Abstract
Objective and design
This study exploits the biological activity of interleukin (IL)-3 to generate high yields of peritoneal mast cells ex vivo in order to examine pro-inflammatory immune responses in ex-vivo culture.
Material or subjects
Mast cells were obtained from the peritoneal cavity of C57BL/6 mice.
Treatment
Mice were injected intraperitoneally twice per day for 5 days with IL-3 (40–50 μg/ml) to increase mast cell numbers.
Methods
Histological studies examined mast cell numbers in the peritoneal cavity, intestine, lung, spleen and skeletal muscle. Peritoneal mast cells cultured ex vivo (PCMCs) were stimulated for 24 h with lipopolysaccharide and Bordetella pertussis antigen and secretion of tumour necrosis factor-α, IL-6, IL-4, IL-5, IL-10 and interferon-γ into supernatant was measured by commercial ELISA. Cell surface marker expression of FcεRI, c-kit, OX40L and TLR2 was measured by flow cytometry. Mast cell degranulation was measured using a β-hexosaminidase assay.
Results
IL-3 treatment increases mast cell numbers in the peritoneal cavity, spleen and muscle but not intestine and lung of C57BL/6 mice. PCMCs generated from IL-3-treated mice exhibit impaired growth, differentiation and responses to activation as measured by decreased cytokine secretion and cell surface marker expression.
Conclusion
Mast cells cultured from IL-3-treated mice show impaired responses.
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Acknowledgments
We would like to thank Eric Callaghan for providing facilities for microscopic work at School of Biology and Environmental Science, University College Dublin, Ireland. We also would like to thank the staff of the Bioresource Unit, at Dublin City University, Ireland, and Sina Heydrich at Charité, Germany, for excellent technical assistance. This project was supported in part by the Ph.D. research and training programme in Target-driven Therapeutic and Theranostics. This programme is being financed by the Higher Education Authority as part of a National Graduate Training Programme in Biopharmaceuticals and Pharmacological Sciences in partnership with University College Dublin, University College Cork, Trinity College Dublin and Queens University Belfast. We also would like to acknowledge support by the COST Action BM1007 “Mast cells and basophils—Targets for innovative therapies”. Tamás Visnovitz was supported by the Irish Research Council for Science, Engineering and Technology (IRCSET).
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Vukman, K.V., Visnovitz, T., Adams, P.N. et al. Mast cells cultured from IL-3-treated mice show impaired responses to bacterial antigen stimulation. Inflamm. Res. 61, 79–85 (2012). https://doi.org/10.1007/s00011-011-0394-6
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DOI: https://doi.org/10.1007/s00011-011-0394-6