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Complement-induced equine neutrophil adhesiveness and aggregation
Abstract:
Equine neutrophils (PMN) were isolated from citrated normal blood by density gradient separation on Ficoll-Hypaque to greater than 96% purity and 98% viability and an average of 3.78 x 10(7) PMN/ml. The agonist C5a des Arg was used in serial dilutions of whole zymosan-activated equine plasma (ZAP) or was partially purified from ZAP by column chromatography. Purified equine PMN exhibited rapid aggregation following incubation with C5a des Arg which was further dependent on the availability of divalent cations, especially Mg++. The microfilament disruptive agent cytochalasin B (5 micrograms/50 microliters) greatly augmented aggregation responses to C5a des Arg. Subaggregating doses of C5a des Arg promoted PMN adhesiveness as assayed on 0.5 x 10 cm borosilicate glass columns containing a 2.0 cm bed of Sephadex G-25. This C5a des Arg-induced increased adhesiveness was inhibitable by prior incubation of the PMN with either non-steroidal (0.065 M phenylbutazone) or steroidal (0.005 M dexamethasone) anti-inflammatory agents. Ultrastructural studies correlated well with functional assays and revealed marked organelle-free lamellipodia formation without PMN-PMN contact at subaggregating doses of the agonist and progressive PMN-PMN contact at aggregating doses. Equine PMN are responsive to C5a des Arg, and induced adhesiveness responses can be manipulated by anti-inflammatory agents.
Insights
Equine neutrophils respond to the C5a des Arg agonist, showing increased aggregation and adhesiveness. Anti-inflammatory drugs like phenylbutazone and dexamethasone can modulate these responses in horses.
Area of Science:
- Immunology
- Veterinary Medicine
- Cell Biology
Background:
- Neutrophils play a crucial role in the equine immune response.
- Understanding neutrophil activation is key to managing inflammatory conditions in horses.
Purpose of the Study:
- To investigate the effects of C5a des Arg on equine neutrophils.
- To assess the modulatory impact of anti-inflammatory agents on neutrophil function.
Main Methods:
- Isolation of highly pure equine neutrophils (PMN) using density gradient centrifugation.
- Stimulation of PMN with C5a des Arg derived from zymosan-activated plasma.
- Assays for PMN aggregation, adhesiveness, and ultrastructural changes.
- Evaluation of anti-inflammatory agents (phenylbutazone, dexamethasone).
Main Results:
- C5a des Arg induced rapid equine PMN aggregation, dependent on Mg++.
- Subaggregating doses of C5a des Arg increased PMN adhesiveness.
- Cytochalasin B enhanced aggregation responses.
- Anti-inflammatory drugs inhibited C5a des Arg-induced PMN adhesiveness.
- Ultrastructural analysis showed lamellipodia formation and PMN-PMN contact.
Conclusions:
- Equine neutrophils are responsive to C5a des Arg, exhibiting aggregation and increased adhesiveness.
- C5a des Arg-induced neutrophil responses can be modulated by both steroidal and non-steroidal anti-inflammatory drugs.
- These findings provide insights into equine neutrophil function and potential therapeutic targets.