PoCXCL8, a teleost chemokine, exerts direct bactericidal, chemotactic/phagocytic, and NETs releasing properties, promoting host anti-bacterial immunity.

As an important CXC chemokine, CXCL8 plays pleiotropic roles in immunological response. In teleost, CXCL8 is involved in cell migration and bacterial invasion. However, the immune antibacterial function of CXCL8 in Japanese flounder (Paralichthys olivaceus) (PoCXCL8) is largely scarce. In this research, we investigated the antibacterial property and leukocyte activation of PoCXCL8. PoCXCL8 consists of 100 amino acid residues, with a conserved chemokine CXC domain. PoCXCL8 was expressed in various tissues, with the highest level in liver and the lowest level in muscle, and sharply induced by V. harveyi or E. tarda in liver, spleen, and head kidney. In vitro, the recombinant PoCXCL8 (rPoCXCL8) could bind to Bacillus subtilis, Edwardsiella tarda, Escherichia coli, Pseudomonas fluorescens, Vibrio anguillarum, Vibrio harveyi, Staphylococcus aureus, and Micrococcus luteus, affect the growth of E. coli, E. tarda, M. luteus, and P. fluorescens, and have a direct bactericidal effect on E. coli and E. tarda. Moreover, rPoCXCL8 was able to bind the outer membranal protein rPilA of E. tarda. In addition, rPoCXCL8 could bind to PBLs, activating the PBLs activity including chemotaxis, proliferation, phagocytosis, reactive oxygen species, acid phosphatase activity. At same time, rPoCXCL8 could induce neutrophil to generate neutrophil extracellular traps (NETs) and promote the expression of inflammatory genes including IL-1β, IL6, MMP13, TNF-α, and NF-κB. In flounder, the presence of rPoCXCL8 could enhance the in vivo resistance to E. tarda in liver, spleen, and head kidney. Moreover, the PoCXCL8-deficient could attenuate the fish defense against E. tarda infection in in spleen and head kidney. In conclusion, these results provided new insights into the antibacterial properties of CXCL8 in P. olivaceus.
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