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Published on: May 31, 2018
Pattern-recognition receptor mRNA expression and function in canine monocyte/macrophages and relevance to canine anal
Arthur K House1, Susan P Gregory, Brian Catchpole
1Department of Veterinary Clinical Sciences, Royal Veterinary College, Hawkshead Lane, North Mymms, Hatfield, AL9 7TA, UK. ahouse@rvc.ac.uk
Abstract:
Pattern-recognition receptors (PRRs) are important components of the innate immune system, enabling early detection of infection. Defective PRR function has been implicated in several infectious and immune-mediated diseases of human beings, including Crohn's disease (CD). Anal furunculosis (AF) is an immune-mediated disease which primarily occurs in German shepherd dogs (GSD) and could result from a similar type of PRR dysfunction. The aim of the current study was to investigate canine PRR responses in vitro and to test the hypothesis that these were altered in AF-affected GSD. The pattern-recognition receptors TLR1, TLR2, TLR4, TLR6, TLR9, NOD1 (nucleotide-binding oligomerisation domain) and NOD2 were evaluated in the DH82 canine monocyte/macrophage cell line. These cells were found to express mRNA for all the selected PRRs with TLR2 mRNA the most and TLR5 mRNA the least abundant. A similar pattern of expression was found in canine blood-derived monocyte/macrophages. Stimulation of DH82 cells and blood-derived monocyte/macrophages using specific PRR-ligands, resulted in expression of pro-inflammatory cytokine mRNA. Quantification of TNFalpha mRNA and protein secretion from stimulated cells demonstrated variable responses with lipopolysaccharide (TLR4 ligand) and PAM(3)CSK4 (TLR1/2 ligand) proving to be the most potent and CpG DNA (TLR9 ligand) the least potent. Comparing PRR responses in blood-derived monocyte/macrophages from healthy blood-donor dogs with those from AF-affected GSD showed a deficiency in the latter in response to LD-MDP (NOD2 ligand) at the mRNA level but not at the protein level. It is possible that dysfunctional NOD2 responses by cells of the monocyte/macrophage lineage are involved in the pathogenesis of AF.
Insights
Canine pattern-recognition receptors (PRRs) were studied in German shepherd dogs with anal furunculosis (AF). AF-affected dogs showed altered NOD2 responses, suggesting a role in disease pathogenesis.
Area of Science:
- Immunology
- Veterinary Medicine
- Molecular Biology
Background:
- Pattern-recognition receptors (PRRs) are crucial for innate immunity and detecting infections.
- Dysfunctional PRRs are linked to immune-mediated diseases in humans, like Crohn's disease.
- Anal furunculosis (AF) in German shepherd dogs (GSD) may involve similar PRR defects.
Purpose of the Study:
- To investigate canine PRR responses in vitro.
- To test if PRR responses are altered in GSD with AF.
- To explore the role of PRR dysfunction in AF pathogenesis.
Main Methods:
- Evaluated mRNA expression of TLR1, TLR2, TLR4, TLR6, TLR9, NOD1, and NOD2 in DH82 canine cell line and blood-derived monocyte/macrophages.
- Stimulated cells with specific PRR ligands to assess pro-inflammatory cytokine mRNA and protein secretion (TNF-alpha).
- Compared PRR responses in healthy GSD versus AF-affected GSD.
Main Results:
- DH82 cells and blood-derived macrophages expressed all tested PRR mRNAs, with TLR2 most abundant.
- Ligand stimulation induced pro-inflammatory cytokine mRNA; LPS and PAM(3)CSK4 were potent, CpG DNA was least potent.
- AF-affected GSD showed deficient NOD2 response at mRNA level (LD-MDP stimulation) but not protein level compared to healthy dogs.
Conclusions:
- Canine monocyte/macrophage cells express key PRRs involved in innate immunity.
- Variable cytokine responses were observed upon PRR stimulation.
- Dysfunctional NOD2 responses in monocyte/macrophages may contribute to the pathogenesis of anal furunculosis in German shepherd dogs.
