Macrophage polarisation: immune responses of carp against parasites

Abstract

In the studies described in this thesis we used a natural host-parasite model of two parasites ( Trypanoplasma borreli and Trypanosoma carassii ) infecting common carp ( Cyprinus carpio L.), to obtain more knowledge about the phenomenon of macrophage polarisation in 'the evolutionary older' teleosts and the consequences of differential activation for the individual host.The general aspects of the teleost immune system are very similar to those of the mammalian immune system. Polarisation of macrophages into classically activated macrophages (caMF) or alternatively activated macrophages (aaMF) have been described for mammals, however, not yet for teleosts. These caMF and aaMF are involved in type I and type II immune responses, respectively. NO production is typically upregulated in caMF while arginase is upregulated in aaMF. Therefore, we propose the use of NO production by caMF and arginase activity from aaMF as markers for type I and type II immune responses in teleosts.To study arginase activity (aaMF) we cloned and sequenced the carp arginase 1 and 2 genes and optimised an arginase activity assay. Furthermore, we developed and characterised a carp macrophage culture system. These head kidney-derived macrophages can be considered the fish equivalent of mammalian bone marrow-derived macrophages. Stimulation of these head kidney-derived macrophages with LPS could induce iNOS expression and NO production (read-out for caMF), while stimulation with the intracellular messenger cAMP could induce arginase 2 expression and activity (read-out for aaMF).Macrophage polarisation has been described during many parasitic infections. T. borreli and T. carassii are kinetoplastid parasites infecting carp. The immune response of carp against these two parasites is different with regard to the production of nitrite by macrophages. We hypothesised that infections with these two parasites in carp provide a good model to study macrophage polarisation in teleosts. We infected carp with T. borreli and with T. carassii and followed the NO production and arginase activity in the head kidney leukocytes (HKL) of these carp. The results indicate that T. borreli -infected carp are more prone to increase nitrite production by caMF, while T. carassii -infected carp are more prone to increase arginase activity by aaMF. We also co-infected carp with both parasites simultaneously. Total parasitaemia was lower and survival was higher in co-infected carp. T. borreli -specific antibody levels were higher in co-infected and in T. carassii -infected carp than in T. borreli -infected carp. Our data indicate a protective effect of co-infection with T. carassii on the resistance to T. borreli , possibly mediated via cross-reactive antibodies. In an attempt to identify the antigenic proteins of the parasites we constructed expression libraries of both parasites and screened these libraries with homologous carp immune serum. Whether the antibodies against the identified antigenic proteins (ribosomal proteins, ubiquitin and activated protein kinase C receptor) are protective and cross-reactive,requires further studies.In conclusion, carp macrophages are able to polarise into classically and alternatively activated macrophages. To some extent this polarisation into caMF and aaMF can be induced by infection with T. borrreli and T. carassii , respectively. The level to which this polarisation in response to these parasites influences the production of (protective) antibodies remains to be elucidated