Related Experiment Videos
Oxygen radicals and nitric oxide production by turkey respiratory macrophages
A E Van Nerom1, R Ducatelle, F Haesebrouck
1Department of Pathology, Bacteriology and Avian Diseases, Faculty of Veterinary Medicine, University of Ghent, Belgium. anne.vannerom@rug.ac.be
Abstract:
The influence of different induction protocols on the recovery of elicited turkey respiratory macrophages (RM), and on their oxygenation activity and nitric oxide (NO) production was examined. RM were induced in three week old specific pathogen free turkeys with Sephadex G-50, Thioglycollate broth, and an emulsion of incomplete Freund's adjuvant (IFA), supplemented either with Mycoplasma hyorhinis grown in Modified Channock broth (IFA-M. hyorhinis) or with Modified Channock broth (IFA-Broth). The RM were recovered by lavage of the lungs and air sacs and were purified by centrifugation through a Percoll suspension. Their oxygenation activity was evaluated in luminol-enhanced chemiluminescence assays, following stimulation with Zymosan A. The NO production was evaluated by incubating the RM with lipopolysaccharide (LPS) from Salmonella enteritidis for 24 or 48 hours. The number of recovered RM was slightly, but not significantly lower for Sephadex G-50 and IFA-Broth than for Thioglycollate broth and IFA-M. hyorhinis. RM elicited with Sephadex G-50 and IFA-Broth showed a significantly higher oxidative burst response to Zymosan A, compared to the Thioglycollate and IFA-M. hyorhinis elicited RM. Although all elicited RM showed a high NO production upon stimulation with LPS, no significant differences were seen in the NO production of the RM obtained following the different induction treatments. Our results point out that care should be taken when applying elicited RM for in vitro assays, as distinct levels of oxygenation activity were obtained using different induction protocols.
Insights
Different induction protocols significantly impact turkey respiratory macrophage oxygenation activity but not nitric oxide production. Careful selection of induction methods is crucial for reliable in vitro assays using these cells.
Area of Science:
- Immunology
- Cell Biology
- Veterinary Science
Background:
- Turkey respiratory macrophages (RM) are crucial for innate immunity.
- Understanding how induction protocols affect RM function is vital for research.
Purpose of the Study:
- To investigate the influence of various induction protocols on turkey RM recovery, oxygenation activity, and nitric oxide (NO) production.
- To determine the optimal induction method for eliciting turkey RM for in vitro studies.
Main Methods:
- Respiratory macrophages were elicited in turkeys using Sephadex G-50, Thioglycollate broth, IFA-M. hyorhinis, or IFA-Broth.
- RM were recovered via lung lavage and purified using Percoll gradients.
- Oxygenation activity was assessed via luminol-enhanced chemiluminescence assays after Zymosan A stimulation.
- Nitric oxide production was measured after LPS stimulation.
Main Results:
- No significant differences in RM recovery numbers were observed between protocols.
- Sephadex G-50 and IFA-Broth elicited RM demonstrated significantly higher oxidative burst responses compared to Thioglycollate and IFA-M. hyorhinis.
- All elicited RM produced substantial NO upon LPS stimulation, with no significant inter-protocol differences.
Conclusions:
- Induction protocols significantly influence the oxygenation activity of elicited turkey RM.
- The choice of induction method is critical for standardizing RM function in in vitro assays.
- Further research may optimize protocols for specific experimental needs.