Conservation of cytokine-mediated responses in innate immunity: a flow cytometric study investigating the effects of human proinflammatory cytokines on phagocytosis in the earthworm Eisenia hortensis

Abstract

This study was aimed at determining the influence of human proinflammatory cytokines on innate immune responses in the earthworm Eisenia hortensis. Preincubation of earthworm coelomocytes in vitro with either interleukin-1 beta (IL-1 beta), granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin-2 (IL-2), or tumor necrosis factor-alpha (TNF-alpha) followed by subsequent bacterial challenge was carried out to investigate whether human proinflammatory cytokines would induce a state of enhanced responsiveness in phagocytic cells derived from the coelomic cavity of E. hortensis. The effect on phagocytosis by large coelomocytes (hyaline amebocytes) was evaluated using flow cytometry where the uptake of Escherichia coli expressing green fluorescence protein in the presence or absence of pretreatment with proinflammatory cytokines was measured. Our results show that proinflammatory cytokines enhanced phagocytosis to a statistically significant (p ≤ 0.05) degree in 10-18 % of earthworms tested for IL-1 beta, 20 % for GM-CSF, 20-27 % for IL-2, and 27-30 % for TNF-alpha, depending on the cytokine concentration used. Our results favor the suggestion that receptor-coding genes have been conserved through evolution between vertebrates and invertebrates.

Keywords

• Eisenia hortensis
• proinflammatory cytokines
• phagocytosis
• innate immunity
• flow cytometry
• hyaline amebocytes