Introduction. Antibiotics have been used for a long time in animal production to protect animals against pathogens. However, EU policy has been adopted to implement a sustainable production without adding antibiotics as growth promoters. Marine algae contain in their cell wall water soluble sulfated polysaccharides with potential biological activities such as anticoagulant, antiviral, antibacterial and immunomodulating activities that are being explored to be used as an effective alternative to antibiotics (1, 2). A crude extract containing sulfated polysaccharides was prepared from the green algae Ulva armoricana. The ability of this extract to stimulate the expression of the immune response mediators was evaluated using an in vitro system of porcine differentiated intestinal epithelial cells IPEC-1.
Materials and methods. The tested algae extract is composed of 11.6% of neutral sugars, 7.3% of proteins, 12.2% of uronic acids and 26.4% of sulfated polysaccharides. The capacity of this extract to stimulate the expression of immune mediators has been evaluated by RT-qPCR, using an in-vitro culture system of differentiated porcine intestinal epithelial cells, IPEC-1. Three doses (1, 0.1 and 0.01 mg/ml) were tested in comparison with E. coli O111:B4 LPS as positive control and with cells incubated alone as negative control. The extract was also tested on a human embryonic renal cell line, HEK293 which expresses TLR4/MD2/CD14, TLR2, TLR5, TLR9, NOD1 and NOD2, in order to identify which TLR or NLR receptor was involved. Expression of IL-8 with ELISA was used as a marker of stimulation. Statistical analysis was done with Kruskal-Wallis and Bonferroni-Dunn tests.
Results. Analysis by RT-qPCR showed that the algae sulfated polysaccharide extract, at 1 mg/ml, induced an increased expression of several cytokines in comparison with non-treated cells. CCL20 chemokine showed the highest stimulation (38 times higher than the control), followed by IL-8 (10.8 times higher), TNF-α (8.3 times higher), IL-1 α (7.1 times higher) and IL-6 (4 times higher). It also induced, to a lower extent, a significant increase of IL-1β, IL-12p40, TGF- β and PPAR γ expression. The algae extract did not seem to influence the expression of IL-10, CCL25, CCL28, TLR2 and TLR4. LPS did not induce the expression of these mediators in comparison with the algae extract.
When tested with HEK293 cells on several membrane receptors, it seemed that the algae extract stimulated the expression of immune factors via the activation of TLR4 membrane receptor
Discussion. These results showed that the sulfated polysaccharide-rich extract of green algae has the capacity to stimulate, in vitro, the expression of cytokines involved in the immune response. This suggests that this extract could be used as a new prophylactic strategy to stimulate the immune response of animals and reduce the use of antibiotics in farms, as this extract can be mixed in the feed or applied directly to drinking water, not disturbing the normal management of the farm.
O’Sullivan L. et al. Marine Drugs, 8, 2038-2064.
Chojnacka K.et al. The Open Conference Proceedings Journal, 3, (Suppl 1-M4), 20-28.
Key words. Algae, immunity, polysaccharides.
Presented at CLANA 2016.