The release of basogranulin in response to IgE-dependent and IgE-independent stimuli: validity of basogranulin measurement as an indicator of basophil activation.
BACKGROUND
Basogranulin, the novel basophil granule protein recognized by the monoclonal antibody BB1, can be released by stimulation with anti-IgE antibody or calcium ionophore. However, the kinetics and regulation of its secretion are unknown.


OBJECTIVE
We quantified basogranulin and histamine release in response to a range of stimuli to assess whether basogranulin secretion is a reliable marker of basophil activation.


METHODS
Isolated peripheral blood basophils were stimulated with anti-IgE antibody, calcium ionophore, N -formyl-Met-Leu-Phe, and complement C5a. The released basogranulin and histamine were quantified by dot blotting with BB1 and a fluorometric method, respectively. Basogranulin localization was confirmed by flow cytometry.


RESULTS
Both basogranulin and histamine displayed a bell-shaped response curve when basophils were challenged with anti-IgE. Half-maximal release occurred within 30 seconds. Basogranulin levels were maximal by 15 minutes, whereas those for histamine continued increasing to 30 minutes. Wortmannin, a PI3-K inhibitor, suppressed the release of both mediators. Basophils from donors with the "nonreleaser" phenotype secreted neither mediator in response to anti-IgE. Non-IgE-dependent stimuli released both mediators in parallel in a concentration-dependent manner. The correlation between the relative amounts of each mediator released was highly significant (r =.901, P <.0001, n = 87). Flow cytometry revealed that some of the secreted basogranulin adhered to the cell surface.


CONCLUSIONS
Basogranulin is secreted along with histamine in response to both FcepsilonR I-related and unrelated stimuli. It is therefore a valid marker of basophil activation and could provide the basis for an immunoassay that distinguishes between basophil and mast cell activation.
Mochizuki
Akinori
A
Immunopharmacology Group, Southampton General Hospital.
McEuen
Alan R
AR
Buckley
Mark G
MG
Walls
Andrew F
AF
D000888:Antibodies, Anti-Idiotypic / Q000494:pharmacology; D001491:Basophils / Q000737:chemistry / Q000502:physiology*; D001798:Blood Proteins / Q000032:analysis / Q000378:metabolism*; D005434:Flow Cytometry; D006636:Histamine Release; D006801:Humans; D007073:Immunoglobulin E / Q000502:physiology*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D017455:Receptors, IgE / Q000502:physiology
D000888:Antibodies, Anti-Idiotypic; D001798:Blood Proteins; D017455:Receptors, IgE; C026408:anti-IgE antibodies; D007073:Immunoglobulin E; D009240:N-Formylmethionine Leucyl-Phenylalanine; D019869:Phosphatidylinositol 3-Kinases