The Regulation Of Ayu(plecoglossus Altivelis)urocortin In The Monocyte/macrophage Function

Urocortin(UCN)is a hormone in the hypothalamic–pituitary–adrenal axis.It is expressed in various immune cells and participates in the regulation of peripheral immune response.However,the function of teleost UCN in the immune system remains unclear.In this study,we cloned the c DNA sequence of UCN from ayu Plecoglossus altivelis(Pa UCN)and analyzed the effect on ayu monocyte/macrophage(MO/MФ)function.Pa UCN consisted of 1722 nucleotides and contained an open reading frame(ORF)which encoded a 170 amino acid sequences(AAs)protein and with a molecular weight(MW)of19.5 k Da.Multiple sequence alighment and phylogenetic tree analyses revealed that Pa UCN contained conservative signal peptide and mature peptide.Pa UCN clustered within the fish urocortin 1(UCN1)group and was most related to the rainbow trout(Oncorhynchus mykiss)UCN(74.7%).Pa UCN was expressed in all tested tissues,including some major immune organs and the expression was increased in the liver,spleen,head kidney,and gill upon Vibrio anguillarum infection.We synthesized mature Pa UCN protein(m Pa UCN)and found that m Pa UCN treatment significantly improved the phagocytosis and bacterial killing of MO/MФ.m Pa UCN exhibited antimicrobial capability against some tested bacteria and showed the strongest activity against Listeria monocytogenes(minimal inhibit concentration,MIC: 6.25 μg/ml).Under the infection of V.anguillarum,m Pa UCN reduced pro-inflammatory cytokine expression and increased anti-inflamamtory cytokine expression in MO/MФ.Glutathione S-transferase(GST)-Pull down indicated that m Pa UCN reduced pro-inflammatory cytokine expression in MO/MФ,which was partially mediated via interaction with Plecoglossus altivelis interleukin-6(Pa IL-6).m Pa UCN reduced bacterial load in liver,spleen,kidney and blood and increased the survival of V.anguillarum-infected ayu.In addition,m Pa UCN significantly decreased the pro-inflammatory cytokine expression and increased the anti-inflammatory cytokine expression in the liver,spleen,and kidney of V.anguillarum-infected ayu.Overall,UCN as an endocrine factor regulates the immune response of ayu after infection by activating MO/MФ,thus contributing to enhance fish survival.