奈米微粒危害測試技術與生物毒性指標之探討

A Study on the Approaches for Hazard Ranking and Identifying the Biomarkers of the Adverse Effects of Nanoparticles

隨著奈米科技的精進發展，奈米材料已經被廣泛的應用在生活環境中。然而，奈米微粒對生物體產生的毒性與危害，至今仍未十分瞭解。奈米微粒有許多獨特的物化特性，而這些特性可能會影響其細胞毒性反應。雖然毒性測試的方法種類甚多，但是目前尚缺少合理且共通的生物毒性指標、毒性測試系統與危害評估平台等，以作為奈米物質管理機制的參考。本研究選用在生活上已廣泛被應用且可能具有生物毒性之奈米銀進行實驗。合成的奈米銀微粒會依照我們所設定的測試流程，先分析其物化特性再以細胞株進行體外實驗。奈米銀經物化特性分析後，皆符合我們進行實驗所需之材料要求。體外試驗中，我們使用人類支氣管上皮細胞（BEAS-2Bb cell line）進行實驗，並利用MTS與Live/Dead cell viability分析方法進行細胞毒性分析。細胞毒性測試結果顯示，奈米銀曝露細胞後會誘導活性氧物種（ROS）生成、血基質氧化酶（HO-1）基因表現、細胞自體吞噬與細胞凋亡的產生。由於在細胞未產生顯著死亡毒性前，便可觀察到一些生物指標的改變，因此我們建議可利用偵測ROS、HO-1之基因表現並藉由分析細胞自體吞噬現象作為危害效應之初步指標。本研究依據過去經驗及文獻提供一個初步細胞毒性篩選流程項目，經由奈米銀的測試結果，本研究所提出之流程可能也可以進一步應用於其它奈米物質的危害效應測試。

Advancement and improvement of nanotechnology, nanomaterials (NMs) have been comprehensively applied in our modern society and greatly influenced our daily life. However, the toxicity and hazardous impacts of nano-scale particles on organisms is still unclear. The different physico-chemical characteristics are identified in the nanofied particles, which may contribute to their toxic effects on the target cells. Currently, there are numerous approaches to carry out toxicity tests but there is a lack of common and reasonable/sensible biomarkers and detection systems for toxicity evaluation as well as a risk management platform for offering the reference database. In this study, we determined to select silver nanoparticles (AgNPs), which have been commonly used in industry, as the candidate toxicants and tend to demonstrate in vitro toxicity to a certain extent. We characterized the physico-chemical properties of the synthetic nanoparticles. We used human bronchial epithelial cells (BEAS-2B cell line) to evaluate the cytotoxic effects by using MTS and Live/Dead cell viability assays. Our results found that AgNPs led to elicit of reactive oxygen species (ROS), upregulated expression of the Heme oxygenase 1 (HO-1) gene and occurrence of autophagy, which could be the bio-indicators for nanotoxicity. Therefore, the results from the preliminary data shows that MTS and Live/Dead cell viability assays could be used for cell line -based nanotoxicity screening and assessment. To conclusion, the outcome of the present work on AgNPs adverse effects might be implicated to other nanomaterials.