Analysis of immune gene expression modulated by benzo[a]pyrene in head kidney of olive flounder (Paralichthys olivaceus)

Poly aromatic hydrocarbons (PAHs) are known to cause functional disorder of fish immune responses. Alteration of inflammatory cytokines and other immune gene expressions by PAHs in immune organs may play a pivotal role in immunotoxicity. Thus this study aimed to elucidate the immunotoxic mechanism of PAH using benzo[a]pyrene (BaP) by analyzing the gene expression of cytokines (IL-1β, TNFα, IL-6, IL-8, IFNγ, Mx), apoptosis (FasL, SOD) and other immune related substances (Lysozyme, IgM) in head kidney and macrophage in olive flounder. In Q-PCR analysis, proinflammatory cytokine (IL-1β, IL-6, IL-8, TNFα) gene expressions were significantly upregulated by BaP while Mx and IgM gene expressions were significantly downregulated in head kidney by a longer exposure to BaP in vivo and in vitro. Lysozyme gene expression was initially upregulated but later downregulated in head kidney in vivo and in vitro. Inhibition test revealed that TNFα gene expression was upregulated by BaP via the AHR pathway as blocked by ANF while IL-6 and IFNγ gene expressions were upregulated by a calcium dependent pathway (i.e. NFAT) as blocked by EGTA. In primary macrophage cells, only IL-8 gene expression was significantly upregulated among proinflammatory cytokines while IFNγ, lysozyme and IgM gene expressions were downregulated by BaP. FasL and SOD expressions were not altered in head kidney cells but significantly upregulated in macrophage cells, indicating apoptosis and oxidative stress. These results indicate that exposure to BaP causes the downregulation of immune response by triggering the death of macrophage cells, the reduction of effectors like IgM and lysozyme, and the decrease of macrophage cell activity.