Abstract
Innate immunity is induced when pathogen-associated molecular patterns (PAMPs) bind host pattern recognition receptors (PRRs). Polyinosinic:polycytidylic acid [poly(I:C)] is a synthetic analogue of viral dsRNA that acts as a PAMP, inducing type I interferons (IFNs) in vertebrates. In the present study, the immunostimulatory effects of high molecular weight (HMW) poly(I:C) in rainbow trout cells were measured when bound to a cationic phytoglycogen nanoparticle (Nano-HMW). The physical characteristics of the nanoparticle itself, when bound to different lengths of dsRNA and when cell associated was evaluated. Optimal concentration and timing for innate immune stimulation was measured using the RTG-P1 reporter cell line. The immunostimulatory effects of HMW poly (I:C) was compared to Nano-HMW in vitro using the RTgutGC cell line cultured in a conventional monolayer or a transwell culture system. The ability of an activated intestinal epithelium to transmit an antiviral signal to macrophages was evaluated using a co-culture of RTgutGC cells and RTSll (a monocyte/macrophage cell). In all culture conditions, Nano-HMW was a more effective inducer of IFN-related antiviral immune responses compared to HMW poly (I:C) alone. This study introduces the use of cationic phytoglycogen nanoparticles as a novel delivery system for immunomodulatory molecules to enhance immune responses in aquatic vertebrates.
Highlights
The innate immune system of vertebrates consists of cells armed with pattern recognition receptors (PRRs) that sense pathogen-associated molecular patterns (PAMPs)
In order to assess the interactions of intestinal epithelial cells with the underlying antigen presenting cells, RTS11 in combination with RTgutGC cells within the transwell culture system was used
NPs are critical quality control parameters for properly characterizing a NP’s formulation. These properties determine the ability for uptake by cells, cellular trafficking and dictate the nature of immunological responses[28,29,30] Smaller NPs, in the size ranges of most viruses, are preferentially taken up via clathrin or caveolae-mediated endocytosis and can effectively deliver drugs across biological barriers[29]
Summary
The innate immune system of vertebrates consists of cells armed with pattern recognition receptors (PRRs) that sense pathogen-associated molecular patterns (PAMPs). We hypothesized that HMW poly(I:C) delivered by cationic phytoglycogen NPs (Nano-HMW) would be a more effective antiviral inducer in rainbow trout cells compared to the free HMW poly(I:C) These formulations were tested using an intestinal epithelial cell line (RTgutGC) cultured in a conventional monolayer and in a transwell culture system that better mimics an intestinal epithelial layer[26,27]. The antiviral responses stimulated by Nano-HMW and HMW poly(I:C) alone were measured by the level of expression of IFN and ISGs transcripts, Mx1 promoter activity, and antiviral protection assays This is the first study to evaluate innate antiviral responses induced by dsRNA in conjunction with cationic phytoglycogen NPs in fish. Our study presents a mechanism of PAMP delivery to fish that increases immune stimulation and represents an avenue for future development of antiviral therapies in an economically relevant fish species
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