Inductive immune responses in the Japanese pufferfish (Takifugu rubripes) treated with recombinant IFN-γ, IFN-γrel, IL-4/13A and IL-4/13B

- Immune induction by four recombinant cytokines was tested in Japanese pufferfish.
- Recombinant IFN-γ, IFN-γrel, IL-4/13A and IL-4/13B induced immune responses.
- Treated fish showed increased phagocytosis and lysozyme activity.
- The treated fish had elevated expression of six inflammatory cytokine genes.
- Recombinant cytokines exerted regulatory effects on pufferfish immune system.

Immune regulation in response to recombinant cytokines (Th1- and Th2-types) is poorly understood in fish. Therefore, we synthesized and purified the Japanese pufferfish, (Takifugu rubripes) recombinant cytokines, namely interferon-gamma (IFN-γ), IFN-γrel, interleukin (IL)-4/13A and IL-4/13B, and then administered by injection to examine the immune regulatory effects on phagocytic activity, superoxide anion production and lysozyme activity, and cytokine gene expressions in treated fish head kidney (HK) at 1, 3 and 5 days post injection (dpi). Pufferfish that received rIFN-γ injection had an elevated phagocytic activity at 1 and 3 dpi, whereas rIFN-γrel and rIL-4/13A treated fish showed an increased phagocytic activity only at 1 and 3 dpi, respectively. Superoxide anion production increased only at 1 dpi in rIFN-γ, rIFN-γrel and rIL-4/13A injected pufferfish. Lysozyme showed a high activity at 1 dpi in rIFN-γ injected fish, at 5 dpi in rIL-4/13A injected fish and all through the time course in rIL-4/13B injected fish. rIFN-γ stimulation caused an elevated IL-1β and tumor necrosis factor-alpha (TNF-α) transcriptions, whereas stimulation with rIFN-γrel resulted in IFN-γ gene induction and a long term increase in IL-6 and IL-12p40 gene expressions. Injection of rIL-4/13A induced transcription of IL-6, IL-12p40 and TNF-α, while rIL-4/13B treatment enhanced IL-6, IL-12p35 and IL-12p40 gene expressions. Our results suggest that each recombinant cytokine has a role in activation of immune cells. More precisely, rIFN-γ may be potentially the most effective immune inducer among the tested cytokines. Therefore, use of recombinant Th1 and Th2 cytokines as immune enhancers could be an efficient technique for disease prevention in fish.