The Interplay of TLR-NFκB Signalling Pathway and Functional Immune-Related Enzymes in the Inflammatory Response of Ciona robusta

Ascidians (Tunicata) are a powerful model for studying the innate immune system. To better understand the dynamics of immune responses under bacterial challenge in Ciona robusta, we exposed ascidians to lipopolysaccharide (LPS) injection. Immunohistochemistry analysis on two components of the nuclear factor kappa B (Nf kappa B) key signalling pathway, the Toll-like receptor 4 (TLR4) and NF kappa B, showed their over-expression on tissue of LPS-injected ascidians. Also, several enzymes related to immune responses were up-modulated following the LPS challenge. Our study suggests a broad and complex innate immune activation in the regulation of tunicate inflammatory responses. The close phylogenetic relationship between ascidians (Tunicata) and vertebrates makes them a powerful model for studying the innate immune system. To better understand the nature and dynamics of immune responses and the mechanisms through which bacterial infections are detected and translated into inflammation in Ciona robusta, we applied an approach combining in vivo lipopolysaccharide (LPS) stimulation, immune-labelling techniques and functional enzymatic analyses. The immunohistochemistry showed that Toll-like receptor 4 (TLR4) and nuclear factor kappa B (NF kappa B) were expressed during the inflammatory pharynx response 4 h post-LPS, with the formation of nodules in pharynx vessel lumen. Also, the endothelium vessels were involved in the inflammatory response. Observations of histological sections from naive and buffer-inoculated ascidians confirmed an immuno-positive response. Enzyme immune parameters-which included the activity of phenoloxidase, glutathione peroxidase, lysozyme, alkaline phosphatase and esterase-showed up-modulation 4 h after LPS injection, confirming their participation during ascidian inflammatory response. These findings provide new insights into the mechanisms underlying the LPS-induced C. robusta response and suggest that a broad innate immune mechanism, as in vertebrates, is involved in the regulation of inflammatory responses. Further findings in this direction are needed to cover knowledge gaps regarding the organized set of molecular and cellular networks involved in universal immune interactions with pathogens.