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刊物信息

期刊名称:药物分析杂志
主管单位:中国科学技术协会
主办单位:中国药学会
承办:中国食品药品检定研究院
主编:金少鸿
地址:北京天坛西里2号
邮政编码:100050
电话:010-67012819,67058427
电子邮箱:ywfx@nicpbp.org.cn
国际标准刊号:ISSN 0254-1793
国内统一刊号:CN 11-2224/R
邮发代号:2-237
 

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三维表皮模型Epikutis?参考品的研制

Development of three-dimensional epidermal model Epikutis? reference material

作者(英文):
分类号:R917
出版年·卷·期(页码):2019,39 (12):2117-2125
DOI: 10.16155/j.0254-1793.2017.01.01
-----摘要:-------------------------------------------------------------------------------------------

目的:考察体外重组三维(3D)表皮模型(Epikutis?)作为标准物质及用于替代模型皮肤刺激性检测方法的可行性。方法:利用由人源性角质形成细胞构建的体外重组3D表皮模型(Epikutis?)作为标准物质的原料,参照经济合作与发展组织439文件(OECD 439):替代模型皮肤刺激性检测方法,选择组织活力和屏障功能指标,考察表皮模型的均匀性和稳定性,并通过多实验协作测定对其组织活力和屏障指数进行分析。其中,组织活力测定采用MTT试验,以阴性对照光密度值作为组织活力指标;屏障功能测定采用1% TritonX-100作用不同时间后,测定半数活性时间,作为屏障指数(ET50)。结果:Epikutis?表皮模型(每批100多个)的均匀性良好,其样品间均匀性不确定度为:ubb=0.013。短期稳定性(模拟保存运输条件下,运输箱内温度4~25℃放置24 h),组织活力(光密度,OD)为1.029,其短期稳定性不确定度为:us=0.018;ET50的平均值为(2.08±0.12)h,其短期稳定性不确定度us=0.159,无明显活性及屏障功能降低趋势。协作测定结果:OD为1.033。其检测不确定度为:uchar=0.036;最终的合成不确定度(含ubbusuchar)为0.043。ET50的协作标定结果为1.70 h;最终的合成不确定度为0.159。选择10种OECD 439提供的参考物质种类,进行皮肤刺激性试验,其结果与OECD439发布的刺激性判定结果一致,符合预期结果。结论:Epikutis?表皮模型的组织活力:1.033±0.086,最终定值范围为:0.9≤OD≤1.2;ED50:(1.70±0.32)h,最终定值范围为:1.4 h≤ET50≤2.0 h。上述研究结果表明Epikutis?表皮皮肤模型满足OECD TG 439所推荐的替代模型皮肤刺激性检测方法的使用,可以用于化妆品、化学试剂、医疗器械等皮肤刺激性定性检测,也为纳米材料的透皮吸收及皮肤毒性研究提供了替代方法模型。

-----英文摘要:---------------------------------------------------------------------------------------

Objective: To investigate the feasibility of Epikutis?,a three-dimensional (3D) epidermal model as reference material and applicability of Epikutis? in in vitro skin irritation-reconstructed human epidermis test. Methods: The raw material used for reference material development was established based on the in vitro system of reconstructed human epidermis,which used human derived non-transformed keratinocytes as cell source to reconstruct an epidermal model. This 3D epidermal model (Epikutis?) represents histology and cytoarchitecture of human skin. According to Organization for Economic Cooperation and Development (OECD) Test Guideline 439,tissue viability and barrier function of Epikutis? were selected as characteristic value of reference material,and using these items to evaluate the uniformity,stability and to definite characteristic value. The tissue viability was assayed by 3-(4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide (MTT,detected optical density,OD) value of negative control,and the barrier function was measured by using 1% Triton X-100 for different time,and the half active time was detected as the barrier index (ET50). Results: The Epikutis? models have good uniformity,and the uncertainty value was ubb=0.013 within over hundred model samples. The short-term stability was designed to mimic 24 h of preservation and transportation at 4-25℃,and the results showed that the tissue viability was 1.029 and uncertainty value was us=0.018,and the ET50 value was (2.08±0.12) h and uncertainty value was us=0.159. Both the tissue viability and the ET50 value have no significant decreases. Based on the results of collaborative examination of several laboratories,the average tissue viability was 1.033 and uncertainty value uchar=0.036,the final composite uncertainty (contains ubb、us and uchar) was 0.043;and the average ET50 value was 1.70 h and the final composite uncertainty value was 0.159. The results of skin irritation tests on 10 chemicals selected from OECD 439 showed that the sensitivity was 100%,the results were consistent with the evaluation results published by the OECD439. Conclusion: The tissue viability of Epikutis? epidermal model was 1.033±0.086,and final range was 0.9 ≤ OD ≤ 1.2;and the ET50 value was (1.70±0.32) h,and final range was 1.4 h ≤ ET50 ≤ 2.0 h. The Epikutis? epidermal model is a suitable model that meets the application of OECD 439 in vitro skin irritation-reconstructed human epidermis test,and it has prospect to be used for qualitative detection skin irritation of cosmetic,chemicals,medical devices and also provide an alternative tool for investigating skin toxicity and permeability of nanomaterials.

-----参考文献:---------------------------------------------------------------------------------------

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