Inhibition of macrophage activity by mitogen-induced goldfish leukocyte deactivating factor

Macrophage activating and deactivating cytokines have been characterized in mammalian systems but little is known about these immunoregulatory molecules in fish. Using gel permeation and chromatofocusing fast performance liquid chromatography (GP-FPLC and C-FPLC) we partially purified a macrophage deactivating factor (MDF) from mitogen-induced goldfish kidney leukocytes. Inhibition of the macrophage-derived nitric oxide (NO) response induced by this MDF was time-, dose- and temperature-dependent. Macrophages pre-treated for 6 or 24 h with MDF before activation with macrophage activating factors (MAF) and/or bacterial lipopolysaccharide (LPS) exhibited a down-regulation in their NO response, while those treated with MDF 24 h after activation with MAF and LPS did not. MDF treatment also impaired the NO response of goldfish macrophages infected with the mammalian protozoan parasite Leishmania major. These results suggest that MDF exhibits its inhibitory effect downstream of the converging intracellular pathways induced by LPS and/or L. major. The novel teleost MDF has an approximate Mr of 15 kD and a pI < 4, and is the first endogenous molecule of teleosts known to down regulate macrophage antimicrobial responses.