An in vitro testing strategy towards mimicking the inhalation of high aspect ratio nanoparticles.
An in vitro testing strategy towards mimicking the inhalation of high aspect ratio nanoparticles.
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DOI：
10.1186/s12989-014-0040-x
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发表时间：
2014-09-23
影响因子：
10
通讯作者：
Clift MJ
中科院分区：
文献类型：
--
作者：
Endes C;Schmid O;Kinnear C;Mueller S;Camarero-Espinosa S;Vanhecke D;Foster EJ;Petri-Fink A;Rothen-Rutishauser B;Weder C;Clift MJ
挑战仍然是通过吸入可靠地模拟人类暴露于高纵横比纳米颗粒（HARN）。复杂的，多细胞的体外模型是一个特别有利的解决方案，这个问题，特别是当考虑到需要提供现实和有效的替代侵入性动物实验的HARN危害评估。通过将材料表征技术的系统试验台，一个特定的空气-液体细胞暴露系统，除了关键的生化终点外，还可以实时监测细胞释放的HARN剂量，在这里，我们证明了一种成功的方法来研究体外HARN气溶胶的危害。纤维素纳米晶体（CNCs）来源于棉花和被囊动物，具有不同的纵横比（~9和~80），被用作模型HARN样品。具体地，使用“气液界面细胞暴露系统”（ALICE）以0.14至1.57 μg/cm 2的实际细胞递送浓度将良好分散和表征的CNC悬浮液雾化。然后使用暴露后24小时的气液界面（ALI）处的人上皮气道屏障的3D多细胞体外模型评估每个HARN样品的生物学影响（细胞毒性、氧化应激水平和促炎作用）。此外，使用结晶石英（DQ 12）作为ALICE系统中的阳性颗粒对照和长纤维铁石棉（LFA）对测试策略进行了验证，以确认体外模型对纤维损伤的敏感性。CNC悬浮液的快速（≤4 min）受控雾化使得剂量受控且空间均匀的CNC沉积到在ALI条件下培养的细胞上。用石英晶体微天平实时监测细胞递送的CNC剂量。与CNC纵横比无关，在最高浓度1.57 μg/cm 2下未观察到显著的细胞毒性（p > 0.05）、氧化应激诱导或（促）炎症反应。DQ 12和LFA在体外亚致死浓度下均引起显著（p < 0.05）的促炎反应。总之，虽然本研究强调了CNC的良性性质，但提出的先进技术和机制方法允许采用最先进的检测策略，以实际有效地确定与吸入暴露HARN有关的体外危害。本文的在线版本（doi：10.1186/s12989-014-0040-x）包含补充材料，可供授权用户使用。
The challenge remains to reliably mimic human exposure to high aspect ratio nanoparticles (HARN) via inhalation. Sophisticated, multi-cellular in vitro models are a particular advantageous solution to this issue, especially when considering the need to provide realistic and efficient alternatives to invasive animal experimentation for HARN hazard assessment. By incorporating a systematic test-bed of material characterisation techniques, a specific air-liquid cell exposure system with real-time monitoring of the cell-delivered HARN dose in addition to key biochemical endpoints, here we demonstrate a successful approach towards investigation of the hazard of HARN aerosols in vitro. Cellulose nanocrystals (CNCs) derived from cotton and tunicates, with differing aspect ratios (~9 and ~80), were employed as model HARN samples. Specifically, well-dispersed and characterised CNC suspensions were aerosolised using an “Air Liquid Interface Cell Exposure System” (ALICE) at realistic, cell-delivered concentrations ranging from 0.14 to 1.57 μg/cm2. The biological impact (cytotoxicity, oxidative stress levels and pro-inflammatory effects) of each HARN sample was then assessed using a 3D multi-cellular in vitro model of the human epithelial airway barrier at the air liquid interface (ALI) 24 hours post-exposure. Additionally, the testing strategy was validated using both crystalline quartz (DQ12) as a positive particulate control in the ALICE system and long fibre amosite asbestos (LFA) to confirm the susceptibility of the in vitro model to a fibrous insult. A rapid (≤4 min), controlled nebulisation of CNC suspensions enabled a dose-controlled and spatially homogeneous CNC deposition onto cells cultured under ALI conditions. Real-time monitoring of the cell-delivered CNC dose with a quartz crystal microbalance was accomplished. Independent of CNC aspect ratio, no significant cytotoxicity (p > 0.05), induction of oxidative stress, or (pro)-inflammatory responses were observed up to the highest concentration of 1.57 μg/cm2. Both DQ12 and LFA elicited a significant (p < 0.05) pro-inflammatory response at sub-lethal concentrations in vitro. In summary, whilst the present study highlights the benign nature of CNCs, it is the advanced technological and mechanistic approach presented that allows for a state of the art testing strategy to realistically and efficiently determine the in vitro hazard concerning inhalation exposure of HARN. The online version of this article (doi:10.1186/s12989-014-0040-x) contains supplementary material, which is available to authorized users.