title: Characterization of Brucella abortus infection of bovine monocyte-derived dendritic cells.
abstract: Brucella abortus is a Gram negative facultative intracellular pathogen of cattle, and an important zoonosis in humans worldwide. Previous studies have shown that dendritic cells (DC) from humans and mice are highly permissive for Brucella survival and proliferation. Impairment of DC activation and maturation by Brucella infection has also been reported in these two species. The aim of this study was to characterize infection of bovine DC with B. abortus. Monocyte-derived DC (mdDC) were cultured from bovine peripheral blood mononuclear cells (PBMC) using the recombinant bovine cytokines IL-4 and GM-CSF. The resulting mdDC were DEC205(+), MHC class II(hi). Approximately 70% of the cultured cells were DEC205(+), MHC II(+). MdDC were infected with B. abortus strain 2308 at an MOI of 1 and 100. Parallel infection experiments were performed in monocyte derived macrophages (mdM) isolated from the same subjects. Bacteria were successfully killed by mdDC by 24 hours post infection (PI) with high and low dose of B. abortus, bacteria persisted in mdM infected with a high dose. Expression of IL-1b, IL-6, IL-10, IL-12p40, IFNγ, iNOS and TNFα in B. abortus infected and LPS stimulated mdDC at 6 and 24 hours PI were evaluated using RT-qPCR. At 6 hours PI all transcripts were increased over control cells and significantly less IL-10, IL-12p40, and IFNγ were expressed in mdDC infected with B. abortus compared to LPS stimulation. Evaluation of mdDC cultures by flow cytometry was performed. Flow cytometric analysis of infected and LPS stimulated mdDC 24 hours PI showed expression of CD80 and CD86 was impaired in two of the three animals analyzed. MHC class II expression was equivocal between the groups. From these results we conclude that cultured bovine mdDC are not permissive for intracellular proliferation of B. abortus, and infected mdDC exhibit some signs of maturational and activational impairment.
authors.affiliation: Department of Veterinary Medicine and Epidemiology, University of California Davis, One Shields Ave, Davis, CA 95616, USA. hellerme@missouri.edu
authors.forename: M C
authors.initials: MC
authors.lastname: Heller
authors.forename: J L
authors.initials: JL
authors.lastname: Watson
authors.forename: M T
authors.initials: MT
authors.lastname: Blanchard
authors.forename: K A
authors.initials: KA
authors.lastname: Jackson
authors.forename: J L
authors.initials: JL
authors.lastname: Stott
authors.forename: R M
authors.initials: RM
authors.lastname: Tsolis
mesh_terms: D000818:Animals; D002003:Brucella abortus / Q000276:immunology*; D002007:Brucellosis, Bovine / Q000276:immunology*; D002417:Cattle; D016207:Cytokines / Q000235:genetics / Q000276:immunology*; D003713:Dendritic Cells / Q000276:immunology*; D005260:Female; D005434:Flow Cytometry; D007963:Leukocytes, Mononuclear / Q000276:immunology; D012313:RNA / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary
chemical_list: D016207:Cytokines; D012313:RNA