The fusion protein BLS-FliC has the capacity to trigger dendritic cells activation in a TLR4 and TLR5 dependent way

ROSSI AH; ERREA A; HIRIART Y; BIEDMA, M.E; MORENO G; GOLDBAUM F; BERGUER P; RUMBO M

Tours

Simposio; DC 2014-13 th International Symposium on Dendritic Cells; 2014

During the last years there has been increasing research in the vaccines field that highlighted the need for adjuvantdevelopment. TLR agonists are adjuvant candidates based on their capacity to boost adaptive responses. With the aim todevelop a new adjuvant system, we have generated a fusion protein combining Lumazine synthase from Brucella spp. (BLS),withTLR4 dependent properties and flagellin (FliC), a TLR5 agonist. We addressed the activation of dendritic cells (DC), anessential step for the induction of the adaptive response elicited by BLS-FliC and studied the contribution of TLR5, TLR4 andMyD88 adaptor protein involved in signaling transduction in BLS-FliC induced DC activation. Bone marrow dendritic cells(BMDC) were derived from C57BL/6 mice (wt), TLR4-defficient, TLR5-/- or MyD88-/- mice with GM-CSF. Cells were stimulatedduring 18 h (30μg/ml BLS-FliC). Resulting response was compared with the one elicited by FliC (1μg/ml) and BLS (9μg/ml).BMDC activation was determined measuring the levels of co-stimulatory molecules by flow cytometry and cytokine secretionby ELISA. BLS-FliC increases the expression of CD80 and CD86 and secretion of IL12p40 and IL-6 on wt BMDC (p< 0.001).Moreover, all measured parameters were higher for BLS-FliC treatment compared to simulation with BLS or FliC as singlemolecules (p<0.001). On the other hand, BMDC activation triggered by BLS-FliC was conserved in absence of TLR5 or TLR4,suggesting TLR4 and TLR5 stimulatory properties of the chimera protein due to BLS and FliC. Finally, we´ve found that BLSFliCstimulatory potential was reduced in MyD88 KO BMDC compared to Wt cells but still remained statistically different fromcontrol treatment as shown by the expression levels of CD86 (p< 0.05). All together our results demonstrate that BLS-FliC isable to activate dendritic cells through a TLR5 and TLR4 pathways and partially dependent on MyD88.