Activity of Granulocytes and Chemokines in the Leukocyte-encapsulation Response of Japanese Flounder Paralichthys olivaceus

The role of leukocytes from the kidney of Japanese flounder Paralichthys olivaceus in the encapsulation response was investigated using an in vitro model. The aggregation response of leukocytes to the target fish pathogen Ichthyophonus hoferi was observed over 12 h. The cellular aggregates mainly comprised granulocytes, which were characterized by intracellular peroxidase expression. Moreover, studies of isolated granulocytes demonstrated their adhesion to I. hoferi and formation of an initial layer in the encapsulation response. We observed that mRNA expression levels of three CC-chemokines (CCL3, CCL4 and CC-CLM) and one CXC-chemokine (IL-8) increased during the encapsulation process. Among these chemokines, CC-CLM and IL-8 were actively produced by granulocytes upon stimulation by I. hoferi. Flounder granulocytes therefore appeared to play an important role in the process by releasing specific chemokines upon pathogen recognition, thereby inducing subsequent cellular recruitment leading to encapsulation.