PDF(Pubmed)
Abstract：
The evaluation of inhalation toxicity, drug safety and efficacy assessment, as well as the investigation of complex disease pathomechanisms, are increasingly relying on in vitro lung models. This is due to the progressive shift towards human-based systems for more predictive and translational research. While several cellular models are currently available for the upper airways, modelling the distal alveolar region poses several constraints that make the standardization of reliable alveolar in vitro models relatively difficult. In this work, we present a new and reproducible alveolar in vitro model, that combines a human derived immortalized alveolar epithelial cell line (AXiAEC) and organ-on-chip technology mimicking the lung alveolar biophysical environment (AXlung-on-chip). The latter mimics key features of the in vivo alveolar milieu: breathing-like 3D cyclic stretch (10% linear strain, 0.2 Hz frequency) and an ultrathin, porous and elastic membrane. AXiAECs cultured on-chip were characterized for their alveolar epithelial cell markers by gene and protein expression. Cell barrier properties were examined by TER (Transbarrier Electrical Resistance) measurement and tight junction formation. To establish a physiological model for the distal lung, AXiAECs were cultured for long-term at air-liquid interface (ALI) on-chip. To this end, different stages of alveolar damage including inflammation (via exposure to bacterial lipopolysaccharide) and the response to a profibrotic mediator (via exposure to Transforming growth factor β1) were analyzed. In addition, the expression of relevant host cell factors involved in SARS-CoV-2 infection was investigated to evaluate its potential application for COVID-19 studies. This study shows that AXiAECs cultured on the AXlung-on-chip exhibit an enhanced in vivo-like alveolar character which is reflected into: 1) Alveolar type 1 (AT1) and 2 (AT2) cell specific phenotypes, 2) tight barrier formation (with TER above 1,000 Ω cm2) and 3) reproducible long-term preservation of alveolar characteristics in nearly physiological conditions (co-culture, breathing, ALI). To the best of our knowledge, this is the first time that a primary derived alveolar epithelial cell line on-chip representing both AT1 and AT2 characteristics is reported. This distal lung model thereby represents a valuable in vitro tool to study inhalation toxicity, test safety and efficacy of drug compounds and characterization of xenobiotics.
摘要：
吸入毒性的评价，药物安全性和有效性评估，以及复杂疾病病理机制的研究，越来越依赖于体外肺模型。这是由于逐步转向基于人类的系统，以进行更具预测性和转化性的研究。虽然目前有几种细胞模型可用于上呼吸道，对肺泡远端区域进行建模会带来一些限制，这使得可靠的肺泡体外模型的标准化相对困难。在这项工作中,我们提出了一种新的可重复的肺泡体外模型，它结合了人类衍生的永生化肺泡上皮细胞系（AxiAEC）和模仿肺泡生物物理环境的芯片上器官技术（AXlung-on-chip）。后者模仿体内肺泡环境的关键特征：类似呼吸的3D循环拉伸（10％线性应变，0.2Hz频率)和超薄，多孔和弹性膜。通过基因和蛋白质表达对芯片上培养的ACiAECs的肺泡上皮细胞标记进行了表征。通过TER(隔栅电阻)测量和紧密连接形成检查电池的阻隔性能。建立远端肺的生理模型,在芯片上的气-液界面(ALI)长期培养AxiAECs。为此,分析了肺泡损伤的不同阶段,包括炎症(通过暴露于细菌脂多糖)和对促纤维化介质的反应(通过暴露于转化生长因子β1).此外,研究了SARS-CoV-2感染相关宿主细胞因子的表达，以评估其在COVID-19研究中的潜在应用。这项研究表明，在AXlung芯片上培养的AxiAECs表现出增强的体内肺泡样特征，反映为：1）肺泡1型（AT1）和2（AT2）细胞特异性表型，2）紧密的屏障形成（TER高于1,000Ωcm2）和3）在几乎生理条件下可重复的长期保存肺泡特征（共培养，呼吸,ALI).据我们所知,这是首次报道同时具有AT1和AT2特征的原代来源的肺泡上皮细胞片上细胞系.因此，该远端肺模型代表了研究吸入毒性的有价值的体外工具。测试药物化合物的安全性和有效性以及异种生物的表征。
