Functional characterization of equine dendritic cells propagated ex vivo using recombinant human GM-CSF and recombinant equine IL-4.
Abstract: Naive T cells can be activated both in vivo and in vitro by specialized antigen presenting cells, dendritic cells (DC), with potent antigen-specific, immunostimulatory activity. Indeed, DC can provide an extremely powerful and important immunological tool by which to potentiate the immune response for specific recognition of foreign antigens. Until recently, the direct isolation of DC from PBMC required laborious procedures with extremely poor yields (<0.1%). Methods have been developed for the human, lower primate, and murine model systems to propagate large numbers of DC from PBMC or bone marrow ex vivo with various cytokines. However, all other model systems, including equine, still require the laborious isolation procedures to obtain DC. In this study, we have adapted the methods developed for the human system to generate large numbers of equine DC from PBMC precursors using recombinant human GM-CSF and recombinant equine IL-4. Our report is the first documentation of ex vivo generated DC from PBMC in a domesticated animal model system. Equine DC derived from PBMC were rigorously characterized by analyzing morphological, phenotypic, and functional properties and were determined to have similar attributes as DC generated from human PBMC. Equine DC appeared stellate with large projectiles and veils and had cell surface antigens at similar levels as those defined on human and murine DC. Furthermore, functional attributes of the DC included rapidly capturing antigens by pinocytosis, receptor-mediated endocytosis, and phagocytosis, activating naive T cells in a mixed leukocyte reaction to a much greater extent than macrophage or lymphoblasts, presenting soluble and particulate antigen 10-100 fold more effectively to T cells on a per cell basis than macrophage or lymphoblasts, and presenting soluble and particulate antigen to both CD4+ and CD8+ T cells. Taken together, our study provides a framework by which equine DC can now be readily produced from PBMC precursors and presents an impetus for and model by which DC can be simply generated in other animal model systems.
Publication Date: 1999-12-10 PubMed ID: 10587301DOI: 10.1016/s0165-2427(99)00094-xGoogle Scholar: Lookup
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- Journal Article
- Research Support
- U.S. Gov't
- P.H.S.
Summary
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The researchers in this study have successfully propagated and studied equine dendritic cells ex vivo (outside the body) using human GM-CSF and equine IL-4, two specific immune system regulators. The approach to study these immune cells, which are integral to mounting an immune response to foreign antigens, can be adapted for use across various animal models.
Background and Purpose
- The immune system uses dendritic cells (DC) to trigger immune responses. In particular, DCs help activate naive T cells, specialized immune cells that respond to specific foreign bodies (antigens).
- Prior to this study, obtaining DCs was a laborious process with low success rates. While significant progress had been made in propagating DCs from human and several animal model blood cells or bone marrow ex vivo using specific cytokines, these techniques hadn’t been adapted for other animals, including horses.
- The research had a two-pronged purpose: to develop a method for propagating equine DCs ex vivo using a modified human approach and to rigorously test these cells for akin functionality to human DCs.
Process and Findings
- The researchers used recombinant human GM-CSF and recombinant equine IL-4, two types of cytokines, to help propagate DCs from equine peripheral blood mononuclear cells (PBMCs).
- These newly grown equine DCs were then observed and tested for similar traits as human DCs. This included their appearance (morphology), surface markers (phenotype), and ability to perform certain functions.
- It was found that equine DCs closely resembled their human counterparts in both appearance and surface antigen levels.
- The DCs also demonstrated the desired functionalities such as efficient antigen capture through various processes (e.g., endocytosis, pinocytosis, and phagocytosis), effective activation of naive T cells, and presentation of antigens to multiple types of T cells more effectively than other cell types.
Impact and Future Applications
- This study is the first to showcase the production of ex vivo generated DCs from PBMCs in a domestic animal model, and the results affirm the potential to generate DCs in other animal model systems.
- The successful propagation and characterization of equine DCs bring forth vast possibilities for future research in immunology. The method and model developed can assist in understanding how DCs function across different species and could have potential implications for cross-species disease prevention or therapeutic interventions.
Cite This Article
APA
Hammond SA, Horohov D, Montelaro RC.
(1999).
Functional characterization of equine dendritic cells propagated ex vivo using recombinant human GM-CSF and recombinant equine IL-4.
Vet Immunol Immunopathol, 71(3-4), 197-214.
https://doi.org/10.1016/s0165-2427(99)00094-x Publication
Researcher Affiliations
- Department of Molecular Genetics and Biochemistry, School of Medicine, University of Pittsburgh, PA 15261, USA.
MeSH Terms
- Animals
- Antigen Presentation
- CHO Cells
- Cells, Cultured
- Cricetinae
- Dendritic Cells / physiology
- Endocytosis
- Granulocyte-Macrophage Colony-Stimulating Factor / pharmacology
- Horses / immunology
- Interleukin-4 / pharmacology
- Phagocytosis
- Recombinant Proteins / pharmacology
- T-Lymphocytes / immunology
Grant Funding
- 5RO1 AI25850 / NIAID NIH HHS
- 5T32 AI07487 / NIAID NIH HHS
Citations
This article has been cited 7 times.- Lee Y, Kiupel M, Soboll Hussey G. Characterization of respiratory dendritic cells from equine lung tissues. BMC Vet Res 2017 Nov 6;13(1):313.
- Dossa RG, Alperin DC, Hines MT, Hines SA. The equine CD1 gene family is the largest and most diverse yet identified. Immunogenetics 2014 Jan;66(1):33-42.
- Cavatorta DJ, Erb HN, Felippe MJ. Activation-induced FoxP3 expression regulates cytokine production in conventional T cells stimulated with autologous dendritic cells. Clin Vaccine Immunol 2012 Oct;19(10):1583-92.
- Fidalgo-Carvalho I, Craigo JK, Barnes S, Costa-Ramos C, Montelaro RC. Characterization of an equine macrophage cell line: application to studies of EIAV infection. Vet Microbiol 2009 Apr 14;136(1-2):8-19.
- Flaminio MJ, Borges AS, Nydam DV, Horohov DW, Hecker R, Matychak MB. The effect of CpG-ODN on antigen presenting cells of the foal. J Immune Based Ther Vaccines 2007 Jan 25;5:1.
- Mauel S, Steinbach F, Ludwig H. Monocyte-derived dendritic cells from horses differ from dendritic cells of humans and mice. Immunology 2006 Apr;117(4):463-73.
- Chan SS, McConnell I, Blacklaws BA. Generation and characterization of ovine dendritic cells derived from peripheral blood monocytes. Immunology 2002 Nov;107(3):366-72.
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