Characterization and bioactivity of grass carp (Ctenopharyngodon idella) interleukin-21: Inducible production and involvement in inflammatory regulation

Abstract

In mammals, interleukin 21 (IL-21) is a broad pleiotropic cytokine that plays critical roles in the development of several inflammatory and autoimmune diseases. In fish, functional information of Il-21 is limited, and its role in immune response is largely unknown. In the present study, we cloned a coding sequence of grass carp (Ctenopharyngodon idella) il21 gene (gcil21). To characterize the release patterns and biological activity of gcIl-21, we prepared recombinant gcIl-21 (rgcIl-21) and obtained the polyclonal antibody with gcIl-21 specificity. Western blotting analysis showed that in grass carp head kidney leukocytes (HKLs), gcIl-21 was undetected in culture supernatant of untreated cells but drastically induced by heat-killed Aeromonas hydrophila (A. hydrophila), uncovering the release features of gcIl-21 and its possible involvement in immune response. Subsequent functional experiments revealed that rgcIl-21 did not affect the mRNA expression of grass carp il1b and tgfb, but induced a strong expression of grass carp il10, and to a lesser extent of grass carp tnfa in HKLs, suggesting a dominant effect of gcIl-21 in modulating Il-10 signaling as seen in rainbow trout and mammals. Furthermore, in vivo studies showed that intraperitoneal injection of rgcIl-21 was able to increase the survival rate of grass carp infected with live A. hydrophila, and reduce the pathological responses caused by the same pathogenic bacteria in head kidney and intestine. Taken together, these results for the first time revealed the close relationship of fish Il-21 production and function with inflammatory responses, and highlighted its anti-bacterial and anti-inflammatory ability, thereby providing a new insight into host defense mechanisms in fish.

Keywords: Anti-bacterial activity; Anti-inflammatory property; Grass carp; Il-21; Protein release.