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Abstract：
BACKGROUND: Steroid insensitivity in Chronic Obstructive Pulmonary Disease (COPD) presents a problem for controlling the chronic inflammation of the airways. The glucocorticoid receptor (GR) mediates the intracellular signaling of inhaled corticosteroids (ICS) by interacting with transcription factors and histone deacetylases (HDACs). The aim of this study was to assess if COPD patients\' response to ICS in vivo, may be associated with the expression of GR, the complex of GR with transcription factors, and the expression of various HDACs in vitro.
METHODS: Primary airway smooth muscle cells (ASMC) were established from endobronchial biopsies obtained from patients with asthma (n = 10), patients with COPD (n = 10) and subjects that underwent diagnostic bronchoscopy without pathological findings and served as controls (n = 6). ASMC were also established from 18 COPD patients, 10 responders and 8 non-responders to ICS, who participated in the HISTORIC study, an investigator-initiated and driven clinical trial that proved the hypothesis that COPD patients with high ASMC in their endobronchial biopsies respond better to ICS than patients with low ASMC. Expression of GR and its isoforms GRα and GRβ and HDACs was investigated in primary ASMC in the absence or in the presence of dexamethasone (10- 8M) by western blotting. The complex formation of GR with transcription factors was assessed by co-immunoprecipitation.
RESULTS: Expression of GR and its isoform GRα but not GRβ was significantly reduced in ASMC from COPD patients as compared to controls. There were no significant differences in the expression of GR, GRα and GRβ between responders and non-responders to ICS. However, treatment with dexamethasone upregulated the expression of total GR (p = 0.004) and GRα (p = 0.005) after 30 min in responders but not in non-responders. Τhe formation of the complex GR-c-Jun was increased 60 min after treatment with dexamethasone only in responders who exhibited significantly lower expression of HDAC3 (p = 0.005) and HDAC5 (p < 0.0001) as compared to non-responders.
CONCLUSIONS: These data suggest that ASMC from COPD patients who do not respond to treatment with ICS, are characterized by reduced GR-c-Jun complex formation and increased expression of HDAC3 and HDAC5.
BACKGROUND: ISRCTN11017699 (Registration date: 15/11/2016).
METHODS: Primary airway smooth muscle cells (ASMC) were established from endobronchial biopsies obtained from patients with asthma (n = 10), patients with COPD (n = 10) and subjects that underwent diagnostic bronchoscopy without pathological findings and served as controls (n = 6). ASMC were also established from 18 COPD patients, 10 responders and 8 non-responders to ICS, who participated in the HISTORIC study, an investigator-initiated and driven clinical trial that proved the hypothesis that COPD patients with high ASMC in their endobronchial biopsies respond better to ICS than patients with low ASMC. Expression of GR and its isoforms GRα and GRβ and HDACs was investigated in primary ASMC in the absence or in the presence of dexamethasone (10- 8M) by western blotting. The complex formation of GR with transcription factors was assessed by co-immunoprecipitation.
RESULTS: Expression of GR and its isoform GRα but not GRβ was significantly reduced in ASMC from COPD patients as compared to controls. There were no significant differences in the expression of GR, GRα and GRβ between responders and non-responders to ICS. However, treatment with dexamethasone upregulated the expression of total GR (p = 0.004) and GRα (p = 0.005) after 30 min in responders but not in non-responders. Τhe formation of the complex GR-c-Jun was increased 60 min after treatment with dexamethasone only in responders who exhibited significantly lower expression of HDAC3 (p = 0.005) and HDAC5 (p < 0.0001) as compared to non-responders.
CONCLUSIONS: These data suggest that ASMC from COPD patients who do not respond to treatment with ICS, are characterized by reduced GR-c-Jun complex formation and increased expression of HDAC3 and HDAC5.
BACKGROUND: ISRCTN11017699 (Registration date: 15/11/2016).
摘要：
背景：慢性阻塞性肺疾病（COPD）中的类固醇不敏感存在控制气道慢性炎症的问题。糖皮质激素受体（GR）通过与转录因子和组蛋白脱乙酰酶（HDAC）相互作用来介导吸入皮质类固醇（ICS）的细胞内信号传导。这项研究的目的是评估COPD患者对ICS的反应是否在体内，可能与GR的表达有关，GR与转录因子的复合物，以及各种HDAC在体外的表达。
方法：从哮喘患者（n=10）的支气管内活检中建立了原发性气道平滑肌细胞（ASMC），COPD患者(n=10)和接受诊断性支气管镜检查但无病理结果并作为对照的受试者(n=6).还从18例COPD患者中建立了ASMC，ICS的10名响应者和8名非响应者，参加历史研究的人，一项由研究者发起并驱动的临床试验证明了以下假设:支气管内活检中ASMC高的COPD患者对ICS的反应优于ASMC低的患者.在不存在或存在地塞米松（10-8M）的情况下，通过蛋白质印迹在原发性ASMC中研究了GR及其同工型GRα和GRβ和HDAC的表达。通过共免疫沉淀评估GR与转录因子的复合物形成。
结果：与对照组相比，COPD患者ASMC中GR及其同工型GRα而非GRβ的表达显著降低。GR的表达无显著差异，ICS应答者和非应答者之间的Grα和Grβ。然而,在30分钟后，地塞米松治疗上调了应答者的总GR（p=0.004）和GRα（p=0.005）的表达，但在非应答者中没有。复合GR-c-Jun的形成在用地塞米松治疗60分钟后增加,仅在与无反应者相比表现出显著较低的HDAC3(p=0.005)和HDAC5(p<0.0001)表达的反应者中。
结论：这些数据表明，对ICS治疗无反应的COPD患者的ASMC，其特征在于减少的GR-c-Jun复合物形成和增加的HDAC3和HDAC5的表达。
背景：ISRCTN11017699（注册日期：2016年11月15日）。
方法：从哮喘患者（n=10）的支气管内活检中建立了原发性气道平滑肌细胞（ASMC），COPD患者(n=10)和接受诊断性支气管镜检查但无病理结果并作为对照的受试者(n=6).还从18例COPD患者中建立了ASMC，ICS的10名响应者和8名非响应者，参加历史研究的人，一项由研究者发起并驱动的临床试验证明了以下假设:支气管内活检中ASMC高的COPD患者对ICS的反应优于ASMC低的患者.在不存在或存在地塞米松（10-8M）的情况下，通过蛋白质印迹在原发性ASMC中研究了GR及其同工型GRα和GRβ和HDAC的表达。通过共免疫沉淀评估GR与转录因子的复合物形成。
结果：与对照组相比，COPD患者ASMC中GR及其同工型GRα而非GRβ的表达显著降低。GR的表达无显著差异，ICS应答者和非应答者之间的Grα和Grβ。然而,在30分钟后，地塞米松治疗上调了应答者的总GR（p=0.004）和GRα（p=0.005）的表达，但在非应答者中没有。复合GR-c-Jun的形成在用地塞米松治疗60分钟后增加,仅在与无反应者相比表现出显著较低的HDAC3(p=0.005)和HDAC5(p<0.0001)表达的反应者中。
结论：这些数据表明，对ICS治疗无反应的COPD患者的ASMC，其特征在于减少的GR-c-Jun复合物形成和增加的HDAC3和HDAC5的表达。
背景：ISRCTN11017699（注册日期：2016年11月15日）。
