Molecular Mechanism underlying Eosinophil Differentiation in Bronchial Asthma

支气管哮喘嗜酸性粒细胞分化的分子机制

• 批准号: 13670591
• 负责人: IWAMOTO Itsuo
• 金额: $ 2.56万
• 依托单位: Chiba University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13670591/
• 关键词: Eosinophils; IL-5 receptor α chain; One hybrid screening; retrovirus-mediated expression cloning; bronchial asthma; transcription factor; IL-5レセプター; プロモーター; 発現制御; one-hybridスクリーニング; レトロウイルス

Allergic late-phase reactions provoked by specific antigens are associated with intense eosinophil infiltration into the site of antigen administration. Because eosinophils contain a large amount of cytotoxic proteins in their granules and is believed to be involved in the pathogenesis of allergic inflammation, the downregulation of eosinophil differentiation is considered to be important for the inhibition of allergic inflammation. While it is well known that IL-5 plays a critical role in the eosinophil development, it is unknown about the molecular mechanisms of IL-5-induced eosinophil development. Recently, it has been demonstrated that, using IL-5Rα transgenic mice, IL-5 does not transduce a specific signal for the eosinophil development but simply transduces a proliferative signal to eosinophil progenitors that express IL-5Rα specifically. Therefore, it is essential to understand the regulatory mechanisms of IL-5Rα expression in eosinophils to prevent allergic inflammation.To determine the molecular mechanism underlying the cell lineage-specific expression of IL-5Rα in eosinophils, we developed a retrovirus-mediated mammalian one hybrid system. In this system, BaF3 cells (Thy1.2) were transfected with Thy1.1 cDNA under the control of IL-5Rα promoter (BaF3-P1-Thy1.1 cells) and then BaF3-P1-Thy1.1 cells were retrovirally transfected with cDNA library of an eosinophilic cell line (AML14.3D10 cells). After 2 weeks of culture, Thy1.1-positive cells were purified by magnetic cell sorting and cDNAs in the retrovirus vector in the Thy1.1-positive cells were isolated by PCR. By two cycles of the screening, we isolated 25 cDNAs. Now, we are performing the experiments to confirm the ability of cDNAs to induce the expression of IL-5Rα. In the near future, we believe that genes that regulate the eosinophil development can be isolated from the cDNAs and that the mechanism of eosinophil development is uncovered.

由特异性抗原引起的过敏性晚期反应与抗原给药部位的嗜酸性粒细胞浸润有关。由于嗜酸性粒细胞在其颗粒中含有大量的细胞毒性蛋白，并且被认为参与过敏性炎症的发病机制，因此嗜酸性粒细胞分化的下调被认为对于抑制过敏性炎症是重要的。虽然众所周知IL-5在嗜酸性粒细胞发育中起关键作用，但IL-5诱导的嗜酸性粒细胞发育的分子机制尚不清楚。近年来，在IL-5 R α转基因小鼠中发现，IL-5并不为嗜酸性粒细胞的发育提供特异性信号，而只是将增殖信号转导给特异性表达IL-5 R α的嗜酸性粒细胞祖细胞。因此，了解IL-5 R α在嗜酸性粒细胞中表达的调控机制对预防过敏性炎症的发生具有重要意义。在此系统中，将IL-5 R α启动子控制的Thy1.1 cDNA转染BaF 3细胞（Thy1.2）（BaF 3-P1-Thy1.1细胞），然后将嗜酸性粒细胞系（AML 14.3D10细胞）的cDNA文库逆转录病毒转染BaF 3-P1-Thy1.1细胞。培养2周后，通过磁性细胞分选纯化Thy1.1阳性细胞，并通过PCR分离Thy1.1阳性细胞中逆转录病毒载体中的cDNA。经过两轮筛选，共分离到25个cDNA。目前，我们正在进行实验以证实cDNA诱导IL-5 R α表达的能力。在不久的将来，我们相信可以从cDNA中分离出调控嗜酸性粒细胞发育的基因，并揭示嗜酸性粒细胞发育的机制。

项目成果

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Suto A、Nakajima H、Hirose K、Suzuki K、Saito Y、Iwamoto I：“Interleukin-21 通过抑制 IL-4 刺激的 B 细胞种系 Cε 转录来防止抗原诱导的 IgE 产生”Blood。 2002）

Seto Y, Nakajima H, Suto A, Saito Y, Iwamoto I.: "Enhanced Th2 cell-mediated allergic inflammation in Tyk2-deficient mice"J.Immunol.. 170. 1077-1083 (2003)

Seto Y、Nakajima H、Suto A、Saito Y、Iwamoto I.：“Tyk2 缺陷小鼠中 Th2 细胞介导的过敏性炎症增强”J.Immunol.. 170. 1077-1083 (2003)

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