Thymic stromal lymphopoietin promotes asthmatic airway remodelling in human lung fibroblast cells through STAT3 signalling pathway
Thymic stromal lymphopoietin promotes asthmatic airway remodelling in human lung fibroblast cells through STAT3 signalling pathway
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DOI：
10.1002/cbf.2926
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发表时间：
2013-08-01
影响因子：
3.6
通讯作者：
Tian, Hong
中科院分区：
文献类型：
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作者：
Wu, Jinxiang;Liu, Fen;Tian, Hong
本研究旨在确定胸腺基质淋巴生成素（TSLP）在哮喘气道重构中的作用和调控。为了鉴定支气管组织中TSLP、平滑肌肌动蛋白（-SMA）和I型胶原蛋白的表达，分别采集哮喘患者和健康对照者的支气管活检标本，并进行特异性抗体染色。为了研究TSLP调控的信号通路，我们通过shRNA方法或转染方法在人肺成纤维细胞（HLF-1）中沉默或过表达TSLP，并通过酶联免疫吸附试验和Western blot分析检测TSLP受体（TSLPR）的表达。在TSLP信号通路中，Western blotting检测总信号换能器和转录激活因子3 （STAT3）、STAT5、STAT3磷酸化蛋白（pSTAT3）和STAT5磷酸化蛋白（pSTAT5）、TSLP、-SMA和I型胶原蛋白的表达。实时反转录法测定-SMA、I型胶原蛋白和mRNA的表达。为了进一步确认TSLP-STAT3信号通路在HLF-1细胞中的作用，我们通过靶向小分子抑制STAT3活性，然后分别通过实时定量反转录和Western blotting检测tslp诱导-SMA和I型胶原在mRNA和蛋白水平上的表达。首先，在哮喘患者的上皮中检测到TSLP、-SMA和I型胶原蛋白的过表达。其次，通过TSLP调控STAT3活性、-SMA和I型胶原蛋白的表达。具体来说，在HLF-1细胞中引入TSLP诱导pSTAT3、-SMA和胶原蛋白I， TSLP沉默后检测到-SMA和胶原蛋白I的抑制。第三，在STAT3抑制剂存在的情况下，pSTAT5没有变化，tslp诱导的-SMA和I型胶原蛋白上调是STAT3依赖性的。如果我们通过STAT3靶向小分子抑制STAT3活性，则无法检测到tslp诱导的-SMA和I型胶原蛋白上调。TSLP在哮喘气道重构中的作用通过STAT3信号通路实现。版权所有：John Wiley & Sons， Ltd。
This study aimed to identify the role and regulation of thymic stromal lymphopoietin (TSLP) in asthmatic airway remodelling. To identify the expression of TSLP, smooth muscle actin (-SMA) and collagen I in bronchial tissues, bronchial biopsy specimens were collected from patients with asthma and healthy controls and stained with specific antibodies, respectively. To characterize the signalling pathways regulated by TSLP, we silenced or overexpressed TSLP in human lung fibroblast (HLF-1) cells by shRNA approaches or transfection and detected the expression of TSLP receptor (TSLPR) by enzyme-linked immunosorbent assay and Western blot analysis. In TSLP signalling pathway, the protein expression of total signal transducer and activator of transcription 3 (STAT3), STAT5, the phosphorylation of STAT3 (pSTAT3) and STAT5 (pSTAT5), TSLP, -SMA and collagen I were also detected by Western blotting. In addition, the -SMA, collagen I and mRNA expression were determined by real-time reverse-transcription. To further confirm the TSLP-STAT3 signalling pathway in HLF-1 cells, we inhibited STAT3 activity by targeted small molecules and then detected TSLP-induced expression of -SMA and collagen I in both mRNA and protein levels by quantitative real-time reverse-transcription and Western blotting, respectively. First, overexpression of TSLP, -SMA and collagen I was detected in epithelium collected from patients with asthma. Second, STAT3 activity and the expression of -SMA and collagen I were controlled, regulated by TSLP. Specifically, the pSTAT3, -SMA and collagen I were induced by the introduction of TSLP in HLF-1 cells, and the repression of -SMA and collagen I was detected after TSLP silencing. Third, no changes of pSTAT5 were found in the presence of the STAT3 inhibitor, and TSLP-induced -SMA and collagen I upregulation is in a STAT3 dependent manner. If we inhibit STAT3 activity by STAT3 targeted small molecules, TSLP-induced -SMA and collagen I upregulation cannot be detected. The functions of TSLP in asthmatic airway remodelling were performed through STAT3 signalling pathway. Copyright (c) 2012 John Wiley & Sons, Ltd.