PDF(Pubmed)
Abstract：
BACKGROUND: TNFα-inducible matrix metalloproteinases play a critical role in the process of airway remodeling in respiratory inflammatory disease including asthma. The cationic host defense peptide LL-37 is elevated in the lungs during airway inflammation. However, the impact of LL-37 on TNFα-driven processes is not well understood. Here, we examined the effect of LL-37 on TNFα-mediated responses in human bronchial epithelial cells (HBECs).
METHODS: We used a slow off-rate modified aptamer-based proteomics approach to define the HBEC proteome altered in response to TNFα. Abundance of selected protein candidates and signaling intermediates was examined using immunoassays, ELISA and Western blots, and mRNA abundance was examined by qRT-PCR.
RESULTS: Proteomics analysis revealed that 124 proteins were significantly altered, 12 proteins were enhanced by ≥2-fold compared to unstimulated cells, in response to TNFα. MMP9 was the topmost increased protein in response to TNFα, enhanced by ∼10-fold, and MMP13 was increased by ∼3-fold, compared to unstimulated cells. Furthermore, we demonstrated that LL-37 significantly suppressed TNFα-mediated MMP9 and MMP13 in HBEC. Mechanistic data revealed that TNFα-mediated MMP9 and MMP13 production is controlled by SRC kinase and that LL-37 enhances related upstream negative regulators, namely, phospho-AKT (T308) and TNFα-mediated TNFAIP3 or A20.
CONCLUSIONS: The findings of this study suggest that LL-37 may play a role in intervening in the process of airway remodeling in chronic inflammatory respiratory disease such as asthma.
METHODS: We used a slow off-rate modified aptamer-based proteomics approach to define the HBEC proteome altered in response to TNFα. Abundance of selected protein candidates and signaling intermediates was examined using immunoassays, ELISA and Western blots, and mRNA abundance was examined by qRT-PCR.
RESULTS: Proteomics analysis revealed that 124 proteins were significantly altered, 12 proteins were enhanced by ≥2-fold compared to unstimulated cells, in response to TNFα. MMP9 was the topmost increased protein in response to TNFα, enhanced by ∼10-fold, and MMP13 was increased by ∼3-fold, compared to unstimulated cells. Furthermore, we demonstrated that LL-37 significantly suppressed TNFα-mediated MMP9 and MMP13 in HBEC. Mechanistic data revealed that TNFα-mediated MMP9 and MMP13 production is controlled by SRC kinase and that LL-37 enhances related upstream negative regulators, namely, phospho-AKT (T308) and TNFα-mediated TNFAIP3 or A20.
CONCLUSIONS: The findings of this study suggest that LL-37 may play a role in intervening in the process of airway remodeling in chronic inflammatory respiratory disease such as asthma.
摘要：
背景：TNFα诱导的基质金属蛋白酶（MMP）在包括哮喘在内的呼吸道炎性疾病的气道重塑过程中起关键作用。阳离子宿主防御肽(CHDP)LL-37在气道炎症期间在肺中升高。然而,LL-37对TNFα驱动过程的影响尚不清楚。这里,我们检查了LL-37对人支气管上皮细胞（HBEC）中TNFα介导的反应的影响。
方法：我们使用了基于慢解离速率修饰的基于适体的蛋白质组学方法来定义响应TNFα而改变的HBEC蛋白质组。使用免疫测定法检查了选定的蛋白质候选物和信号传导中间体的丰度，ELISA和蛋白质印迹,通过qRT-PCR检测mRNA丰度。
结果：蛋白质组学分析显示124种蛋白质发生了显著改变，与未刺激的细胞相比，12种蛋白质增强了≥2倍，对TNFα的反应。MMP9是响应TNFα的最高增加蛋白，增加~10倍，MMP13增加~3倍，与未刺激的细胞相比。此外，我们证明LL-37在HBEC中显著抑制TNFα介导的MMP9和MMP13。机制数据显示，TNFα介导的MMP9和MMP13的产生受SRC激酶控制，LL-37增强相关的上游负调节因子，即磷酸化AKT（T308）和TNFα介导的TNFAIP3或A20。
结论：这项研究的结果表明，LL-37可能在慢性炎症性呼吸道疾病如哮喘的气道重塑过程中起干预作用。
方法：我们使用了基于慢解离速率修饰的基于适体的蛋白质组学方法来定义响应TNFα而改变的HBEC蛋白质组。使用免疫测定法检查了选定的蛋白质候选物和信号传导中间体的丰度，ELISA和蛋白质印迹,通过qRT-PCR检测mRNA丰度。
结果：蛋白质组学分析显示124种蛋白质发生了显著改变，与未刺激的细胞相比，12种蛋白质增强了≥2倍，对TNFα的反应。MMP9是响应TNFα的最高增加蛋白，增加~10倍，MMP13增加~3倍，与未刺激的细胞相比。此外，我们证明LL-37在HBEC中显著抑制TNFα介导的MMP9和MMP13。机制数据显示，TNFα介导的MMP9和MMP13的产生受SRC激酶控制，LL-37增强相关的上游负调节因子，即磷酸化AKT（T308）和TNFα介导的TNFAIP3或A20。
结论：这项研究的结果表明，LL-37可能在慢性炎症性呼吸道疾病如哮喘的气道重塑过程中起干预作用。
