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Published on: June 8, 2012
Masson Pine pollen (Pinus massoniana) activate HD11 chicken macrophages invitro
Daniel Brugger1, Belinda Wilhelm2, Benjamin Schusser3
1Institute of Animal Nutrition and Dietetics, Vetsuisse-Faculty, University of Zurich, Winterthurerstrasse 270, 8057, Zurich, Switzerland; Chair of Animal Nutrition, TUM School of Life Sciences Weihenstephan, Technical University of Munich, Liesel-Beckmann-Strasse 2, 85354, Freising, Germany.
Ethnopharmacological Relevance:
Masson Pine pollen (Pinus massoniana; MP) are used in Traditional Chinese Medicine to treat gut conditions. Early in vivo work supports this claim and suggests interaction of the material with the gastrointestinal immune system.
Aim Of The Study:
The present study tested if and how MP material activates HD11 chicken macrophages in vitro using material from different production sites and harvest years.
Material & Methods:
We applied twelve batches of MP from different Chinese production sites and harvest years. Materials were subjected to LAL tests (endotoxic activity), GC-MS (fatty acid analysis), and plate techniques (microbiological background, antimicrobial activity). Furthermore, HD11 chicken macrophages were challenged (6 h, 37 °C) with MP or LPS (E. coli O111:B4), respectively, to quantify nitric oxide (NO) production and immune gene expression (RT-qPCR).
Results:
MP material promoted strong signals in LAL tests and contained significant amounts of 3-hydroxydodecanoic acid and 3-hydroxymyristic acid, irrespective of processing, harvest year, or origin. The pollen material activated HD11 chicken macrophages, which was confirmed by spikes of NO release and k-means cluster analysis of TLR-signaling pathway gene expression data. Response of NO production to Log2-titration of MP and LPS-treated media was in any case linear and significant. The response was reduced by polymyxin-B (PMB) and the inhibition was twice as strong for LPS than MP. No or minor microbiological background was detected on the majority of MP samples. Three samples showed presence of spoilage microorganisms and Gram-negative bacteria, but this did not correlate to LAL data or bacterial DNA counts. No antimicrobial activity of MP was evident.
Conclusion:
Pollen of the Masson Pine activated HD11 chicken macrophages in vitro, which is likely partially due to a background of bacterial LPS associated with the pollen material. However, as most of the effect (appr. 80%) could not be blocked by PMB this is certainly due to other stimuli. We hypothesize that polysaccharides and oligosaccharides of the pollen matrix have the potential to interact with certain immune receptors presented on the plasma membrane of chicken macrophages.
Insights
Masson Pine pollen activates chicken macrophages in vitro, likely due to bacterial lipopolysaccharides (LPS) and other pollen components. Further research is needed to identify these specific immune-stimulating compounds.
Area of Science:
- Immunology
- Pharmacology
- Traditional Chinese Medicine
Background:
- Masson Pine pollen (Pinus massoniana; MP) is used in Traditional Chinese Medicine (TCM) for gut conditions.
- In vivo studies suggest MP interacts with the gastrointestinal immune system.
Purpose of the Study:
- To investigate the in vitro activation of HD11 chicken macrophages by MP.
- To analyze the influence of production site and harvest year on MP's immunomodulatory effects.
Main Methods:
- MP batches from various origins and years were analyzed for endotoxic activity (LAL test), fatty acids (GC-MS), and microbial contamination.
- HD11 chicken macrophages were stimulated with MP or lipopolysaccharide (LPS) to measure nitric oxide (NO) production and immune gene expression (RT-qPCR).
Main Results:
- MP activated HD11 chicken macrophages, evidenced by increased NO release and altered immune gene expression.
- MP contained significant levels of 3-hydroxydodecanoic acid and 3-hydroxymyristic acid.
- While bacterial LPS contributed to the effect, approximately 80% of MP's activity was independent of LPS, as shown by polymyxin-B (PMB) inhibition.
Conclusions:
- Masson Pine pollen activates chicken macrophages in vitro.
- Bacterial LPS is a partial activator, but other pollen-derived components, potentially polysaccharides and oligosaccharides, are major contributors.
- MP's immunomodulatory potential warrants further investigation into specific active constituents.
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