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In vitro toxicity of asbestos substitutes evaluated by cytomagnetometry

通过细胞磁力测定法评估石棉替代品的体外毒性

基本信息

批准号：
11470099
负责人：
AIZAWA Yoshiharu
金额：
$ 4.48万
依托单位：
Kitasato University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

Recently, some asbestos substitutes have been used in Japan, as health disorders occurred by asbestos had been known. However, such substitutes have not proved their safety yet, and it is important to establish screening methods for evaluating health disorders. Then we examined a few evaluations of health disorders for silicon carbide whisker (SiC), titanium dioxide whisker (TiO_2) and rock wool (RW) not only by cytomagnetometry, but also by LDH release assays, morphological observations and apoptotic observations.Both 20, 40, and 60 μg/ml of SiC and TiO_2 in the culture medium showed that relaxation was delayed dose-dependently and decay constants were smaller than those of negative controls. LDH release assay showed that releasing rates were increased dose-dependently. Morphological observations showed multiple vacuolations in cytosol, degenerations of nuclei, and agglutinations of chromatin. And apoptotic observations by using TUNEL method and DNA ladder method showed slight apoptotic cells in a sample added to SiC.On the other hand, 50, 100, and 150μg/ml of RW did not show delays of relaxation and decreasing of decay constants in comparison with negative controls. Results of LDH release assay, morphological observations including TUNEL method and DNA ladder method, and apoptotic observations supported results of cytomagnetometry.Then, it is found that an acute cytotoxicity of RW to alveolar macrophages is less than that of SiC and TiO_2 in in vitro experiments. In future, we are going to use subculture cells instead of alveolar macrophages obtained from bronchoalveolar lavage, to avoid changes in conditions of experiments. In order to observe a mechanism of relaxation optically, we will examine intracellular movements of magnetized particles after magnetization by using a video-enhanced microscope. In addition, cytotoxic evaluations of asbestos substitutes except SiC, TiO_2 and RW will be examined.

最近，日本已使用了一些石棉替代品，因为人们知道石棉会引起健康问题。然而，这些替代品尚未证明其安全性，重要的是要建立评估健康障碍的筛查方法。然后，我们不仅通过细胞磁性测定法，而且还通过LDH释放测定法、形态学观察和凋亡观察，对碳化硅晶须（SiC）、二氧化钛晶须（TiO_2）和岩棉（RW）的健康障碍进行了一些评价。60 μg/ml的SiC和TiO_2使细胞的弛豫时间呈剂量依赖性延长，衰减常数小于阴性对照。乳酸脱氢酶释放实验表明，释放速率呈剂量依赖性增加。形态学观察显示胞浆中有多个空泡，细胞核变性，染色质凝集。用TUNEL法和DNA梯状条带法观察细胞凋亡，发现加入SiC的样品中有轻微的细胞凋亡，而50、100和150μg/ml的RW与阴性对照组相比，没有出现弛豫延迟和衰减常数降低。LDH释放实验、形态学观察（TUNEL法和DNA Ladder法）和细胞凋亡观察结果均支持细胞磁测量的结果，并发现RW对肺泡巨噬细胞的急性毒性低于SiC和TiO_2。今后，我们将使用传代细胞代替从支气管肺泡灌洗获得的肺泡巨噬细胞，以避免实验条件的变化。为了观察光学弛豫机制，我们将使用视频增强显微镜检查磁化后磁化颗粒的细胞内运动。此外，还将对除SiC、TiO_2和RW以外的石棉替代品进行细胞毒性评价。