Abstract
The overexpression of GATA-3, T-bet and TGF-ß may theoretically induce IL-4/A, IFN-γ and IL-17A expression, respectively. Whether this also applies to fish is not yet known. The plasmid vectors encoding reporter gene (RFP)-tagged T-bet, GATA-3 and TGF-ß were used as overexpression tools, transfected into cells or injected intramuscularly to monitor the expression of IFN-γ, IL-4/13A and IL-17A. In addition, the fish were either experimentally challenged with Vibrio anguillarum (VA group) or Piscirickettsia salmonis (PS group). The reporter gene (RFP) inserted upstream of the GATA-3, T-bet and TGF-ß genes, was observed in muscle cell nuclei and in inflammatory cells after intramuscular (i.m.) injection. PS group: following the injection of GATA-3 and T-bet-encoding plasmids, the expression of GATA-3 and T-bet was high at the injection site. The spleen expression of IFN-γ, following the injection of a T-bet-encoding plasmid, was significantly higher on day 2. VA group: The T-bet and GATA-3-overexpressing fish expressed high T-bet and GATA-3 mRNA levels in the muscles and on day 4 post-challenge. The expression of TGF-ß in the muscles of fish injected with TGF-ß-encoding plasmids was significantly higher on days 7 (8 days pre-challenge) and 19 (4 days after challenge). The protective effects of the overexpression of T-bet, GATA-3 and TGF-ß on both bacterial infections were negligible.
Highlights
The aquaculture production of Atlantic salmon is threatened by infections from a wide array of pathogens
To determine whether the expression vectors induced gene expression in vitro, and to gain information on the sub-cellular localization of T-bet, GATA-3 and TGF-β expression following transfection, fluorescence microscopy of cells transfected with T-bet, GATA-3 and TGF-β-encoding plasmids that were fused to red fluorescent protein (RFP) was carried out
The analysis revealed the the presence of RFP-elicited fluorescence thesections sections obtained obtained from pTagRFP-Tpresence of RFP-elicited fluorescence ininallallthe fromfish fishthat thatreceived received pTagRFP-T-bet, bet, pTagRFP-GATA-3 and pTagRFP-TGF-ß
Summary
The aquaculture production of Atlantic salmon is threatened by infections from a wide array of pathogens. Some of them are not effectively controlled by current vaccines (consisting of inactivated pathogens in water-in-oil formulations), which mainly show effects through their B-cell response, whereas a robust T cell response is suboptimal, limiting the vaccine efficacy. One way to increase vaccine efficacy is to develop targeting molecular adjuvants [1,2,3], which may induce T cell activation. It is unknown whether the T cell differentiation process in fish is equal to that described in higher vertebrates and mammalian species. Two of Biology 2020, 9, 82; doi:10.3390/biology9040082 www.mdpi.com/journal/biology
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