Changing expression of IL-3 and IL-5 receptors in cultured human eosinophils q Chitose Yoshimura-Uchiyama, a,b Masao Yamaguchi, a Hiroyuki Nagase, a Kouji Matsushima, c Takashi Igarashi, b Tsutomu Iwata, b Kazuhiko Yamamoto, a and Koichi Hirai d, * a Department of Allergy and Rheumatology, University of Tokyo Graduate School of Medicine, Tokyo, Japan b Department of Pediatrics, University of Tokyo Graduate School of Medicine, Tokyo, Japan c Department of Molecular Preventive Medicine, University of Tokyo Graduate School of Medicine, Tokyo, Japan d Department of Bioregulatory Function, University of Tokyo Graduate School of Medicine, Tokyo, Japan Received 24 July 2003 Abstract IL-3, IL-5, and GM-CSF exert overlapping functions in eosinophils via a shared receptor b-chain, and IL-3Ra transcript ex- pression is the weakest in blood eosinophils. We investigated the long-term regulation of surface expression of IL-3Ra. IL-3 was the most potent inducer of CD69 expression after 24-h stimulation, but not after 1-h stimulation. Expression of IL-5Ra and GM- CSFRa was significantly downregulated by culturing with their respective ligands, while IL-3Ra expression was not. IL-3 at 30 pM significantly increased IL-3Ra expression and IL-3Ra expression was also upregulated by both IL-5 and GM-CSF. In parallel with the surface protein expression, IL-3Ra mRNA was also upregulated by IL-3, IL-5, and GM-CSF. These results demonstrated that long-term culturing of eosinophils with CSFs induced a change in the potency order of CSFs, with IL-3 coming to exert the strongest effect. They thus suggest that IL-3 plays more important roles in local eosinophil activation than previously recognized. Ó 2003 Elsevier Inc. All rights reserved. Keywords: Eosinophils; IL-3; IL-5; GM-CSF; CD69; HLA-DR; Kinetics; Receptor Massive eosinophil infiltration is especially prominent at the sites of inflammation associated with allergic diseases. Eosinophils promote allergic inflammation, at least in part, through the release of an array of histo- toxic mediators including several granule-associated proteins [1]. Cytokines are thought to play critical roles in allergic inflammation, in view of their abilities to modulate immune responses within the allergic inflam- matory cell network. Especially eosinophil-directed he- mopoietins, i.e., IL-3, IL-5, and GM-CSF, have been strongly implicated in the pathogenesis of eosinophilic inflammation [2]. In addition to their induction of pro- liferation of progenitors of eosinophil lineage, these colony-stimulating factors (CSFs) stimulate terminally differentiated eosinophils to enhance their various bio- logical functions, which include mediator release, adhesion molecule expression, and longevity [2]. The receptors for IL-3, IL-5, and GM-CSF all possess a heterodimeric structure having a distinct a-chain but sharing a common b-chain (bc) [3]. Each CSF binds specifically to its corresponding receptor a-chain but does not interact directly with the bc. However, bc is fully responsible for transducing intracellular signals, thereby resulting in the overlapping of biological func- tions exhibited by the three cytokines [3,4]. On the other hand, the expression levels of the a-chains determine the biological potency of each CSF. We recently demon- strated that the expression order of a-chain transcripts was IL-5Ra P GM-CSFRa > IL-3Ra in freshly isolated eosinophils: eosinophils express almost equivalent levels of IL-5Ra and GM-CSFRa, but IL-3Ra was the least Biochemical and Biophysical Research Communications 309 (2003) 26–31 www.elsevier.com/locate/ybbrc BBRC q Abbreviations: CSF, colony-stimulating factor; bc, common b-chain; MESF, molecules of equivalent soluble fluorochrome units. * Corresponding author. Fax: +81-3-3815-5954. E-mail address: hiraiko-tky@umin.ac.jp (K. Hirai). 0006-291X/$ - see front matter Ó 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0006-291X(03)01526-2