Azaspiracids (AZAs) are marine toxins that are produced by Azadinium and Amphidoma dinoflagellates that can contaminate edible shellfish inducing a foodborne poisoning in humans, which is characterized by gastrointestinal symptoms. Among these, AZA1, -2, and -3 are regulated in the European Union, being the most important in terms of occurrence and toxicity. In vivo studies in mice showed that, in addition to gastrointestinal effects, AZA1 induces liver alterations that are visible as a swollen organ, with the presence of hepatocellular fat droplets and vacuoles. Hence, an in vitro study was carried out to investigate the effects of AZA1, -2, and -3 on liver cells, using human non-tumor IHH hepatocytes.
Exposure of IHH cells to AZA1, -2 or -3 (5×10-12 ‒ 1×10-7 M) for 24 h did not affect cell viability and proliferation (SRB and 3H-Thymidine incorporation assays), but induced a significant concentration-dependent increase of mitochondrial dehydrogenases activity (MTT reduction assay). This effect depends on the activity of mitochondrial electron transport chain complex I and II, being counteracted by rotenone and tenoyl trifluoroacetone, respectively. Furthermore, AZAs-increased mitochondrial dehydrogenase activity was almost totally suppressed in K+-, Cl-- and Na+-free media and sensitive to specific inhibitors of KATP and hERG potassium channels, Na+/K+ ATPase and CFTR chloride channels. Conclusions: These results suggest that AZA mitochondrial effects in hepatocytes derive from an imbalance of intracellular levels of K+ and in particular Cl- ions, as demonstrated by the selective reduction of toxin effects by CFTR chloride channel inhibition.
