Genotyping of Toll-like receptor 4, myeloid differentiation factor 2 and CD-14 in the horse: an investigation into the influence of genetic polymorphisms on the LPS induced TNF-alpha response in equine whole blood.
Abstract: The inter- and intra-species differences in the response to lipopolysaccharides (LPS) are well recognised in mammalian species. It has been hypothesized that these differences can be attributed to genetic polymorphisms in the components involved in LPS signal transduction. These components include the cluster of differentiation factor 14 (CD-14), a membrane bound protein on the surface of mononuclear cells that recognises LPS and a receptor complex consisting of Toll-like receptor-4 (TLR-4) and myeloid differentiation factor-2 (MD-2). Sequencing of these three proteins in humans and mice revealed that all three are susceptible to polymorphic alterations, influencing the response to LPS. Previous experiments in the horse showed large inter-individual variations in the response to LPS. With the aim to assess this inter-individual variation, we performed a whole blood assay in 10 healthy horses as a functional assay to study the responsiveness to LPS. In 3 out of the 10 horses, LPS-induced TNF-alpha production was significantly lower compared to the overall mean. Subsequently the entire cDNA sequence encoding for the TLR-4, MD-2 and CD-14 protein was documented for each horse. Although mutations were observed in the sequence of TLR-4, these could not be related to an altered response to LPS in the concentration used in this study, as determined in the whole blood assay. Despite the various mutations found in the TLR-4 receptor protein, no alterations could be found in either the MD-2 or CD-14 gene, which are obviously more conserved structures.
Publication Date: 2006-02-14 PubMed ID: 16476493DOI: 10.1016/j.vetimm.2005.12.003Google Scholar: Lookup
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- Journal Article
Summary
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This study investigates the genetic differences in horses that might affect their response to lipopolysaccharides (LPS), a bacterial by-product. The focus is on three areas: the Toll-like receptor-4 (TLR-4), myeloid differentiation factor-2 (MD-2), and CD-14 genes. Genetic changes were found in the TLR-4 gene, but changes in these did not seem to affect how the horses respond to LPS. Changes in the MD-2 and CD-14 genes were not found, indicating they are more stable structures.
Background
- The research investigates the various individual differences seen in the response to lipopolysaccharides (LPS) among horses – a situation attributable to genetic differences.
- The proteins under study include the Toll-like receptor-4 (TLR-4) and myeloid differentiation factor-2 (MD-2), and cluster of differentiation factor 14 (CD-14). In previous investigations, these proteins have shown potential for mutative change influencing the response to LPS.
Procedure and Findings
- A whole blood assay was conducted on 10 healthy horses to study LPS responsiveness.
- Results indicated that three out of the 10 horses showed significantly lower LPS-induced TNF-alpha production compared to the overall average.
- The complete cDNA sequence encoding for the TLR-4, MD-2, and CD-14 proteins was documented for each horse.
- Mutations were observed in the TLR-4 sequence, however, these mutations did not demonstrate a correlation with a modified response to LPS, as determined in the whole blood assay.
Conclusions
- The TLR-4 receptor protein revealed various mutations, however, these mutations did not impact the horse’s response to LPS for the concentrations used in the study.
- The MD-2 and CD-14 genes displayed more structure conservation and did not exhibit any alterations, thus providing no evidence of a differential response to LPS resulting from possible genetic changes.
Cite This Article
APA
Werners AH, Bull S, Vendrig JC, Smyth T, Bosch RR, Fink-Gremmels J, Bryant CE.
(2006).
Genotyping of Toll-like receptor 4, myeloid differentiation factor 2 and CD-14 in the horse: an investigation into the influence of genetic polymorphisms on the LPS induced TNF-alpha response in equine whole blood.
Vet Immunol Immunopathol, 111(3-4), 165-173.
https://doi.org/10.1016/j.vetimm.2005.12.003 Publication
Researcher Affiliations
- Department of Veterinary Pharmacology, Pharmacy and Toxicology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 16, 3584 CM, Utrecht, The Netherlands. a.h.werners@vet.uu.nl
MeSH Terms
- Animals
- Cytotoxicity Tests, Immunologic / veterinary
- Female
- Horses / blood
- Horses / genetics
- Horses / immunology
- Lipopolysaccharide Receptors / genetics
- Lipopolysaccharide Receptors / immunology
- Lipopolysaccharides / immunology
- Lipopolysaccharides / pharmacology
- Lymphocyte Antigen 96 / genetics
- Lymphocyte Antigen 96 / immunology
- Male
- Polymorphism, Single Nucleotide
- RNA / chemistry
- RNA / genetics
- Reverse Transcriptase Polymerase Chain Reaction / veterinary
- Sequence Analysis, DNA
- Toll-Like Receptor 4 / genetics
- Toll-Like Receptor 4 / immunology
- Tumor Necrosis Factor-alpha / immunology
Citations
This article has been cited 9 times.- Mukhopadhyay A, Cook SR, SanMiguel P, Ekenstedt KJ, Taylor SD. TLR4 and MD2 variation among horses with differential TNFα baseline concentrations and response to intravenous lipopolysaccharide infusion. Sci Rep 2023 Jan 27;13(1):1486.
- Hellman S, Hjertner B, Morein B, Fossum C. The adjuvant G3 promotes a Th1 polarizing innate immune response in equine PBMC. Vet Res 2018 Oct 22;49(1):108.
- Tarlinton RE, Alder L, Moreton J, Maboni G, Emes RD, Tötemeyer S. RNA expression of TLR10 in normal equine tissues. BMC Res Notes 2016 Jul 19;9:353.
- Winfield LS, Dechant JE. Primary gastric rupture in 47 horses (1995-2011). Can Vet J 2015 Sep;56(9):953-8.
- Zamani F, Zare Shahneh F, Aghebati-Maleki L, Baradaran B. Induction of CD14 Expression and Differentiation to Monocytes or Mature Macrophages in Promyelocytic Cell Lines: New Approach. Adv Pharm Bull 2013;3(2):329-32.
- Bonin CP, Baccarin RY, Nostell K, Nahum LA, Fossum C, de Camargo MM. Lipopolysaccharide-induced inhibition of transcription of tlr4 in vitro is reversed by dexamethasone and correlates with presence of conserved NFκB binding sites. Biochem Biophys Res Commun 2013 Mar 8;432(2):256-61.
- Lewis DH, Chan DL, Pinheiro D, Armitage-Chan E, Garden OA. The immunopathology of sepsis: pathogen recognition, systemic inflammation, the compensatory anti-inflammatory response, and regulatory T cells. J Vet Intern Med 2012 May-Jun;26(3):457-82.
- Werling D, Coffey TJ. Pattern recognition receptors in companion and farm animals - the key to unlocking the door to animal disease?. Vet J 2007 Sep;174(2):240-51.
- Stejskalova K, Janova E, Splichalova P, Futas J, Oppelt J, Vodicka R, Horin P. Twelve toll-like receptor (TLR) genes in the family Equidae - comparative genomics, selection and evolution. Vet Res Commun 2024 Apr;48(2):725-741.
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