In vitro immune-depression and anti-inflammatory activities of cantharidin on gilthead seabream ( Sparus aurata ) leucocytes activated by λ-carrageenan

Cantharidin is a natural compound with known therapeutic applications in humans. The aim of this study was to investigate the in vitro effects of cantharidin on gilthead seabream ( Sparus aurata ) head kidney leucocytes (HKL) stimulated with lambda- carrageenan. HKLs were incubated for 24 h with cantharidin (0, 2.5 and 5 mu g mL -1 ) and lambda- carrageenan (0 and 1000 mu g mL -1 ). The results showed that HKL viability only decreased by 15.2% after incubated with 5 mu g mL -1 of cantharidin and lambda- carrageenan. Cantharidin increased the peroxidase activity of HKLs only when incubated in combination with lambda- carrageenan. Besides this, cantharidin inhibited the respiratory burst and phagocytic activities. Furthermore, cantharidin induced morphological changes in HKLs (apoptotic and vacuolization signs) that were enhanced when incubated with lambda- carrageenan . Considering the analysis of the selected gene expression studied in HKLs [NF- kappa B subunits ( rela , relb , crel , nfkb1 , nfkb2 ), proinflammatory cytokines ( il1b, tnfa ), anti-inflammatory cytokines ( il10 , tgfb ) and caspases ( casp1 , casp3 , casp8 , casp9 )], although lambda- carrageenan up-regulated the expression of the proinflammatory gene il1b , lambda- carrageenan and cantharidin down-regulated its expression in HKLs. In addition, cantharidin up-regulated casp3 and casp9 expression. The casp3 and casp9 gene expression was down-regulated while casp1 gene expression was up-regulated in HKLs incubated with both cantharidin and lambda- carrageenan. All the effects of cantharidin are related to its inhibitory effect on protein phosphatases, which induce apoptosis at long exposure times, and minimize the effects of lambda- carrageenan. The present results provide detailed insight into the immune -depressive and anti-inflammatory properties of cantharidin on immune cells, which could be of interest to the aquaculture sector.