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Migration of dog polymorphonuclear neutrophilic leukocytes to formylated peptides
D Linnekin1, C A Bowles, G Murano
1Armed Forces Radiobiology Research Institute, Department of Experimental Hematology, Bethesda, Maryland.
Abstract:
Formylated peptides are potent stimulants of polymorphonuclear neutrophilic leukocyte (PMN) migration from species such as humans and rabbits. Interestingly, PMNs from dogs, cats, pigs and cows have been reported as refractory to N-formyl-l-methionyl-l-leucyl-l-phenylalanine (FMLP) and generally are believed not to express formylpeptide receptors. Formylpeptides are a major component of conditioned media from E. coli cultures and believed to be a significant element in inflammatory responses elicited by E. coli. Our studies have found that E. coli filtrate was a potent stimulant of dog PMN migration. Inhibition of migration to E. coli filtrates by the antagonist t-botyloxycarbonyl-l-methionyl-l-leucyl-l-phenylalanine (t-boc-MLP) demonstrated that the migration was mediated through the formylated peptide receptor. Migration in response to peptides with higher affinity for the formylpeptide receptor than FMLP was further evidence for these receptors on the dog PMN. PMNs from dogs migrated in response to FMLP at high concentrations (100 microM); however, pretreatment with phorbol myristate acetate resulted in increased migration of dog PMNs in response to concentrations of FMLP as low as 1 pM. These results demonstrate that dog PMNs are responsive to formylpeptides and that these responses can be up-regulated by PMA. Thus PMNs from a species previously thought incapable of responding to formylpeptides can respond to formylpeptide analogs with high affinity for the receptor as well as be primed for enhanced migration to FMLP by PMA.
Insights
Dog neutrophils (PMNs) can respond to formylated peptides, contrary to previous beliefs. Their migration is mediated by formylpeptide receptors and can be enhanced by phorbol myristate acetate (PMA).
Area of Science:
- Immunology
- Cell Biology
- Veterinary Medicine
Background:
- Formylated peptides are known stimulants of polymorphonuclear neutrophilic leukocyte (PMN) migration in humans and rabbits.
- PMNs from several species, including dogs, are generally considered refractory to N-formyl-l-methionyl-l-leucyl-l-phenylalanine (FMLP) and lack formylpeptide receptors.
- Formylpeptides from E. coli are significant in inflammatory responses.
Purpose of the Study:
- To investigate the responsiveness of dog PMNs to formylated peptides.
- To determine if dog PMNs express functional formylpeptide receptors.
- To explore the priming effect of phorbol myristate acetate (PMA) on dog PMN migration.
Main Methods:
- Stimulation of dog PMN migration using E. coli filtrate.
- Inhibition assays using the formylpeptide receptor antagonist t-botyloxycarbonyl-l-methionyl-l-leucyl-l-phenylalanine (t-boc-MLP).
- Testing migration in response to FMLP and high-affinity formylpeptide analogs, with and without PMA pretreatment.
Main Results:
- E. coli filtrate potently stimulated dog PMN migration.
- Inhibition by t-boc-MLP confirmed formylpeptide receptor mediation.
- Dog PMNs migrated to high-affinity analogs and, at low concentrations, to FMLP after PMA priming.
Conclusions:
- Dog PMNs are responsive to formylated peptides via formylpeptide receptors.
- These responses can be significantly up-regulated by PMA.
- Previous assumptions about canine PMN refractoriness to formylpeptides are challenged.