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Identification of a MHC class I-restricted CD8+ immune response to foot-and-mouth disease virus in cattle
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Abstracts / Veterinary Immunology and Immunopathology 128 (2009) 211–347
8. Mediators of recruitment and function of cells of the immune system; Fc receptors and immunoglobulins; signal transduction and gene expression in cells of the immune system Production and characterization of monoclonal antibodies to bovine IgG-Fc Tânia R. Penha 1,∗ , Ernesto R. Krüger 2 , Vanete Soccol 3 , JorgeV.B. Agottani 1 , Flávio H. Itano 1 , Ludmila D.C. Troiano 1 , Josiane Brodzinski 1 1
Instituto de Tecnologia do Paraná (TECPAR), Curitiba, PR, Brazil 2 Centro de Diagnóstico Marcos Enrietti, Curitiba, PR, Brazil 3 Universidade Federal do Paraná, Curitiba, PR, Brazil Keywords: IgG; Monoclonal antibodies E-mail address: [email protected] (T.R. Penha). Species: Ruminants Monoclonal antibodies directed to bovine IgG, were produced and characteriszed. From a total of seven hybridomas with a good performance, two of them were chosen and designated B4F11 and B3H12, taking in account the optical density evaluated on ELISA test, where culture medium was the start material. These monoclonals were probed in a western blot to detect IgG fragments originated by papaine proteolises, making known that B4H11 reacted with a conformational epitope, while B3H12 interacted in a specific manner with the Fc (fragment cristalizable) from bovine IgG. This monoclonal anti Fc was utilized in a conjugation procedure with fluorescein isothyocianate and peroxidase in order to develop indirect immunofluorescence and ELISA, respectively. doi:10.1016/j.vetimm.2008.10.213 Identification of a MHC class I-restricted CD8+ immune response to foot-and-mouth disease virus in cattle Efrain Guzman ∗ , Geraldine Shirley Ellis
Taylor, Bryan
Charleston,
Institute for Animal Health, Division of Immunology, Compton, Newbury RG20 7NN, United Kingdom Keywords: Foot-and-mouth disease virus; CD8+ T cells; ␣ CD4+ T cells; Interferon-␥; MHC class I-restricted CD8+ epitopes E-mail address: [email protected] (E. Guzman). Species: Ruminants Foot-and-mouth disease virus (FMDV), an apthovirus, is a member of the genus picornaviridae and can be classified into seven serotypes. It causes a highly contagious viral infection in ruminants, pigs and cloven-hooved animals with very important economic consequences. The disease is characterised by the formation of vesicles on the mouth, tongue, nose and feet. Control of the disease is achieved by vaccination with a chemically inactivated whole virus vaccine emulsified with adjuvant, providing only short-
term, serotype specific protection. A better understanding of protective immune mechanisms may help in development of novel vaccines. While much attention has been devoted to humoral responses to FMDV, less is known about the role of cell-mediated responses in protective immunity. An in vitro assay for the detection of antigen-specific interferon-gamma (IFN-␥) release by CD8+ T cells was used to determine the level of CD8+ T cell activity in vaccinated and infected cattle of known MHC type. MHC-restricted CD8+ T cell recognition of the structural proteins (P1) of FMDV was detected in vaccinated cattle. A significant cellular immune response to both live and UV-inactivated FMDV and to the P1 region of FMDV was also detected in FMDVinfected animals. Using mouse cells expressing individual cattle MHC class I genes, we have identified specific alleles responsible for the presentation of FMDV P1 peptides to CD8+ T cells. We are currently mapping MHC class Irestricted CD8+ epitopes which will enable us to carry out a detailed study of the role of CD8+ T cells following vaccination or infection in cattle. doi:10.1016/j.vetimm.2008.10.214 Identification of novel receptors for IgG Fc in bovine lymphocytes Matt A. Firth ∗ , Kuldeep S. Chattha, Douglas C. Hodgins, Patricia E. Shewen University of Guelph, Department of Pathobiology, Ontario Veterinary College, Guelph, Ontario, Canada Keywords: Soluble Fc receptor; CD16; CD32 E-mail address: mfi[email protected] (M.A. Firth). Species: Ruminants Receptors for the Fc portion of immunoglobin molecules (FcR) provide an important and vital link between circulating antibody and cellular effector functions. Cell surface FcR for IgG, IgE, IgA and IgM have been identified on many cell types. Much of the current literature focuses on FcR sequence and function in the murine and human Species. To date, only a handful of these FcRs have been sequenced and characterized in livestock species. Fc[gamma]RII (CD32) is an FcR present in several isoforms on a wide variety of cells including B cells, monocytes and neutrophils. On B cells, the Fc[gamma]RIIb isoform acts to down regulate IgG production in direct opposition to the B cell co-receptor complex consisting of CD19, CD21 and CD81 by means of an intracellular ITIM motif. A recent examination of the expression profile of CD32 in bovine blood mononuclear cells revealed the presence of two previously uncharacterized CD32-like sequences. Sequencing of the first uncharacterized transcript indicated it to be a CD32 mRNA splice variant, lacking a 120 bp region corresponding to the transmembrane region in the full-length molecule. Sequence homology with the published bovine Fc[gamma]RII sequence was 99% and 100% for the extracellular and intracellular domains, respectively. It is hypothesized that the resulting protein is a soluble isoform of CD32; this phenomenon has been described previously only in mice and humans. Analysis
