Host defense peptide LEAP-2 contributes to monocyte/macrophage polarization in barbel steed (Hemibarbus labeo)

The liver-expressed antimicrobial peptide 2 (LEAP-2) plays a vital role in host immunity against pathogenic organisms. In the present study, cDNA of the LEAP-2 gene was cloned and sequenced from the barbel steed (Hemibarbus labeo). The predicted amino acid sequence of the barbel steed LEAP-2 comprises a signal peptide and a prodomain, which is followed by the mature peptide. Sequence analysis revealed that barbel steed LEAP-2 belongs to the fish LEAP-2A cluster and that it is closely related to zebrafish LEAP-2A. We found that barbel steed LEAP-2 transcripts were expressed in a wide range of tissues, with the highest mRNA levels detected in the liver. In response to lipopolysaccharide (LPS) treatment, LEAP-2 was significantly upregulated in the liver, head kidney, spleen, gill, and mid intestine. A chemically synthesized LEAP-2 mature peptide exhibited selective antimicrobial activity against several bacteria in vitro. Moreover, LEAP-2, alone or in combination with LPS or phorbol 12-myristate 13-acetate, strongly induced a pro-inflammatory reaction in barbel steed monocytes/macrophages (MO/MФ), involving the induction of iNOS activity, respiratory burst, and the pro-inflammatory cytokines IFN-γ, TNF-α, and IL-1β. Collectively, the results of this study indicate the importance of fish LEAP-2 in the M1-type polarization of MO/MΦ.