Macrophages are phagocytes that have a central role in the organization of the immune system after an
infection. These cells can recognize specific molecular components of micro-organisms (
pathogen-associated molecular patterns,
PAMPs) via specific receptors (
PRRs) and elicit specific cellular responses. In the past, the expression of immune genes in response to different
PAMPs has been characterized in different fish species. However, little is known about actual
cytokine release. We characterized the secretion of tumour
necrosis factor (TNF)-α in primary macrophage cultures of rainbow trout (Oncorhynchus mykiss) in response to several
PAMPs by Western blot and compared this to the induction of TNF-α gene expression as well as other pro-inflammatory
cytokines such as
interleukin (IL)-1β and
IL-6 and anti-viral molecules such as INF-α and
Mx protein (Mx). We show that
lipopolysaccharide (LPS) and
zymosan are major inducers of TNF-α secretion, which is not initially linked to the induction of TNF-α
mRNA expression. The secretion of TNF-α, but intriguingly not the expression, is also stimulated by ultrapure LPS meaning that, in fish, contaminants of commercial LPS preparations are better inducers of the inflammatory response. Moreover, we have characterized the signaling pathways that are activated by different
PAMPs and the link between those pathways and the final step of TNF-α secretion: TNF-α shedding by TNF-α converting
enzyme (TACE/ ADAM17). For the first time, we show that, in fish macrophages, TNF-α is processed by TACE-like activity and this cleavage is dependent upon the activation of ERK, p38MAPK and JNK signaling pathways by LPS.