| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2002: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 2001: ¥1,900,000 (Direct Cost: ¥1,900,000)
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| Research Abstract |
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 [^3H]LTD_4. Purified eosinophils loaded with Fura-2-acetoxymethyl ester was 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℃ in the presence or absence of IL-5, and EDN release was evaluated.
Results : The expression of the mRNA and protein of CysLT_1 on eosinophils, and [^3H]LTD_4-specific binding to eosinophils, were observed. Neither Th1 cytokine (IFN-g) 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.
Conclusion : CysLTs induce migration and enhance degranulation in eosinophils via CysLT_1. Accordingly, interaction of CysLTs and CysLT_1 on eosinophils has potential to play a prominent role in the pathophysiology of asthma.
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