title: Analysis of the antigen-specific IFN-gamma producing T-cell subsets in cattle experimentally infected with Mycobacterium bovis.
abstract: Three 10 months old cattle were infected by the intratracheal route with 10(6)cfu of a field strain of Mycobacterium bovis. Blood samples were regularly collected for in vitro IFN-gamma production after antigenic stimulation. Peripheral blood cells of infected animals produced IFN-gamma in response to crude M. bovis antigens (live and heat-inactivated BCG and protein-purified derivative (PPD)) 3-4 weeks after infection. The ratio of the response to bovine PPD versus avian PPD indicated a specific sensitisation for M. bovis antigens. Three months post-infection (PI), animals were culled and M. bovis was cultured from tubercle lesions. At different time points, the frequency of specific M. bovis IFN-gamma producing CD4+, CD8+ and WC1+ T-cells in the peripheral blood was examined by flow cytometry. Two colour immunofluorescence staining of intracellular IFN-gamma and bovine cell surface molecules showed that both CD4+ and CD8+, but not WC1+, T-cells produced IFN-gamma following stimulation with PPD, live or killed BCG. In two animals analysed, the relative percentage of circulating IFN-gamma producing CD8+ cells decreased between week 5 and week 9 PI. The same evolution was not observed for IFN-gamma secreting CD4+ cells. Magnetic positive selection of T-cells from infected animals showed that CD4+ T-cells produced specific IFN-gamma only in the presence of antigen presenting cells (APCs). Positively selected CD8+ T-cells secreted IFN-gamma only in the presence of recombinant human IL-2 and APCs. In vitro depletion of the CD4+ T-cells, but not the depletion of CD8+ or WC1+ T-cells, resulted in abrogation of the specific IFN-gamma production showing the key role of this cell population for the specific IFN-gamma production.
authors.affiliation: Veterinary and Agrochemical Research Center, Groeselenberg 99, B-1180, Brussels, Belgium. kawal@var.fgov.be
authors.forename: K
authors.initials: K
authors.lastname: Walravens
authors.forename: V
authors.initials: V
authors.lastname: Wellemans
authors.forename: V
authors.initials: V
authors.lastname: Weynants
authors.forename: F
authors.initials: F
authors.lastname: Boelaert
authors.forename: V
authors.initials: V
authors.lastname: deBergeyck
authors.forename: J-J
authors.initials: JJ
authors.lastname: Letesson
authors.forename: K
authors.initials: K
authors.lastname: Huygen
authors.forename: J
authors.initials: J
authors.lastname: Godfroid
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D002417:Cattle; D007371:Interferon-gamma / Q000096:biosynthesis*; D008297:Male; D016176:T-Lymphocyte Subsets / Q000276:immunology*; D014380:Tuberculosis, Bovine / Q000276:immunology*
chemical_list: D007371:Interferon-gamma