Interleukin-6 (IL-6) is a pleiotropic cytokine that regulates hematopoiesis, inflammation, immune responses and bone homeostasis in mammals. Fish IL-6 has been cloned in recent years but to date no functional studies have been reported. Thus, in this paper we present for the first time in fish the functional characterisation of IL-6, using rainbow trout (Oncorhynchus mykiss) as the fish model and with a focus on macrophage effects. Trout IL-6 (tIL-6) expression in macrophages could be induced by proinflammatory agents (LPS, polyI:C, and IL-1β) and recombinant tIL-6 (rtIL-6) rapidly induced STAT3 phosphorylation and expression of SOCS-1 to -3, CISH and IRF-1, as seen in mammals. However, three findings in this study suggest a novel role of tIL-6 in fish. Firstly, macrophage growth was enhanced by rtIL-6 in vitro, suggesting that IL-6 produced during inflammatory events may promote macrophage proliferation locally. Secondly, rtIL-6 induced the expression of cathelicidin-2, an antimicrobial peptide with immune-modulatory function, but down-regulated the expression of IL-1β and TNF-α, indicating a role of IL-6 in host defence and also in limiting inflammation. Thirdly, rtIL-6 induced the expression of hepcidin in macrophages. In mammals hepcidin is antimicrobial but also regulates iron homeostasis by inhibiting iron absorption, and its expression is induced by IL-6 only in hepatocytes but not macrophages. Thus, in fish if IL-6 is induced in patrolling macrophages during sepsis this may act to reduce iron availability by induction of hepcidin expression and lead to iron deficiency, as a means to limit the spread of infection.