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ABSTRACT: The aim of this work was to develop mAbs against porcine CD205 and to conduct a comparative analysis of the CD205 protein expression on lymphoid tissues, monocyte-derived dendritic cells (DCs) and DCs isolated from the porcine skin. To conduct this study, we generated a monoclonal antibody, designated 1.F6F6, against the C-type lectin-like domain-5 of the porcine CD205 and showed that it recognizes a protein band of ∼200 kDa by Western Blot analysis in mesenteric lymph nodes cells. Flow cytometric analysis showed that the mAb 1.F6F6 recognized 28.5%, 28.1% and 34.1% of cells from tonsil, inguinal and mesenteric lymph nodes, respectively, and 6% of cells from thymus. Analysis of monocyte-derived DCs showed that approximately 20% were positive and activation of the cells with LPS increased the positive population to 36%. Analysis of DCs isolated from the porcine skin showed that approximately 70% of the cell population expressed the CD205 receptor. The development of a monoclonal antibody capable of recognizing the CD205 receptor in swine opens up possibilities of applying new strategies for enhancing vaccine efficacy by using the anti-CD205 antibody for DC antigen-targeting to enhance priming of immune responses.
Veterinary Immunology and Immunopathology 03/2012; 146(1):74-80. · 2.08 Impact Factor
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ABSTRACT: Dendritic cell antigen targeting primes robust immune responses in mouse models. Optimizing this immunization strategy in the actual hosts that require protection will advance development of efficacious contemporary vaccines. In a proof-of-concept study, we tested the immunogenicity of a single, low dose of a novel multi-component DNA construct expressing a CD205-targeted antigen fused to a CD40L minimal functional domain for linked DC activation. The DNA construct was formulated with DNA-encoded Flt3L and GM-CSF for DC recruitment and the formulation was evaluated in MHC class II-matched calves. Immunization of the calves with the CD205 antigen-targeting construct mixed with the cytokine constructs induced significant IFN-γ-secreting CD4(+) T-cells, CD4(+) T-cell proliferation, and antibody responses detectable within one week post-immunization. CD205 antigen-targeting significantly expanded IFN-γ-secreting CD4(+) T-cells, CD4(+) T-cell proliferation, and IgG antibody responses three weeks post-immunization. Nineteen weeks post-priming, the IFN-γ-secreting CD4(+) T-cells, CD4(+) T-cell proliferation, and the IgG titers were waning, but they remained significant. Following boosting at nineteen weeks post-immunization, the immune responses primed by the CD205-targeted antigen underwent rapid recall and the mean response tripled within one week post-boost. Comparative analysis of the immune responses observed one week post-priming versus the responses detected one week post-boost revealed that the average number of the IFN-γ-secreting CD4(+) T-cells observed in the calves immunized with the CD205 antigen targeting construct increased five-fold, the mean CD4(+) T-cell proliferation increased three-fold, whereas the mean IgG antibody titer increased two hundred-fold. These promising outcomes support testing the protective efficacy of CD205-targeted antigens in the calf model.
Vaccine 02/2012; 30(9):1624-35. · 3.77 Impact Factor
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Duncan M Mwangi,
Yoshikazu Honda,
Simon P Graham,
Roger Pelle,
Evans L N Taracha,
James Gachanja,
John K Nyanjui, Jocelyn Bray,
Guy H Palmer,
Wendy C Brown,
Waithaka Mwangi
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ABSTRACT: Theileria parva antigens recognized by cytotoxic T lymphocytes (CTLs) are prime vaccine candidates against East Coast fever in cattle. A strategy for enhancing induction of parasite-specific T cell responses by increasing recruitment and activation of dendritic cells (DCs) at the immunization site by administration of bovine Flt3L and GM-CSF prior to inoculation with DNA vaccine constructs and MVA boost was evaluated. Analysis of immune responses showed induction of significant T. parva-specific proliferation, and IFN-γ-secreting CD4(+) and CD8(+) T cell responses in immunized cattle. However, antigen-specific CTLs were not detected. Following lethal challenge, 5/12 immunized cattle survived by day 21, whereas all the negative controls had to be euthanized due to severe disease, indicating a protective effect of the vaccine (p<0.05). The study demonstrated the potential of this technology to elicit significant MHC class II and class I restricted IFN-γ-secreting CD4(+) and CD8(+) T cells to defined vaccine candidate antigens in a natural host, but also underscores the need to improve strategies for eliciting protective CTL responses.
Veterinary Immunology and Immunopathology 01/2011; 140(3-4):244-51. · 2.08 Impact Factor
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ABSTRACT: CD40 is mainly expressed by professional antigen-presenting cells (APCs). Its ligand, CD40L, is transiently expressed on activated CD4(+) T-cells. CD40-CD40L interactions mediate T-cell help to APCs and provide crucial signals for affinity maturation and B-cell class switching. In mammals, agonistic monoclonal anti-CD40 antibodies (mAbs) mimic the effects of CD40L on APCs, leading to enhanced T-cell priming and expansion, increased antibody production and class switching. In this study, we describe agonistic anti-chicken CD40 mAb 2C5. This mAb detected CD40 on primary chicken B-cells and macrophages, DT40 B-cells, and HD11 macrophages, induced NO synthesis in HD11 macrophages, and stimulated DT40 B-cell proliferation. These observations demonstrated at least partial functional equivalence of 2C5 to chicken CD154. This mAb may therefore constitute a new tool to study the role of CD40 in the chicken immune system, and its agonistic effects suggest that it could also be used as an adjuvant.
Developmental and comparative immunology 11/2010; 34(11):1139-43. · 3.29 Impact Factor
