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A comparative study of the FcepsilonRI molecule on human mast cell and basophil cell lines
B M Jensen1, S Dissing, P S Skov
1Allergy Clinic, National University Hospital, Copenhagen, Denmark. bettinamjensen@hotmail.com
Insights
Human mast cell line LAD2 and basophil KU812 subclones show differences in high-affinity IgE receptor (FcepsilonRI) regulation and activation. These cell lines are valuable tools for studying FcepsilonRI biology and its effects.
Area of Science:
- Immunology
- Cell Biology
Background:
- Mast cells and basophils express the high-affinity IgE receptor, FcepsilonRI.
- Investigating human mast cell line LAD2 and basophil KU812 subclones reveals differences in FcepsilonRI regulation and activation.
Purpose of the Study:
- To analyze FcepsilonRI surface regulation, anti-IgE-triggered activation, FcepsilonRIalpha protein stability, and mRNA levels.
- To determine the utility of LAD2 and KU812 cell lines in FcepsilonRI biology research.
Main Methods:
- Flow cytometry for FcepsilonRI surface expression.
- Intracellular Ca2+ measurement for cellular activation.
- Western blot and radioimmunoassay for FcepsilonRIalpha protein stability.
- Real-time PCR for mRNA levels of FcepsilonRIalpha, beta, and beta(T) chains.
Main Results:
- LAD2 and two KU812 subclones exhibited FcepsilonRI surface expression capable of inducing cellular activation.
- IgE presence increased FcepsilonRI expression and FcepsilonRIalpha protein stability.
- All cell lines expressed FcepsilonRIalpha, beta, and beta(T) mRNA, with LAD2 showing the highest expression; no difference in beta/beta(T) ratio was observed.
Conclusions:
- LAD2 and KU812 cell lines are important tools for investigating the FcepsilonRI molecule.
- These cell lines aid in studying the effects induced by FcepsilonRI activation.
Background:
Mast cells and basophils express the high-affinity IgE receptor FcepsilonRI. We have analysed the human mast cell line LAD2 and four subclones of the basophil cell line KU812 in order to reveal possible differences concerning the FcepsilonRI surface regulation, anti-IgE-triggered activation, FcepsilonRIalpha protein stability and the mRNA level of FcepsilonRIalpha-, beta- and the truncated beta-chain (beta(T)), and thereby determine the utility of these cell lines in investigations of the FcepsilonRI biology.
Methods:
The surface expression of FcepsilonRI was assessed by flow cytometry, using the monoclonal antibody CRA1. The FcepsilonRI-induced cellular activation (i.e. cross-linking of FcepsilonRI) was determined by changes in the intracellular level of Ca2+, which was measured by fluorescence of Fura-2. The level of the FcepsilonRIalpha protein was determined by a Western blot technique and by a radioimmunoassay. The mRNA level of FcepsilonRIalpha, beta- and beta(T)-chain was analysed using real-time PCR.
Results:
Two KU812 subclones and especially LAD2 had FcepsilonRI surface expression which was capable of inducing cellular activation. Both the FcepsilonRI expression and stability of the FcepsilonRIalpha protein were increased when IgE was present. All the cell lines expressed mRNA of FcepsilonRIalpha-, beta- and beta(T), with LAD2 tending to have the highest expression. However, a determination of the beta/beta(T) ratio demonstrated no difference between any of the cell clones.
Conclusion:
These cell lines are important tools in the investigation of both the FcepsilonRI molecule and the effects induced by its activation.
