Utility of mast cell P815 for evaluating FcεRI-dependent and FcεRI-independent allergic effects.

INTRODUCTION AND

OBJECTIVES

The degranulation and release of inflammatory mediators mediated by the high-affinity immunoglobulin E (IgE) receptor (FcεRI) on mast cells in response to allergen contact is the driving force of anaphylaxis. This study shows that P815 cells, which were previously thought not to express FcεRI, cause a reaction similar to FcεRI-mediated degranulation in the presence of antigen and IgE.

MATERIALS AND

METHODS

The kinetics of degranulation were evaluated by comparing P815 cells with FcεRI-expressing a rat basophilic leukemia (RBL-2H3) (that typically indicates the specific clone or subline within that cell line) cell lines. Degranulation activity was measured using the release rate of β-hexosaminidase as an indicator. P815 cells showed significant degranulation when compound 48/80 or anti-dinitrophenyl (DNP)-IgE antibody and DNP-human serum albumin (HSA) antigen were added simultaneously. Gene expression analysis confirmed the expression of each FcεRI subunit-specifically, the γ subunit expressed markedly. Moreover, the expression of the phosphorylation enzymes Lyn, spleen tyrosine kinase (Syk), Fyn, and Bruton tyrosine kinase (Btk), which are involved in degranulation, was upregulated.

RESULTS

FcεRI has three subunits: α, β, and γ. P815 cells do not express FcεRI because they have messenger ribonucleic acid (mRNA) for the γ subunit but not for the α and β subunits. However, P815 expressed each subunit protein (α, β, and γ), as detected in the western blotting analysis of cell extracts in the presence of DNP-HSA antigen and anti-DNP-IgE.

CONCLUSION

These results suggest that P815 may cause degranulation via FcεRI. Therefore, P815 is considered to be a cell model that can evaluate both FcεRI-mediated and FcεRI-independent degranulation reactions in response to allergens.