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Published on: November 1, 2014
Intracellular expression and release of Fc epsilon RI alpha by human eosinophils
M C Seminario1, S S Saini, D W MacGlashan
1Department of Medicine, Division of Clinical Immunology, Johns Hopkins Asthma and Allergy Center, Baltimore, MD 21224, USA.
Insights
Human eosinophils have low surface levels of IgE receptors (Fc epsilon RI). Instead, they store Fc epsilon RI alpha intracellularly and release it into surrounding fluids, suggesting a novel immune function.
Area of Science:
- Immunology
- Cell Biology
- Allergy Research
Background:
- Fc epsilon Receptors (Fc epsilon R) are crucial in allergic responses.
- Previous studies showed conflicting data on Fc epsilon R presence on human eosinophils.
Purpose of the Study:
- To clarify the presence and levels of IgE, Fc epsilon RI, and Fc epsilon RII on human eosinophils.
- To investigate the regulation of Fc epsilon R expression on eosinophils.
Main Methods:
- Immunofluorescence and flow cytometry were used to analyze cell surface receptors.
- Immunoprecipitation and Western blotting were employed on eosinophil lysates and supernatants.
- Surface biotinylation was performed to assess cell surface receptor expression.
Main Results:
- Little to no surface IgE or IgE receptors were detected on eosinophils.
- Culturing eosinophils did not induce detectable surface Fc epsilon R.
- Fc epsilon RI alpha was detected intracellularly and released into the supernatant.
Conclusions:
- Human eosinophils possess an intracellular pool of Fc epsilon RI alpha that can be released.
- Surface levels of Fc epsilon RI alpha are undetectable on eosinophils, even in allergic conditions.
- The biological significance of soluble Fc epsilon RI alpha from eosinophils requires further investigation.
Abstract:
Although Fc epsilon R have been detected on human eosinophils, levels varied from moderate to extremely low or undetectable depending on the donor and methods used. We have attempted to resolve the conflicting data by measuring levels of IgE, Fc epsilon RI, and Fc epsilon RII in or on human eosinophils from a variety of donors (n = 26) and late-phase bronchoalveolar lavage fluids (n = 5). Our results demonstrated little or no cell surface IgE or IgE receptors as analyzed by immunofluorescence and flow cytometry. Culture of eosinophils for up to 11 days in the presence or absence of IgE and/or IL-4 (conditions that enhance Fc epsilon R on other cells) failed to induce any detectable surface Fc epsilon R. However, immunoprecipitation and Western blot analysis of eosinophil lysates using mAb specific for Fc epsilon RI alpha showed a distinct band of approximately 50 kDa, similar to that found in basophils. Western blotting also showed the presence of FcR gamma-chain, but no Fc epsilon RI beta. Surface biotinylation followed by immunoprecipitation again failed to detect surface Fc epsilon RI alpha, although surface FcR gamma was easily detected. Since we were able to detect intracellular Fc epsilon RI alpha, we examined its release from eosinophils. Immunoprecipitation and Western blotting demonstrated the release of Fc epsilon RI alpha into the supernatant of cultured eosinophils, peaking at approximately 48 h. We conclude that eosinophils possess a sizable intracellular pool of Fc epsilon RI alpha that is available for release, with undetectable surface levels in a variety of subjects, including those with eosinophilia and elevated serum IgE. The biological relevance of this soluble form of Fc epsilon RI alpha remains to be determined.
