Background

The establishment of reliable and robust in vitro models for hazard assessment, a prerequisite for moving away from animal testing, requires the evaluation of model transferability and reproducibility. Lung models that can be exposed via the air, by means of an air-liquid interface (ALI) are promising in vitro models for evaluating the safety of nanomaterials (NMs) after inhalation exposure. We performed an inter-laboratory comparison study to evaluate the transferability and reproducibility of a lung model consisting of the human bronchial cell line Calu-3 as a monoculture and, to increase the physiologic relevance of the model, also as a co-culture with macrophages (either derived from the THP-1 monocyte cell line or from human blood monocytes). The lung model was exposed to NMs using the VITROCELL® Cloud12 system at physiologically relevant dose levels.

Results

Overall, the results of the 7 participating laboratories are quite similar. After exposing Calu-3 alone and Calu-3 co-cultures with macrophages, no effects of lipopolysaccharide (LPS), quartz (DQ12) or titanium dioxide (TiO₂) NM-105 particles on the cell viability and barrier integrity were detected. LPS exposure induced moderate cytokine release in the Calu-3 monoculture, albeit not statistically significant in most labs. In the co-culture models, most laboratories showed that LPS can significantly induce cytokine release (IL-6, IL-8 and TNF-α). The exposure to quartz and TiO₂ particles did not induce a statistically significant increase in cytokine release in both cell models probably due to our relatively low deposited doses, which were inspired by in vivo dose levels. The intra- and inter-laboratory comparison study indicated acceptable interlaboratory variation for cell viability/toxicity (WST-1, LDH) and transepithelial electrical resistance, and relatively high inter-laboratory variation for cytokine production.

Conclusion

The transferability and reproducibility of a lung co-culture model and its exposure to aerosolized particles at the ALI were evaluated and recommendations were provided for performing inter-laboratory comparison studies. Although the results are promising, optimizations of the lung model (including more sensitive read-outs) and/or selection of higher deposited doses are needed to enhance its predictive value before it may be taken further towards a possible OECD guideline.

中文翻译：

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暴露的气液界面共培养肺模型的可转移性和再现性

背景

建立可靠且稳健的体外危害评估模型是摆脱动物试验的先决条件，需要评估模型的可转移性和再现性。可以通过气液界面（ALI）通过空气暴露的肺模型是有前途的体外模型，用于评估吸入暴露后纳米材料（NM）的安全性。我们进行了一项实验室间比较研究，以评估由人支气管细胞系 Calu-3 组成的肺模型作为单一培养物的可转移性和再现性，并为了增加模型的生理相关性，也作为与巨噬细胞的共培养物（源自 THP-1 单核细胞系或人血单核细胞）。使用 VITROCELL® Cloud12 系统以生理相关剂量水平将肺模型暴露于 NM。

结果

总体而言，7个参与实验室的结果相当相似。单独暴露 Calu-3 以及 Calu-3 与巨噬细胞共培养后，未检测到脂多糖 (LPS)、石英 (DQ12) 或二氧化钛 (TiO ₂ ) NM-105 颗粒对细胞活力和屏障完整性的影响。LPS 暴露在 Calu-3 单一培养物中诱导了适度的细胞因子释放，尽管在大多数实验室中没有统计学意义。在共培养模型中，大多数实验室表明LPS可以显着诱导细胞因子释放（IL-6、IL-8和TNF-α）。在两种细胞模型中，暴露于石英和TiO ₂颗粒并没有引起细胞因子释放统计上显着的增加，这可能是由于我们的沉积剂量相对较低，这是受到体内剂量水平的启发。实验室内和实验室间比较研究表明，细胞活力/毒性（WST-1、LDH）和跨上皮电阻的实验室间差异可接受，细胞因子产生的实验室间差异相对较高。

结论

评估了肺共培养模型的可转移性和再现性及其在 ALI 中暴露于雾化颗粒的情况，并为进行实验室间比较研究提供了建议。尽管结果很有希望，但在进一步朝着可能的经合组织指南迈进之前，需要优化肺部模型（包括更灵敏的读数）和/或选择更高的沉积剂量，以增强其预测价值。