Institute of Bioengineering and Nanotechnology, A*STAR, The Nanos, #04-01, 31 Biopolis Way, Singapore 138669, Singapore.
Biomaterials. 2011 Feb;32(4):1229-41. doi: 10.1016/j.biomaterials.2010.09.064. Epub 2010 Oct 23.
Hepatotoxicity evaluation of pharmaceutical lead compounds in early stages of drug development has drawn increasing attention. Sandwiched hepatocytes exhibiting stable functions in culture represent a standard model for hepatotoxicity testing of drugs. We have developed a robust and high-throughput hepatotoxicity testing platform based on the sandwiched hepatocytes for drug screening. The platform involves a galactosylated microfabricated membrane sandwich to support cellular function through uniform and efficient mass transfer while protecting cells from excessive shear. Perfusion bioreactor further enhances mass transfer and cellular functions over long period; and hepatocytes are readily transferred to 96-well plate for high-throughput robotic liquid handling. The bioreactor design and perfusion flow rate are optimized by computational fluid dynamics simulation and experimentally. The cultured hepatocytes preserved 3D cell morphology, urea production and cytochrome p450 activity of the mature hepatocytes for 14 days. When the perfusion-cultured sandwich is transferred to 96-well plate for drug testing, the hepatocytes exhibited improved drug sensitivity and low variability in hepatotoxicity responses amongst cells transferred from different dates of perfusion culture. The platform enables robust high-throughput screening of drug candidates.
在药物开发的早期阶段，对药物先导化合物的肝毒性评估受到了越来越多的关注。在培养中表现出稳定功能的夹心法肝细胞是药物肝毒性测试的标准模型。我们已经开发了一种基于夹心法肝细胞的强大的高通量肝毒性测试平台，用于药物筛选。该平台涉及一种半乳糖化的微加工膜夹层，通过均匀有效的质量传递来支持细胞功能，同时保护细胞免受过度剪切。灌注式生物反应器进一步增强了长期的质量传递和细胞功能；并且肝细胞易于转移到 96 孔板进行高通量机器人液体处理。通过计算流体动力学模拟和实验优化了生物反应器设计和灌注流速。培养的肝细胞在 14 天内保持了 3D 细胞形态、尿素生成和成熟肝细胞的细胞色素 p450 活性。当灌注培养的夹层转移到 96 孔板进行药物测试时，与从不同灌注培养日期转移的细胞相比，肝细胞表现出提高的药物敏感性和肝毒性反应的低变异性。该平台能够实现稳健的高通量药物候选物筛选。
