoints studied were protein content, neutral red uptake and the tetrazolium salt, MTS, reduction. The biomarkers used for the oxidative stress study were reactive oxygen species (ROS) content, reduced glutathione content and γ-glutamylcysteine synthetase activity.The cytotoxicity endpoints revealed a decrease in the cellular viability in a time and concentration-dependent way. Moreover, when cells were exposed to pure CYN, an increase in the ROS content was observed, being more marked at the higher concentrations used. Finally, the present work shows alterations in GSH content and synthesis due to CYN. Moreover, a relationship between cytotoxic effects and ROS production has been evidenced. The results obtained confirm the alteration on fish liver cells, which should be considered relevant to what it may happen in real scenarios since fish are frequently in contact with this cyanotoxin.