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Development of bioassays for gaseous chemicals using air-liquid interface culture of human lung epithelial cells

利用人肺上皮细胞气液界面培养物开发气态化学物质生物测定法

基本信息

批准号：
13480163
负责人：
SAKAI Yasuyuki
金额：
$ 7.55万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13480163/
关键词：
Alveolus Air way ir-liquid interface culture damage to the lung intake Suspended particular matter Cytochrome P450 Numerical simulation 浮遊粒子状物体肺胞上皮肺気道上皮バイオアッセイ有機溶媒

项目摘要

To establish a series of toxicitiy evaluation systems for chemicals or environmental samples in gas phase, we employed air-liquid interface culture (ALIC) of human bronchial (air way) and alveolus epithelial cells as a basic culture method, and investigated their applications to volatile organic compounds and suspended paniculate matter (SPM).First, using a human bronchial epithelial cell line, Calu-3, we developed a simple closed system for direct gas exposure, and tested the toxicity expression of Calu-3 to several organic compounds during 48 hours of loading. The toxicity in ALIC exposure was higher than that in conventional exposure in the liquid phase. The reason was largely explained by numerical estimation that chemical concentration on the cell surface in the liquid culture is lower than that in ALIC culture, in the cause of the diffusion process of molecules in the surface liquid layer. These results indicate that basic concept of the combination of ALIC of lung cells and a si … More mple closed system is promising as a testing cytotoxicrly to gaseous compounds.Second, in vivo mimicking exposure of actual SPM samples to human lung alveolar epithelia was achieved using an air-liquid interface culture of human lung alveolar A549 cells, resulting in successful observation of the toxicity in terms of the cell survivability after 48 hours exposure. In addition, an integrated use of the ALIC-based A549 cell assay for general toxicity and the EROD-based Hep G2 cell assay for PAHs was recommended to evaluate the primary toxicity of SPM. We thereby understood the permeation kinetics of SPM toxicity across the alveolar epithelia in terms of the EROD capacity of Hep G2 cells, and were able to obtain basic information on SPM toxicity in humans.Third, we developed a numerical simulation model that describes elution of chemicals initially adsorbed onto SPM samples, intake and accumulation of these chemicals into alveolus cell layer, and their final permeation to the systemic blood circulation. By focusing on the EROD activity of SPM samples, we quantified the comprehensive toxicity of the relevant samples and used these toxicitis in determining parameters involved in these numerical simulations. In addition, we extended the numerical models in actual situations that reflect low-dose and long-term exposure of SPM to humans across the very thin layer of the alveolar tissue. Less

为了建立一系列气相化学品或环境样品的毒性评价体系，以人支气管上皮细胞和肺泡上皮细胞的气液界面培养（ALIC）为基础，研究了其在挥发性有机物和悬浮颗粒物（SPM）中的应用。我们开发了一个简单的直接气体暴露的封闭系统，并在48小时的加载期间测试了Calu-3对几种有机化合物的毒性表达。在液相中，ALIC暴露的毒性高于常规暴露。其原因主要是通过数值估计来解释的，即液体培养中细胞表面的化学浓度低于ALIC培养中的化学浓度，这是由于分子在表面液体层中的扩散过程。这些结果表明，肺细胞ALIC与细胞增殖抑制剂结合的基本概念是可行的。 ...更多信息第二，利用人肺泡A549细胞的气液界面培养，在体内模拟实际SPM样品暴露于人肺泡上皮细胞，成功地观察到暴露48小时后细胞的存活率。此外，建议综合使用基于ALIC的A549细胞试验的一般毒性和基于EROD的Hep G2细胞试验的多环芳烃评价SPM的主要毒性。我们由此了解了SPM毒性通过Hep G2细胞EROD能力的肺泡上皮细胞的渗透动力学，并能够获得SPM在人体中毒性的基本信息。第三，我们开发了一个数值模拟模型，该模型描述了最初吸附在SPM样品上的化学物质的洗脱，这些化学物质进入肺泡细胞层的摄入和积累，以及它们最终渗透到全身血液循环中。通过关注SPM样品的EROD活性，我们量化了相关样品的综合毒性，并将这些毒性用于确定这些数值模拟中涉及的参数。此外，我们扩展了数值模型，在实际情况下，反映低剂量和长期暴露的SPM人类在非常薄的肺泡组织层。少