Interleukin-8 Gene Repression by Clarithromycin is mediated by AP-1 Binding Site in Human Bronchial Epithelial Cells

克拉霉素对白细胞介素 8 基因的抑制是由人支气管上皮细胞中 AP-1 结合位点介导的

• 批准号: 09670597
• 负责人: NAKAMURA Hidenori
• 金额: $ 1.86万
• 依托单位: YAMAGATA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09670597/
• 关键词: Interleukin-8; Bronchial Epithelial Cells; Macrolide Antibiotics; Promoter; Transcription Factors

Macrolide antibiotics are known to be effective for the treatment of chronic inflammatory airway diseases including diffuse panbronchiolitis, chronic bronchitis and bronchial asthma. Other than anti-microbial activities, macrolides have "anti-inflammatory" effects, such as the inhibition of cytokine production. In the present study, we investigated the effects of clarithromycin (CAM) on the interleukin-8 (IL-8) gene expression and its protein levels using a human bronchial epithelial cell line, BET-1A cells. Northern analyses demonstrated that CAM inhibited the tumor necrosis factor alpha (TNF)-induced IL-8 gene expression in dose and incubation time-dependent fashions. The half-life of IL-8 mRNA transcripts in TN F-treated BET-1A cells did not change with CAM.Transfection studies with BET-1A cells, using fusion genes composed of the 5'-flanking sequences of the IL-8 gene and a luciferase reporter gene, demonstrated a potent promoter activity in a 174.bp segment (-130 to +44 bp relative to the transcription start site). This segment includes AP-1 and NF-_<lambda>B-like sites, and exhibited a strongest response to TNF.TNF-induced promoter activity in this segment showed a significant repression by CAM.However, a 156-bp segment (-112 to +44 bp), which does not include an AP-1 site but includes a NF-_<lambda>B-like site, did not show a significant repression of TNF-induced promoter activity by CAM.Consistent with promoter analyses, an electrophoretic mobility shift assay demonstrated that CAM repressed the AR-1 binding in TNF-treated BET-1A cells, however, TNF induced both AP-1 and NF-_<lambda>B binding activities in BET-1A cells. These data suggest that macrolidessuch as CAM repress the IL-8 gene transcription mainly via the AP-1 binding site in human bronchial epithelial cells. Our findings provide a novel mechanism of the anti-inflammatory function of macrolides, implicating a target for the development of a new drug for the treatment of chronic airway inflammation.

已知大环内酯类抗生素可有效治疗慢性炎性气道疾病，包括弥漫性泛细支气管炎、慢性支气管炎和支气管哮喘。除了抗菌活性外，大环内酯类化合物还具有“抗炎”作用，如抑制细胞因子的产生。在本研究中，我们利用人支气管上皮细胞系β - 1a细胞研究了克拉霉素（CAM）对白细胞介素-8 （IL-8）基因表达及其蛋白水平的影响。Northern分析表明，CAM抑制肿瘤坏死因子α (TNF)诱导的IL-8基因表达，其剂量和孵育时间依赖于模型。在TN f处理的β - 1a细胞中，IL-8 mRNA转录物的半衰期未因CAM而改变。用IL-8基因5′侧序列和荧光素酶报告基因组成的融合基因转染β - 1a细胞，在174个细胞中显示出强大的启动子活性。Bp片段（相对于转录起始位点-130至+44 Bp）。该片段包括AP-1和NF-_<lambda> b样位点，对TNF表现出最强的反应。tnf诱导的该片段启动子活性受到CAM的显著抑制。然而，一个156 bp的片段（-112至+44 bp）不包括AP-1位点，但包括NF-_<lambda> b样位点，CAM对tnf诱导的启动子活性没有明显的抑制作用。与启动子分析一致，电泳迁移率转移实验表明，CAM抑制了TNF处理的β - 1a细胞中AR-1的结合，然而，TNF诱导β - 1a细胞中AP-1和NF-_< λ >B的结合活性。这些数据表明，大环内酯类药物如CAM主要通过人支气管上皮细胞AP-1结合位点抑制IL-8基因转录。我们的发现提供了大环内酯类抗炎功能的新机制，为开发治疗慢性气道炎症的新药提供了靶点。

项目成果

Kawasaki S,Takizawa H,Ohtoshi T,Takeuchi N,Kohyama T,Nakamura H,Kasama T,Kobayashi K,Kasahara K,Morita Y,Yamamoto K.: "Roxithromycin inhibits cytokine production by and neutrophil attachment to human bronchial epithelial cells in vitro." Antimicrob Agents

Kawasaki S、Takizawa H、Ohtoshi T、Takeuchi N、Kohyama T、Nakamura H、Kasama T、Kobayashi K、Kasahara K、Morita Y、Yamamoto K.：“罗红霉素在体外抑制人支气管上皮细胞的细胞因子产生和中性粒细胞附着

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Kawasaki S et al.: "Roxithromycin inhibits cytokine production and neutrophil attachment to human bronchial cells in vitro" Antimicrob Agents Chem. 42. 1499-1502 (1998)

Kawasaki S 等人：“罗红霉素在体外抑制细胞因子的产生和中性粒细胞与人支气管细胞的附着”Antimicrob Agents Chem。

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