Variability in the response of canine and human dendritic cells stimulated with Brucella canis

Abstract

Brucella canis is a small intracellular Gram-negative bacterium whose primary host is the dog, but it also can cause mild human brucellosis. One of the main causes of an inefficient immune response against other species of Brucella is their interaction with dendritic cells (DCs), which affects antigen presentation and impairs the development of an effective Th1 immune response. This study analysed the cytokine pattern production, by RT-qPCR and ELISA, in human and canine DCs against whole B. canis or its purified LPS. Human and canine DCs produced different patterns of cytokines after stimulation with B. canis. In particular, while human DCs produced a Th1-pattern of cytokines (IL-1β, IL-12, and TNF-α), canine cells produced both Th1 and Th17-related cytokines (IL-6, IL-12, IL-17, and IFN-γ). Thus, differences in susceptibility and pathogenicity between these two hosts could be explained, at least partly, by the distinct cytokine patterns observed in this study, where we propose that human DCs induce an effective Th1 immune response to control the infection, while canine DCs lead to a less effective immune response, with the activation of Th17-related response ineffective to control the B. canis infection.Electronic supplementary materialThe online version of this article (10.1186/s13567-017-0476-8) contains supplementary material, which is available to authorized users.

Brucella canis induces canine CD4+ T cells multi-cytokine Th1/Th17 production via dendritic cell activation

“…We recently described the expression and secretion of Th1 and Th17-type cytokines, including IL-17A in canine MoDCs stimulated with B. canis or its purified LPS [9]. Although some innate immune cells, including γδ T lymphocytes, iNKT and NK cells produce IL-17A [14], there were no records of IL-17A production in MoDCs from canines or other species prior to this work.…”

“…Bacterial concentration was estimated by comparing the OD at 560 nm with a standard curve and verified by plating on 5% bovine serum tryptic soy agar (TSA). LPS was isolated by TRIzol reagent (TRIzol ® Plus; Invitrogen Corp., Barcelona, Spain) as previously described [9]. The preparation was treated with enzymes (proteinase K, RNAse and DNAse) and subjected to a purification protocol using alcohols and Folch reagent.…”

“…Human and canine MoDCs were generated as previously described [9]. Monocytes were isolated from peripheral blood mononuclear cells (PBMCs) using an anti-human CD14 monoclonal antibody (clone TÜK4) conjugated to magnetic beads (MACS ® ;…”

“…Total cytoplasmic RNA was isolated from stimulated cells using 400 μL of ice-cold lysis buffer containing 0.5% Igepal ® CA-630 (Sigma-Aldrich, Saint Louis, MO, USA) as previously described [9]. First-strand cDNA strand was synthesized using a reverse obtained by normalizing the cytokine expression to endogenous control expression using the 2 -ΔΔCt method.…”

“…We previously proposed that although both human and canine MoDCs are activated after stimulation with B. canis or its purified LPS, their profiles of cytokine production are different. Th1 polarizing cytokines are produced by human MoDCs, while in canine cells Th1 and Th17-type cytokines are produced [9]. However, the nature of the responses that those stimulated MoDCs trigger remains unexplained for both species.…”