基于THP-1细胞的皮肤致敏体外检测方法

目的建立基于人细胞系的替代皮肤致敏动物实验的体外检测方法(h-CLAT),并对化学品、日用化学产品和化妆品植物原料进行皮肤致敏性检测。方法体外培养人急性单核细胞白血病细胞(THP-1),不同浓度受试物与细胞共孵育24 h,通过流式细胞仪检测细胞表面标志物CD86和CD54活化的差异,并对11种已知皮肤致敏化学物质和9种未知样品进行皮肤致敏性预测。同时对未知样品进行豚鼠局部封闭涂皮法验证。结果使用h-CLAT方法准确区分了11种参考物质的皮肤致敏性,9种受试样品中,7种样品被判断为阴性皮肤致敏物质,2种植物提取物被确认为皮肤致敏疑似物质。9种样品的预测结果与动物实验一致。结论 h-CLAT体外检测方法可以代替部分动物测试,用于可溶性皮肤致敏物质的预测。

An in vitro skin sensitization test based on THP-1 cell line

Objective To establish an in vitro skin sensitization test, human cell line activation test ( h-CLAT), based on THP-1 cell line (a human acute monocytic leukemia cell line), and to assess the sensitizing potency of plant raw materials of chemical and cosmetic products by this in vitro skin sensitization test. Method THP-1 cells were cultured in vitro and exposed to 11 reference skin sensitization chemicals and 9 samples, by monitoring the cell viability, cell surface marker CD54/CD86 and relative fluorescence intensity of cells surface after the cells was exposures to the substances, and to discover whether there is a positive reaction. At the same time, Buehler test was used to validate the results of samples tested by h-CLAT. Results 11 reference chemicals were distinguished correctly by h-CLAT. Among the 9 samples tested, 7 samples were recognized as negative sensitizer and 2 plant extracted substances were identified as suspicious skin sensitizer. The qualitative classification of the 9 samples by h-CLAT test was consistent with the results obtained by animal test. Conclusions The h-CLAT-in vitro test can be used to replace some animal tests for the prediction of soluble skin sensitizing substances.