A functional study on CysLT(1) receptors in human eosinophils.
BACKGROUND
The cysteinyl leukotrienes (CysLTs) mediate their biological actions through two receptors: CysLT(1) receptor and CysLT(2) receptor.


OBJECTIVE
This study was undertaken to examine the direct effects of CysLTs on eosinophils, such as chemotaxis and degranulation, focusing on CysLT(1).


METHODS
Eosinophils were isolated from venous blood from normal volunteers who had no history of allergy (purity >99%). They were subjected to reverse transcription-PCR analysis and flow-cytometric analysis for CysLT(1). Binding assays were performed with [(3)H]LTD(4). Purified eosinophils loaded with Fura-2 acetoxymethyl ester were stimulated with CysLTs, and Ca(2+) influx was measured. Eosinophil migration in response to CysLTs was measured using a 96-well multiwell Boyden chamber. Eosinophils were treated with LTD(4) at 10(-6) M for 60 min followed by incubation for 4 h at 37 degrees C in the presence or absence of IL-5 and eosinophil-derived neurotoxin (EDN) release was evaluated.


RESULTS
The expression of the mRNA and protein of CysLT(1) on eosinophils and [(3)H]LTD(4)-specific binding to eosinophils were observed. Neither Th1 cytokine (IFN-gamma) nor Th2 cytokines (IL-4 or IL-5) affected CysLT(1) expression in eosinophils. CysLTs induced an increase in intracellular free Ca(2+) in eosinophils via CysLT(1), as suggested by the efficient inhibition by a CysLT(1) antagonist, pranlukast, in addition to the rank order of potency being LTD(4), LTC(4) and LTE(4). LTD(4) stimulated eosinophils to migrate at 10(-6) M via CysLT(1). LTE(4) also induced significant eosinophil migration at 10(-6) M. LTD(4) enhanced EDN release induced by IL-5 via CysLT(1).


CONCLUSIONS
CysLTs induce migration and enhance degranulation in eosinophils via CysLT(1). Accordingly, interaction of CysLTs and CysLT(1) on eosinophils has the potential to play a prominent role in the pathophysiology of asthma.
Ohshima
Nobuharu
N
Department of Respiratory Medicine, University of Tokyo Graduate School of Medicine, Tokyo, Japan.
Nagase
Hiroyuki
H
Koshino
Takeshi
T
Miyamasu
Misato
M
Yamaguchi
Masao
M
Hirai
Koichi
K
Yamamoto
Kazuhiko
K
Fujisawa
Takao
T
Nakagawa
Naoki
N
Kishikawa
Katsuya
K
Morita
Yutaka
Y
D002118:Calcium / Q000378:metabolism; D015550:Cell Degranulation; D002465:Cell Movement / Q000187:drug effects; D002633:Chemotaxis / Q000187:drug effects; D003545:Cysteine / Q000494:pharmacology; D016207:Cytokines / Q000494:pharmacology; D004804:Eosinophils / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D017998:Leukotriene D4 / Q000378:metabolism; D015289:Leukotrienes / Q000494:pharmacology; D008565:Membrane Proteins; D012333:RNA, Messenger / Q000032:analysis; D018077:Receptors, Leukotriene / Q000235:genetics / Q000502:physiology*
D016207:Cytokines; D015289:Leukotrienes; D008565:Membrane Proteins; D012333:RNA, Messenger; D018077:Receptors, Leukotriene; C112381:cysteinyl-leukotriene; D017998:Leukotriene D4; C412375:cysteinyl leukotriene receptor 2; D003545:Cysteine; C083169:leukotriene D4 receptor; D002118:Calcium