Flamand Nicole, Marrot Laurent, Belaidi Jean-Philippe, Bourouf Linda, Dourille Emilie, Feltes Michèle, Meunier Jean-Roch
L'Oréal, Advanced Research, Life Sciences, Safety Research Department, Phototoxicity-Genotoxicity-Carcinogenicity Group, 1, avenue E. Schueller, F-93600 Aulnay Sous Bois, France.
Mutat Res. 2006 Jul 14;606(1-2):39-51. doi: 10.1016/j.mrgentox.2006.02.009. Epub 2006 May 3.
Today reconstructed skin models that simulate human skin, such as Episkin, are widely used for safety or efficacy pre-screening. Moreover, they are of growing interest for regulatory purposes in the framework of alternatives to animal testing. In order to reduce and eventually replace results of in vivo genotoxicity testing with in vitro data, there is a need to develop new complementary biological models and methods with improved ability to predict genotoxic risk. This can be achieved if these new assays do take into account exposure conditions that are more relevant than in the current test systems. In an attempt to meet this challenge, two new applications using a human reconstructed skin model for in vitro genotoxicity assessment are proposed. The skin is the target organ for dermally exposed compounds or environmental stress. Although attempts have been made to develop genotoxicity test procedures in vivo on mouse skin, human reconstructed skin models have not been used for in vitro genotoxicity testing so far, although they present clear advantages over mouse skin for human risk prediction. This paper presents the results of the development of a specific protocol allowing to perform the comet assay, a genotoxicity test procedure, on reconstructed skin. The comet assay was conducted after treatment of Episkin with UV, Lomefloxacin and UV or 4-nitroquinoline-N-oxide (4NQO). Treatment with the sunscreen Mexoryl was able to reduce the extent of comet signal. A second approach to use reconstructed epidermis in genotoxicity assays is also proposed. Indeed, the skin is a biologically active barrier driving the response to exposure to chemical agents and their possible metabolites. A specific co-culture system (Figure 1) using Episkin to perform the regular micronucleus assay is presented. Micronucleus induction in L5178Y cells cultured underneath Episkin was assessed after treatment of the reconstructed epidermis with mitomycin C, cyclophosphamide or apigenin. This second way of using human reconstructed skin for genotoxicity testing aims at improving the relevance of exposure conditions in in vitro genotoxicity assays for dermally applied compounds.
如今，诸如Episkin等模拟人类皮肤的重建皮肤模型被广泛用于安全性或有效性的预筛选。此外，在动物试验替代方法的框架内，它们在监管目的方面的关注度也日益增加。为了用体外数据减少并最终取代体内遗传毒性测试的结果，有必要开发新的补充性生物学模型和方法，以提高预测遗传毒性风险的能力。如果这些新检测方法确实考虑到比当前测试系统更相关的暴露条件，就能实现这一目标。为应对这一挑战，本文提出了两种使用人类重建皮肤模型进行体外遗传毒性评估的新应用。皮肤是经皮暴露化合物或环境压力的靶器官。尽管已尝试在小鼠皮肤上开展体内遗传毒性测试程序，但人类重建皮肤模型迄今尚未用于体外遗传毒性测试，尽管它们在预测人类风险方面比小鼠皮肤具有明显优势。本文介绍了开发一种特定方案的结果，该方案允许在重建皮肤上进行彗星试验（一种遗传毒性测试程序）。在用紫外线、洛美沙星以及紫外线或4-硝基喹啉-N-氧化物（4NQO）处理Episkin后进行彗星试验。用防晒剂麦素宁滤光环处理能够降低彗星信号的程度。本文还提出了在遗传毒性试验中使用重建表皮的第二种方法。事实上，皮肤是一种生物活性屏障，可驱动对化学剂及其可能代谢物暴露的反应。本文介绍了一种使用Episkin进行常规微核试验的特定共培养系统（图1）。在用丝裂霉素C、环磷酰胺或芹菜素处理重建表皮后，评估在Episkin下方培养的L5178Y细胞中的微核诱导情况。这种将人类重建皮肤用于遗传毒性测试的第二种方法旨在提高体外遗传毒性试验中经皮应用化合物暴露条件的相关性。
