Hsu Hao-Hsiang, Kracht John-Kevin, Harder Laura Elisabeth, Rudnik Kerstin, Lindner Gerd, Schimek Katharina, Marx Uwe, Pörtner Ralf
Institute of Bioprocess and Biosystems Engineering, Hamburg University of Technology.
Institute of Biotechnology, Department Medical Biotechnology, Technische Universität Berlin.
J Vis Exp. 2018 Feb 23(132):56412. doi: 10.3791/56412.
In vitro cultivated skin models have become increasingly relevant for pharmaceutical and cosmetic applications, and are also used in drug development as well as substance testing. These models are mostly cultivated in membrane-insert systems, their permeability toward different substances being an essential factor. Typically, applied methods for determination of these parameters usually require large sample sizes (e.g., Franz diffusion cell) or laborious equipment (e.g., fluorescence recovery after photobleaching (FRAP)). This study presents a method for determining permeability coefficients directly in membrane-insert systems with diameter sizes of 4.26 mm and 12.2 mm (cultivation area). The method was validated with agarose and collagen gels as well as a collagen cell model representing skin models. The permeation processes of substances with different molecular sizes and permeation through different cell models (consisting of collagen gel, fibroblast, and HaCaT) were accurately described. Moreover, to support the above experimental method, a simulation was established. The simulation fits the experimental data well for substances with small molecular size, up to 14 x 10 m Stokes radius (4,000 MW), and is therefore a promising tool to describe the system. Furthermore, the simulation can considerably reduce experimental efforts and is robust enough to be extended or adapted to more complex setups.
体外培养的皮肤模型在制药和化妆品应用中变得越来越重要，也用于药物开发和物质测试。这些模型大多在膜插入系统中培养，其对不同物质的渗透性是一个关键因素。通常，用于测定这些参数的应用方法通常需要大样本量（例如，弗兰兹扩散池）或繁琐的设备（例如，光漂白后荧光恢复（FRAP））。本研究提出了一种直接在直径为4.26毫米和12.2毫米（培养面积）的膜插入系统中测定渗透系数的方法。该方法用琼脂糖和胶原蛋白凝胶以及代表皮肤模型的胶原蛋白细胞模型进行了验证。准确描述了不同分子大小的物质的渗透过程以及通过不同细胞模型（由胶原蛋白凝胶、成纤维细胞和HaCaT组成）的渗透。此外，为了支持上述实验方法，建立了一个模拟。对于斯托克斯半径高达14×10米（4000MW）的小分子物质，该模拟与实验数据拟合良好，因此是描述该系统的一个有前途的工具。此外，该模拟可以大大减少实验工作量，并且足够稳健，可以扩展或适应更复杂的设置。
