title: Analysis of immune responses directed toward a recombinant early secretory antigenic target six-kilodalton protein-culture filtrate protein 10 fusion protein in Mycobacterium bovis-infected cattle.
abstract: Cell-mediated immune responses are critical for protective immunity to mycobacterial infections. Recent progress in defining mycobacterial antigens has determined that region of difference 1 (RD1) gene products induce strong T-cell responses, particularly the early secretory antigenic target 6-kDa (ESAT-6) protein and culture filtrate protein 10 (CFP10). However, comprehensive analysis of the immune response towards these antigens is incompletely characterized. To evaluate recall responses to ESAT-6 and CFP10, peripheral blood mononuclear cells from M. bovis-infected cattle were stimulated in vitro with a recombinant ESAT-6 (rESAT-6)-CFP10 fusion protein and compared to responses induced by M. bovis-derived purified protein derivative. Following antigenic stimulation, activation marker expression was evaluated. Significant proliferative responses (P < 0.05) were evident in CD4(+), CD8(+), immunoglobulin M-positive, and CD172a(+) cell fractions after 6 days of culture. Expression of CD25 and CD26 was increased (P < 0.05) on CD4(+), CD8(+), and gammadelta T-cell-receptor-positive cells. CD4(+) and CD8(+) cells also exhibited significant changes (P < 0.05) in expression of CD45 isoforms. Using a flow cytometry-based proliferation assay, it was determined that CD45R expression is downregulated (P < 0.05) and that CD45RO expression is upregulated (P < 0.05) on proliferating (i.e., activated) CD4(+) cells. Collectively, data indicate that recall immune responses directed toward the rESAT-6-CFP10 fusion protein or purified protein derivative are comparable and that recall to mycobacterial antigens correlates with a CD45RO(+) phenotype.
authors.affiliation: Department of Molecular Microbiology and Immunology, University of Missouri, Columbia, 65211, USA.
authors.forename: Alexander C
authors.initials: AC
authors.lastname: Maue
authors.forename: W Ray
authors.initials: WR
authors.lastname: Waters
authors.forename: William C
authors.initials: WC
authors.lastname: Davis
authors.forename: Mitchell V
authors.initials: MV
authors.lastname: Palmer
authors.forename: F Chris
authors.initials: FC
authors.lastname: Minion
authors.forename: D Mark
authors.initials: DM
authors.lastname: Estes
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D002417:Cattle; D049109:Cell Proliferation; D018819:Dipeptidyl Peptidase 4 / Q000032:analysis / Q000378:metabolism; D015536:Down-Regulation; D017493:Leukocyte Common Antigens / Q000032:analysis / Q000378:metabolism; D008213:Lymphocyte Activation / Q000276:immunology; D009163:Mycobacterium bovis / Q000276:immunology*; D015375:Receptors, Interleukin-2 / Q000032:analysis / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000276:immunology*; D014380:Tuberculosis, Bovine / Q000276:immunology*; D015854:Up-Regulation
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; C505464:ESAT-6-CFP10 fusion protein; D015375:Receptors, Interleukin-2; D011993:Recombinant Fusion Proteins; D017493:Leukocyte Common Antigens; D018819:Dipeptidyl Peptidase 4