Cylindrospermopsin effects on cell viability and redox milieu of Neotropical fish Hoplias malabaricus hepatocytes.

Cylindrospermopsin (CYN) is a cyanotoxin that is cytotoxic to a wide variety of cells, particularly to the hepatocytes. In this study, the toxic effects of purified CYN were investigated in primary cultured hepatocytes of Neotropical fish Hoplias malabaricus. After isolation, attachment, and recovery for 72 h, the cells were exposed for 72 h to 0, 0.1, 1.0, 10, and 100 µg l-1 of CYN. Then, cell viability and a set of oxidative stress biomarker responses were determined. Catalase, superoxide dismutase, glucose-6-phosphate dehydrogenase, and glutathione S-transferase activities were not affected by exposure to CYN. Concentration-dependent decrease of glutathione reductase activity occurred for most CYN-exposed groups, whereas non-protein thiol content increased only for the highest CYN concentration. Lipid peroxidation, protein carbonylation, and DNA damage levels were not altered, but reactive oxygen species levels increased in the cells exposed to the highest concentration of CYN. Cell viability decreased in all the groups exposed to CYN. Thus, CYN may cause a slight change in redox balance, but it is not the main cause of cell death in H. malabaricus hepatocytes.

Subchronic effects of dipyrone on the fish species Rhamdia quelen.

The use of the non-steroidal anti-inflammatory drugs such as dipyrone is so widespread that this drug and its metabolites have been detected in effluents and surface water. This study aimed to evaluate the potential toxic effects of dipyrone on the aquatic environment, using a native fish species, Rhamdia quelen. Fish were exposed to three concentrations of dipyrone, 0.5, 5 and 50 µg/L, in the water for 15 days, and hematological, biochemical, genetic and morphological biomarkers were evaluated. The glutathione S-transferase activity decreased in the highest concentration in relation to the control group. In addition, hematocrit, red blood cells and thrombocyte counts were decreased in all three exposed groups in relation to the control group. The comet assay showed DNA damage at the lowest concentration of dipyrone and significant kidney damage. Those results suggest that a constant exposure of aquatic organisms to dipyrone presents potential toxic effects.

Genotoxic evaluation of different doses of inorganic lead (PbII) in Hoplias malabaricus.

Different genetic biomarkers have been used to evaluate the pollution effects of mutagenic agents such as metals and also a great variety of chemicals delivered on the environment by human activities. This way, the aim of the present report was to evaluate the effects of inorganic lead in fishes through the frequency of piscine micronuclei and nuclear morphological alterations in peripheral cells, chromosomal aberration frequency and comet assays in blood and kidney cells. Specimens of Hoplias malabaricus received different doses of lead by intra-peritoneal injections at time of 96 h. There was not a significant difference between control and treated groups for the piscine micronucleus and chromosomal aberration assays. In the comet assays there was a significant difference between control and contaminated groups. However, a significant difference between the applied doses was not observed. The results obtained with the comet assays also show that blood presented a higher sensibility than the kidney tissue, possibly due to the acute contamination. Although the results showed the genotoxic potential of lead at the 21 and 63 microg Pb(2 + )/g doses for both tissues, the lowest dose is considered more appropriate for future bioassays.

Multibiomarker in fish to evaluate a river used to water public supply.

We aimed to evaluate the ecological integrity of a large river, which receives agricultural and urban effluents and is used to water public supply. The fish species Astyanax bifasciatus was used as bioindicator during winter and spring 2012, and summer 2013 at the Middle Iguaçu River basin in Paraná state, Brazil. Water chemical and physical measures and ecotoxicological tests were carried out as well biochemical and genetic biomarkers in sampled fish in each period. The studied area was divided in three sample points: SP1, located where the water is collected to public supply; SP2, located in an urbanized area, and SP3, located at an urbanized area with the discharge of the sewage treatment. Although water chemical and physical analyzes were range of the Brazilian law to hydric bodies, anticholinesterasic effects were found in winter, oxidative stress in summer and spring. The higher genotoxic effect was in winter to all sample points. The temporal variation in biomarkers and the detection of caffeine in the water call attention to the water quality in this river mainly to be used to public supply.

Cellular responses of Prochilodus lineatus hepatocytes after cylindrospermopsin exposure.

Cylindrospermopsin is a potent toxicant for eukaryotic cells produced by several cyanobacteria. Recently, primary hepatocyte cultures of Neotropical fish have been established, demonstrating to be a quite efficient in vitro model for cellular toxicology studies. In the current study, a protocol for culture of Prochilodus lineatus hepatocytes was established and utilized to investigate the cellular responses to purified cylindrospermopsin exposure. Hepatocytes were successfully dissociated with dispase, resulting in a cell yield of 6.36 × 10(7)cells g(-1) of liver, viability of 97% and attachment on uncoated culture flasks. For investigation of cylindrospermopsin effects, hepatocytes were dissociated, cultured during 96 h and exposed to three concentrations of the toxin (0.1, 1.0 or 10 µgl(-1)) for 72 h. Cylindrospermopsin exposure significantly decreased cell viability (0.1 and 1 µgl(-1)) and multixenobiotic resistance mechanism, MXR (all exposed groups), but increased reactive oxygen/nitrogen species levels (all exposed groups) and lipid peroxidation (10 µgl(-1)). On the other hand no significant alterations were observed for other biochemical biomarkers as 2GSH/GSSG ratio, protein carbonyl levels and DNA strand breaks or glutathione S-transferase and glucose 6-phosphate dehydrogenase activities. In conclusion, hepatocytes might be made sensitive to cylindrospermopsin, at least in part, due to reduction of xenobiotics and endobiotics efflux capacity by MXR. Additionally, the toxin exposure suggests important issues regarding hepatocytes survival at the lowest cylindrospermopsin concentrations.