Overexpression of grass carp (Ctenopharyngodon idella) antimicrobial peptide Hepcidin activates NF-κB pathway and its effect on the growth Vibrio mimicus

Abstract:

[Background] Vibrio mimicus is a pathogenic bacterium that severely impacts aquaculture. The application of antibiotics to the control of vibriosis caused by this pathogen leads to drug resistance, environment pollution, and food safety problems. [Objective] To explore whether overexpression of the antimicrobial peptide Hepcidin (CiHep) from Ctenopharyngodon idella affects V. mimicus growth and NF-κB pathway at the cellular level. [Methods] The recombinant plasmids pEGFP-N1- Cihep and pcDNA3.1(+)- Cihep were constructed for eukaryotic expression. The expression of the fusion protein EGFP-CiHep was detected by immunofluorescence assay and Western blotting at different time points post transfection of pEGFP-N1- Cihep in C. idella kidney (CIK) cells. Then, the effect of the supernatant of the cells transfected with pcDNA3.1(+)- Cihep on the growth of V. mimicus was examined. The transcription levels of the key genes for NF-κB pathway activation and the downstream immunity-related genes in the transfected cells were measured. [Results] EGFP-CiHep was well expressed in the CIK cells and reached the highest expression level 48 h post transfection. The supernatant of the transfected cells significantly inhibited the growth of V. mimicus. CiHep overexpression significantly altered the transcription levels of key genes for NF-κB pathway activation and downstream immunity-related genes. [Conclusion] CiHep can inhibit V. mimicus growth and activate NF-κB pathway. Our results provide a theoretical basis for developing fish-derived Hepcidin and a new perspective on regulation of antimicrobial peptide expression by activating NF-κB pathway for prevention and treatment of fish vibriosis.