组织工程表皮与单核白血病细胞共培养的致敏性检测模型

A co-culture model of tissue engineering epidermis and THP-1 cells for sensitization testing

背景:大量研究表明体外构建的组织工程皮肤可用于化学物质的刺激性检测。目的:构建组织工程表皮和单核白血病细胞共培养体外模型,并用致敏物初步验证。方法:利用人源性角质形成细胞构建组织工程表皮,和单核白血病细胞进行共培养,干预24h后,ELISA法检测空白对照组,溶剂对照组,二硝基氟苯处理组和十二烷基硫酸钠处理组培养液中白细胞介素1β的质量浓度,流式细胞术检测各组中单核白血病细胞表面标记物CD86和CD54的平均荧光强度。结果与结论:角质形成细胞气液界面培养14d后,苏木精-伊红染色结果显示构建的组织工程表皮含有8～10层细胞结构,泛角蛋白免疫组化染色阳性。二硝基氟苯处理组培养液中白细胞介素1β质量浓度,单核白血病细胞表面标记物CD86和CD54的平均荧光强度均高于溶剂对照组(P<0.05)。实验成功构建了组织工程表皮和单核白血病细胞共培养的致敏物检测模型。

BACKGROUND：Several studies have shown that tissue engineering skin are used for skin irritation testing.OBJECTIVE：To construct a co-culture model of tissue engineering epidermis and THP-1 cells,and to identify the specificity of the co-culture model to sensitizers.METHODS：Tissue engineering epidermis was constructed by human keratinocytes.Then,THP-1 cells were co-cultured tissue engineering epidermis.After 24 hours treatment on tissue engineering epidermis,the cultured medium and THP-1 cells were collected,respectively.ELISA was used to detect the concentration of interleukin 1beta （IL-1β） in the culture medium,and the mean fluorescence intensities （MFI） measured by flow cytometry were used to evaluate the expression of CD86 and CD54 on THP-1 cells.RESULTS AND CONCLUSION：After 14 days keratinocytes were cultured at air-liquid interface,HE staining showed 8-10 layers in tissue engineering epidermis.And pan-keratin immuohistochemistry staining was positive.After 24 hours treatment of chemicals,the concentration of IL-1β and MFIs of CD86 and CD54 in DNFB-treated group were higher that that in vehicle control group （P〈0.05）.The results prove that a co-culture model of tissue engineering epidermis and THP-1 cells was successfully established,and it can be used for sensitization test,which gives a new method for toxicity testing in vitro.