Synthetic double-stranded RNA oligonucleotides are immunostimulatory for chicken spleen cells

Toll-like receptors (TLRs) are an integral part of the innate immune system that recognize microbe-derived molecular patterns and initiate innate and adaptive defenses against invading pathogens. TLR3 and TLR7 are involved in sensing virus-associated single-stranded and double-stranded RNA (dsRNA) molecules in cellular endosomes to activate the type I interferon pathway in mammals. Although certain synthetic dsRNA molecules have been identified to show immunostimulation in mammalian cells, very little is known about the ability of these sequences to stimulate avian cells. The current study investigated immunostimulatory properties of four synthetic oligonucleotide sequences using chicken splenocytes. Expression of TLR3 and 7, type I interferons and several other cytokines as well as TLR signaling pathway-related genes at different time points post-stimulation was quantified by real-time PCR. A dose-dependent increase in expression of TLR3 was observed in splenocytes treated with poly-UGUGU (poly-UG) and β-galactosidase dsRNA molecules. TLR3 and TLR7 gene expression was significantly up-regulated upon stimulation with all four dsRNA molecules. Furthermore, in a time course study, a significant increase was noted in the expression of TLR3, TLR7, interferon (IFN)-α, IFN-β, interleukin (IL)-1β, IL-6 as well as 2′,5′-OAS in splenocytes treated with poly-UG. In conclusion, the present study demonstrated the immunostimulatory properties of dsRNA oligonucleotides, especially those that contain a poly-UG motif, in chickens.