Department of Dermatology, VU University Medical Centre, Amsterdam, The Netherlands.
Toxicol In Vitro. 2013 Apr;27(3):1170-9. doi: 10.1016/j.tiv.2012.05.016. Epub 2012 Jun 7.
This manuscript describes methodology and a prediction model for the MUTZ-LC migration assay. The assay represents the physiological change in Langerhans cell (LC) behavior after exposure to a sensitizing chemical, resulting in LC migration from the epidermis to the dermis. MUTZ-LC are derived from the commercially available MUTZ-3 cell line. Upon exposure to a sensitizer MUTZ-LC migrate preferentially towards CXCL12 whereas upon exposure to a non-sensitizer MUTZ-LC migrate towards CCL5. A CXCL12/CCL5 ratio >1.10 in 2/3 independent experiments is indicative of a sensitizer, whereas a CXCL12/CCL5 ratio ≤1.10 is indicative of a non-sensitizer. At non cytotoxic chemical concentrations 9 sensitizers (2,4-dinitrochlorobenzene, paraphenylendiamine, cinnamaldehyde, isoeugenol, nickel-sulfate, tetramethylthiuram disulfide, eugenol, cinnamic-alcohol, ammonium-hexachloroplatinate) were distinguished from 4 non sensitizers (sodium lauryl sulfate, salicylic acid, phenol, octanoic acid). Critical points in assay performance are (i) MUTZ-3 passage number after thawing (p6-p40); (ii) cell viability (>80%); (iii) standard curve to optimize correlation of fluorescence with cell number; and (iv) optimization of the concentration of rhCXCL12 and rhCCL5 in transwell. The protocol has been tested in three European laboratories and results suggest that it may provide working conditions for performing the DC migration assay which is aimed at distinguishing sensitizers from non sensitizers.
本文描述了 MUTZ-LC 迁移测定法的方法和预测模型。该测定法代表了朗格汉斯细胞（LC）在暴露于敏化化学物质后行为的生理变化，导致 LC 从表皮迁移到真皮。MUTZ-LC 源自市售的 MUTZ-3 细胞系。暴露于敏化剂时，MUTZ-LC 优先向 CXCL12 迁移，而暴露于非敏化剂时，MUTZ-LC 向 CCL5 迁移。在 2/3 个独立实验中，CXCL12/CCL5 比值>1.10 表明为敏化剂，而 CXCL12/CCL5 比值≤1.10 表明为非敏化剂。在非细胞毒性化学物质浓度下，从 9 种敏化剂（2,4-二硝基氯苯、对苯二胺、肉桂醛、异丁香酚、硫酸镍、四甲基秋兰姆二硫化物、丁香酚、肉桂醇、六氯铂酸铵）中区分出 4 种非敏化剂（月桂基硫酸钠、水杨酸、苯酚、辛酸）。测定法性能的关键点是：(i) 解冻后 MUTZ-3 的传代数（p6-p40）；(ii) 细胞活力（>80%）；(iii) 优化荧光与细胞数相关性的标准曲线；以及 (iv) 优化 rhCXCL12 和 rhCCL5 在 Transwell 中的浓度。该方案已在三个欧洲实验室进行了测试，结果表明它可能为进行 DC 迁移测定提供工作条件，旨在区分敏化剂和非敏化剂。
