Identification of messenger RNA for IL-4 in human eosinophils with granule localization and release of the translated product.
Human eosinophils are cytokine-producing cells that are prominent in IgE-dependent allergic tissue reactions. IL-4 promotes the development of the Th2-type phenotype in T cells and is an essential cofactor for IgE production by B cells. We detected mRNA for IL-4 by reverse transcription-PCR in blood eosinophils from atopic asthmatics. By specific ELISA, 108 +/- 20 pg of IL-4 protein/10(6) cells could be extracted from whole cells, and approximately 30% of the IL-4 was released after incubation with serum-coated particles. Using immunocytochemistry, eosinophils from atopic asthmatics and nonatopic controls showed IL-4 immunoreactivity using an anti-IL-4 mAb. IL-4 was located predominantly in the eosinophil granules, as shown by both immunogold electron microscopy and a cell fractionation technique that dissociated cell granules from membrane and cytosolic components. IL-4 mRNA colocalized with eosinophils (using sequential immunocytochemistry with an eosinophil-specific (EG2) mAb and in situ hybridization using an IL-4-specific antisense riboprobe) in both cell cytospins from bronchoalveolar lavage fluid from asthmatics as well as skin biopsies obtained from allergen-induced late phase (6-h) reactions in atopic subjects. Using double immunocytochemistry on skin biopsies with eosinophil- and IL-4-specific mAb, 83.5 +/- 3.5% of eosinophils were IL-4+. Conversely, eosinophils accounted for 46.5 +/- 3.9% of the total cells expressing IL-4 immunoreactivity. Thus, human eosinophils express mRNA for IL-4, and the translated product is contained within the crystalloid granule from which it is released after stimulation with serum-coated particles. These observations are consistent with the hypothesis that eosinophils contribute to the development of the Th2 phenotype by T cells infiltrating atopic allergic reactions as well as to IgE synthesis.
Moqbel
R
R
Department of Allergy and Clinical Immunology, National Heart and Lung Institute, Royal Brompton Hospital, London, United Kingdom.
Ying
S
S
Barkans
J
J
Newman
T M
TM
Kimmitt
P
P
Wakelin
M
M
Taborda-Barata
L
L
Meng
Q
Q
Corrigan
C J
CJ
Durham
S R
SR
Kay
A B
AB
D001249:Asthma / Q000378:metabolism; D001483:Base Sequence; D002478:Cells, Cultured; D003594:Cytoplasmic Granules / Q000378:metabolism* / Q000648:ultrastructure; D004804:Eosinophils / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D015847:Interleukin-4 / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis* / Q000648:ultrastructure; D013347:Subcellular Fractions
D012333:RNA, Messenger; D015847:Interleukin-4