Evolution of cytokine responses: IL-1β directly affects intracellular Ca2+ concentration of teleost fish leukocytes through a receptor-mediated mechanism

In this work we studied the biological activities of recombinant IL-1β from the teleosts sea bass (Dicentrarchus labrax) and rainbow trout (Oncorhynchus mykiss) by investigating the effects induced on intracellular Ca2+ concentrations ([Ca2+]i) of spleen leucocytes. Splenocytes were loaded with the Ca2+-permeant Fura-2AM, and then stimulated with rIL-1β. The emitted fluorescence was read for 5 min at 1 min intervals on a dual excitation fluorescence fluorimeter. Results showed that rIL-1β induced in both species a rise in [Ca2+]i, and a subsequent decrease until 5 min after stimulation. The stimulating effect was dose-dependent in both species reaching a plateau at 200 ng/ml of rIL-1β, was abolished by heat-treatment of rIL-1β, and affected in a dose-dependent fashion by treatment of leucocytes with trypsin. These features suggested a functional IL-1 receptor was involved in the binding. The observed rise in [Ca2+]i was not detected in human PBMC and was species-specific, since rIL-1β from sea bass, trout, and human were unable to interfere each other in the assay. Moreover, incubation of splenocytes with rIL-1β induced a rapid tyrosine phosphorylation of a 24 kDa polypeptide in both species. This work represents the first evidence of a direct effect on [Ca2+]i induced by IL-1β and suggests that in the evolution of IL-1 activities, teleost fishes display a peculiar IL-1-associated behaviour that is lacking in mammals.