Abstracts / Veterinary Immunology and Immunopathology 128 (2009) 211–347
8. Mediators of recruitment and function of cells of the immune system; Fc receptors and immunoglobulins; signal transduction and gene expression in cells of the immune system Production and characterization of monoclonal antibodies to bovine IgG-Fc Tânia R. Penha 1,∗ , Ernesto R. Krüger 2 , Vanete Soccol 3 , JorgeV.B. Agottani 1 , Flávio H. Itano 1 , Ludmila D.C. Troiano 1 , Josiane Brodzinski 1 1
Instituto de Tecnologia do Paraná (TECPAR), Curitiba, PR, Brazil 2 Centro de Diagnóstico Marcos Enrietti, Curitiba, PR, Brazil 3 Universidade Federal do Paraná, Curitiba, PR, Brazil Keywords: IgG; Monoclonal antibodies E-mail address: [email protected] (T.R. Penha). Species: Ruminants Monoclonal antibodies directed to bovine IgG, were produced and characteriszed. From a total of seven hybridomas with a good performance, two of them were chosen and designated B4F11 and B3H12, taking in account the optical density evaluated on ELISA test, where culture medium was the start material. These monoclonals were probed in a western blot to detect IgG fragments originated by papaine proteolises, making known that B4H11 reacted with a conformational epitope, while B3H12 interacted in a specific manner with the Fc (fragment cristalizable) from bovine IgG. This monoclonal anti Fc was utilized in a conjugation procedure with fluorescein isothyocianate and peroxidase in order to develop indirect immunofluorescence and ELISA, respectively. doi:10.1016/j.vetimm.2008.10.213 Identification of a MHC class I-restricted CD8+ immune response to foot-and-mouth disease virus in cattle Efrain Guzman ∗ , Geraldine Shirley Ellis
Taylor, Bryan
Charleston,
Institute for Animal Health, Division of Immunology, Compton, Newbury RG20 7NN, United Kingdom Keywords: Foot-and-mouth disease virus; CD8+ T cells; ␣ CD4+ T cells; Interferon-␥; MHC class I-restricted CD8+ epitopes E-mail address: [email protected] (E. Guzman). Species: Ruminants Foot-and-mouth disease virus (FMDV), an apthovirus, is a member of the genus picornaviridae and can be classified into seven serotypes. It causes a highly contagious viral infection in ruminants, pigs and cloven-hooved animals with very important economic consequences. The disease is characterised by the formation of vesicles on the mouth, tongue, nose and feet. Control of the disease is achieved by vaccination with a chemically inactivated whole virus vaccine emulsified with adjuvant, providing only short-
term, serotype specific protection. A better understanding of protective immune mechanisms may help in development of novel vaccines. While much attention has been devoted to humoral responses to FMDV, less is known about the role of cell-mediated responses in protective immunity. An in vitro assay for the detection of antigen-specific interferon-gamma (IFN-␥) release by CD8+ T cells was used to determine the level of CD8+ T cell activity in vaccinated and infected cattle of known MHC type. MHC-restricted CD8+ T cell recognition of the structural proteins (P1) of FMDV was detected in vaccinated cattle. A significant cellular immune response to both live and UV-inactivated FMDV and to the P1 region of FMDV was also detected in FMDVinfected animals. Using mouse cells expressing individual cattle MHC class I genes, we have identified specific alleles responsible for the presentation of FMDV P1 peptides to CD8+ T cells. We are currently mapping MHC class Irestricted CD8+ epitopes which will enable us to carry out a detailed study of the role of CD8+ T cells following vaccination or infection in cattle. doi:10.1016/j.vetimm.2008.10.214 Identification of novel receptors for IgG Fc in bovine lymphocytes Matt A. Firth ∗ , Kuldeep S. Chattha, Douglas C. Hodgins, Patricia E. Shewen University of Guelph, Department of Pathobiology, Ontario Veterinary College, Guelph, Ontario, Canada Keywords: Soluble Fc receptor; CD16; CD32 E-mail address: mfi[email protected] (M.A. Firth). Species: Ruminants Receptors for the Fc portion of immunoglobin molecules (FcR) provide an important and vital link between circulating antibody and cellular effector functions. Cell surface FcR for IgG, IgE, IgA and IgM have been identified on many cell types. Much of the current literature focuses on FcR sequence and function in the murine and human Species. To date, only a handful of these FcRs have been sequenced and characterized in livestock species. Fc[gamma]RII (CD32) is an FcR present in several isoforms on a wide variety of cells including B cells, monocytes and neutrophils. On B cells, the Fc[gamma]RIIb isoform acts to down regulate IgG production in direct opposition to the B cell co-receptor complex consisting of CD19, CD21 and CD81 by means of an intracellular ITIM motif. A recent examination of the expression profile of CD32 in bovine blood mononuclear cells revealed the presence of two previously uncharacterized CD32-like sequences. Sequencing of the first uncharacterized transcript indicated it to be a CD32 mRNA splice variant, lacking a 120 bp region corresponding to the transmembrane region in the full-length molecule. Sequence homology with the published bovine Fc[gamma]RII sequence was 99% and 100% for the extracellular and intracellular domains, respectively. It is hypothesized that the resulting protein is a soluble isoform of CD32; this phenomenon has been described previously only in mice and humans. Analysis