In vitro screening of immunomodulatory properties of synbiotics in chicken DT40 cell line*

Synbiotics (prebiotic & probiotic) are used to stimulate the proper composition of the intestinal microflora in farm animals. One of the desired properties of synbiotics is connected with the immunostimulatory potential that allows for maturation of the immune system and regulation of its functions. This study aimed at selecting synbiotics with the most pronounced immunostimulatory properties, expressed by regulation of immune-related genes. For this purpose we used an in vitro assay based on the DT40 cell line stimulation with different synbiotics and downstream analysis of gene expression. We used a combination of three prebiotics: RFO, inulin, Bi2tos, and three probiotic bacteria strains of Lactococcus lactis. Stimulation was carried out at 37°C and in a 5% CO2 atmosphere for 9 hours. Downstream analysis of gene expression was performed at the mRNA level, using the RT-qPCR method. Each of the 20 analysed genes belonged to (1) cytokines/chemokines/ regulatory molecules regulated by preand probiotics (i.e. IFN-γ, IFN-β, IL-4, IL-6, IL-8, IL-12p40, STAT4, CD3, CD80; iNOS), (2) the TLR2 signalling pathway (TLR2, MyD88) or (3) genes associated with the response of chickens to molecular patterns derived from gram-positive bacteria, i.e. lipoteichoic acid (MAPK8IP3, MAP2K4, MAP2K3, ITGB4, KLHL6, UNC13D and CARD11). The combination of the prebiotic inulin and probiotic Lactococcus lactis subsp. lactis SL2 provided the strongest regulation of the immune-related genes, which proves the immunostimulatory potential of this synbiotic.