Tirella Annalisa, Marano Mauro, Vozzi Federico, Ahluwalia Arti
Centro Interdipartimentale di Ricerca "E.Piaggio", University of Pisa, via Diotisalvi 2, 56126 Pisa, Italy.
Toxicol In Vitro. 2008 Dec;22(8):1957-64. doi: 10.1016/j.tiv.2008.09.016. Epub 2008 Oct 1.
A great deal of effort is being dedicated to the development of new devices able to conduct effective in vitro toxicology analyses. This paper describes the use of a microfluidic gradient maker for the toxicological analysis of two conventional local anesthetics, bupivacaine and lidocaine on cell cultures. The microfluidic device was designed and simulated using COMSOL Multiphysics and the concentration gradient in the microfluidic network was analysed through a fluidodynamic and diffusive study. Subsequently the device was fabricated with soft lithography, casting PDMS in a master to obtain channels about 250 microm deep. Both drugs were tested on C2C12 myoblasts and an analysis was performed using propidium iodide staining followed by an imaging processing routine to obtain quantitative dose-response profiles in the gradient maker. The system was critically compared with microwell-based toxicity testing. The results show that the GM is a more sensitive method for detection of cell toxicity, and compared with testing of drug toxicity using microwells with individual cell cultures, allows one shot testing with a single cell culture exposed to a large number of concentrations. However, the flow rates required to obtain a suitable concentration range across the device may damage shear sensitive cells.
大量的努力正致力于开发能够进行有效的体外毒理学分析的新设备。本文描述了一种微流控梯度发生器在两种传统局部麻醉药布比卡因和利多卡因对细胞培养物的毒理学分析中的应用。该微流控设备使用COMSOL Multiphysics进行设计和模拟，并通过流体动力学和扩散研究分析微流控网络中的浓度梯度。随后，采用软光刻技术制造该设备，在模具中浇铸聚二甲基硅氧烷（PDMS）以获得约250微米深的通道。两种药物均在C2C12成肌细胞上进行测试，并使用碘化丙啶染色进行分析，随后进行图像处理程序以在梯度发生器中获得定量剂量反应曲线。该系统与基于微孔板的毒性测试进行了严格比较。结果表明，梯度发生器是一种检测细胞毒性更灵敏的方法，与使用单个细胞培养的微孔板进行药物毒性测试相比，它允许对单个细胞培养物暴露于大量浓度的药物进行一次性测试。然而，在整个设备上获得合适浓度范围所需的流速可能会损害对剪切敏感的细胞。
