NLRP3 inflammasome activity is upregulated in an in-vitro model of COPD exacerbation

BackgroundChronic obstructive pulmonary disease (COPD) is an inflammatory disease characterized by a progressive and irreversible deterioration of lung function. Exacerbations of COPD have prolonged negative effects on pulmonary function and a major impact on health status and outcomes. NLRP3 inflammasome is a cardinal component of the inflammatory response, with marked evidence in stable and exacerbations of COPD. The aim of our study was to evaluate the NLRP3 inflammasome activity during COPD exacerbation by using an in vitro model.MethodsA549 cells were stimulated with different concentrations (10%, 4%, 2%) of cigarette smoke extract (CSE) with or without LPS (0.1μg/ml) for 24 hours. Cell viability was assessed by using XTT test. Levels of inflammatory cytokines (IL-8, MCP-1, and IL-1β) were measured by ELISA and the activity level of NLRP-3 was evaluated by flow cytometry.ResultsCells exposed to CSE present an increase in inflammatory cytokines (IL-8 and MCP-1) production in a dose-dependent manner. Incubation with LPS to these cells results in higher levels of IL-8 and MCP-1 compared to stimulation of CSE alone. NLRP3 inflammasome activity and IL-1β levels were significantly increased in cells exposed to both CSE and LPS compared to CSE alone.ConclusionsNLRP3 inflammasome is upregulated in an in-vitro model of COPD and COPD exacerbation. Our findings provide novel biomarkers for COPD exacerbation and may present new targets for future research.背景：慢性阻塞性肺疾病（Chronic obstructive pulmonary disease, COPD）是一类以肺功能进行性、不可逆退化为特征的炎性疾病。慢阻肺急性加重会对肺功能产生长期负面影响，并对健康状况与转归造成重大影响。NLRP3炎性小体（NLRP3 inflammasome）是炎性反应的核心组分，在慢阻肺稳定期与急性加重期均有明确相关证据。本研究旨在通过体外模型评估慢阻肺急性加重期的NLRP3炎性小体活性。 方法：将A549细胞以不同浓度（10%、4%、2%）的香烟烟雾提取物（cigarette smoke extract, CSE）进行刺激，分别联合或不联合脂多糖（lipopolysaccharide, LPS，0.1μg/ml），刺激时长为24小时。采用XTT试验检测细胞活力；通过酶联免疫吸附试验（ELISA）检测炎性细胞因子（IL-8、MCP-1及IL-1β）的水平；采用流式细胞术评估NLRP3的活性水平。 结果：经CSE刺激的细胞，其炎性细胞因子IL-8与MCP-1的生成量呈剂量依赖性升高。与仅使用CSE刺激相比，联合LPS处理的细胞中IL-8与MCP-1水平更高。相较于仅CSE刺激组，同时暴露于CSE与LPS的细胞中，NLRP3炎性小体活性与IL-1β水平均显著升高。 结论：在慢阻肺及慢阻肺急性加重的体外模型中，NLRP3炎性小体表达上调。本研究结果为慢阻肺急性加重提供了新型生物标志物，也可为未来相关研究提供全新的干预靶点。