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Functional studies of bovine alveolar neutrophils elicited with recombinant bovine IL-1 beta
J R Heidel1, H M Sassenfeld, C R Maliszewdki
1Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman 99164-7040.
Insights
Bovine interleukin-1 beta (IL-1) effectively recruits neutrophils to the lungs for functional analysis. Elicited neutrophils show enhanced release of specific arachidonate metabolites but similar superoxide release and bacterial killing compared to circulating neutrophils.
Area of Science:
- Veterinary Immunology
- Pulmonary Inflammation
- Cellular Immunology
Background:
- Pulmonary inflammation involves complex cellular responses, with neutrophils playing a key role.
- Interleukin-1 beta (IL-1) is a critical cytokine in inflammatory processes.
- Understanding neutrophil function in the lung is vital for diagnosing and treating respiratory diseases in cattle.
Purpose of the Study:
- To investigate the efficacy of bovine recombinant interleukin-1 beta (rbIL-1 beta) in eliciting alveolar neutrophils for functional studies.
- To compare the functional capacity of rbIL-1 beta-elicited alveolar neutrophils with circulating neutrophils, focusing on arachidonate metabolism, superoxide release, and bacterial killing.
Main Methods:
- Intrabronchial instillation of rbIL-1 beta into steer lungs to induce neutrophil recruitment.
- Bronchoalveolar lavage (BAL) to collect alveolar cells before and after IL-1 beta administration.
- Functional assays on elicited and circulating neutrophils, including stimulation with calcium ionophore A23187.
- Measurement of leukotriene B4 (LTB4), 5-hydroxyeicosatetraenoic acid (5-HETE), and other arachidonate metabolites using High-Performance Liquid Chromatography (HPLC).
- Assessment of superoxide anion release and bacterial killing capacity.
Main Results:
- Intrabronchial rbIL-1 beta successfully elicited a significant influx of neutrophils into the alveolar space within 4 hours.
- Elicited alveolar neutrophils released significantly greater amounts of 5-hydroxyeicosatetraenoic acid and LTB II compared to circulating neutrophils when stimulated.
- Circulating neutrophils pre-incubated or concurrently incubated with rbIL-1 beta and stimulated with A23187 showed increased release of 5-lipoxygenase metabolites.
- No significant differences were observed in superoxide anion release or bacterial killing activity between elicited alveolar neutrophils and circulating neutrophils.
Conclusions:
- Bovine IL-1 beta is an effective agent for recruiting neutrophils to the bovine lung for functional analysis.
- rbIL-1 beta enhances the production of specific arachidonate metabolites by alveolar neutrophils, suggesting a role in modulating inflammatory responses.
- Despite altered arachidonate metabolism, the primary effector functions of superoxide release and bacterial killing remain comparable between elicited alveolar and circulating neutrophils.
Abstract:
Bovine rIL-1 beta (rbIL-1 beta) was instilled intrabronchially into the lungs of steers to elicit harvestable alveolar neutrophils for functional analysis. Before instillation, bronchoalveolar lavage samples from the steers consisted of 96.4 +/- 1.5% (mean +/- SEM) macrophages, with the remaining cells neutrophils and occasional lymphocytes. Four hours after instillation of 1.0 and 10.0 nmol of IL-1, the lavage samples consisted of 96.3 +/- 0.8% and 91.0 +/- 5.7% neutrophils, respectively. Alveolar neutrophils elicited with rbIL-1 beta and challenged with the calcium ionophore, A23187, released similar amounts of leukotriene B4 (LTB4) and its nonenzymatic isomer LTB I, and significantly greater amounts of 5-hydroxyeicosatetraenoic acid and the nonenzymatic isomer LTB II, when compared with circulating neutrophils. The rbIL-1 beta did not, by itself, stimulate release of arachidonate metabolites from circulating neutrophils in quantities that were detectable by HPLC. Circulating neutrophils, preincubated with rbIL-1 beta and stimulated with A23187, released significantly greater amounts of 5-hydroxyeicosatetraenoic acid and total 5-lipoxygenase metabolites when compared with control cells not incubated with rbIL-1 beta. Incubation of circulating neutrophils with rbIL-1 beta and A23187 concurrently resulted in a significantly increased release of all 5-lipoxygenase metabolites of arachidonate. However, both the release of superoxide anion and bacterial killing by rbIL-1 beta-elicited bovine alveolar neutrophils did not differ from the values obtained for circulating neutrophils.