COMBINED ACTIVATION WITH AGONISTS OF TLR4, TLR9 AND NOD2 RECEPTORS SYNERGISTICALLY INCREASES PRODUCTION OF CYTOKINE-PROTEINS IN MOUSE MACROPHAGES
The possibility of enhancing the production of cytokines by macrophages differentiated by GM-CSF from the bone marrow of Balb/c mice has been studied under influence of single, paired and triple combinations of agonists of innate immunity receptors: TLR4, TLR9 and NOD2. The following agonists were used: for TLR4 - LPS from E. coli serotype 055: B5 at concentrations of 0, 20 and 100 ng/ml, for TLR9 - ODN CpG1826, for NOD2 - L18-MDP, both in concentrations of 0, 0.2 and 1 μg/ml. The synergism of IL-12p40, IL-10 and RANTES production is manifested in all paired and triple combinations of the TLR4, TLR9 and NOD2 agonists. Under influence of paired combinations of TLR4+TLR9, TLR4+NOD2 and TLR9+NOD2 agonists the production of IL-12p40 was increased 2.4, 4 and 7.3 times, of IL-10 - 2, 2.7 and 2 times, and of RANTES - 1.7; 2.6 and 1.9 times, respectively. The triple TLR4+TLR9+NOD2 agonistic composition induced 2,3-fold production of IL-12p40, 2.2-fold of IL-10, and 2.1-fold of RANTES as compared to the sum of responses to TLR4+TLR9 pair plus the one to NOD2. Neither paired nor triple combinations of TLR4, TLR9 and NOD2 agonists induced synergistic production of IL-1α, MIP-1α or MCP-1. The maximum production of IL-1α was observed upon exposure to TLR4 alone (125 ± 22 pg/ml at LPS 20 ng/ml), the maximum production of MIP-1α was observed upon exposure to TLR9 alone (378 ± 66 pg ml for ODN CpG1826 1 μg/ml) and the maximum production of MCP-1 was observed upon exposure to TLR4 alone (8436 ± 309 pg/ml at LPS 20 ng/ml).