Department of Chemical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061, USA.
Tissue Eng Part A. 2011 Mar;17(5-6):677-89. doi: 10.1089/ten.TEA.2010.0423. Epub 2010 Dec 19.
A critical hepatic function is the maintenance of optimal bile acid (BA) compositions to achieve cholesterol homeostasis. BAs are rarely quantified to assess hepatic phenotype in vitro since existing analytical techniques have inadequate resolution. We report a detailed investigation into the biosynthesis and homeostasis of eight primary rat BAs in conventional in vitro hepatocyte cultures and in an engineered liver mimic. The three-dimensional (3D) liver mimic was assembled with layers of primary rat hepatocytes and liver sinusoidal endothelial cells. A high-pressure liquid chromatography and mass spectrometry technique was developed with a detection limit of 1 ng/mL for each BA, which is significantly lower than previous approaches. Over a 2-week culture, only 3D liver mimics exhibited the ratio of conjugated cholic acid to chenodeoxycholic acid that has been observed in vivo. This ratio, an important marker of BA homeostasis, was significantly higher in stable collagen sandwich cultures indicating significant deviation from physiological behavior. The biosynthesis of tauro-β-muricholic acid, a key primary rat BA, doubled only in the engineered liver mimics while decreasing in the other systems. These trends demonstrate that the 3D liver mimics provide a unique platform to study hepatic metabolism.
一项关键的肝脏功能是维持最佳胆汁酸 (BA) 组成以实现胆固醇的体内平衡。由于现有的分析技术分辨率不足，很少对 BA 进行定量以评估体外的肝脏表型。我们报告了一项详细的研究，该研究调查了在传统的体外肝细胞培养物和工程化的肝模拟物中八种主要大鼠 BA 的生物合成和体内平衡。三维 (3D) 肝模拟物由单层原代大鼠肝细胞和肝窦内皮细胞组装而成。开发了一种高效液相色谱和质谱技术，每个 BA 的检测限为 1ng/mL，明显低于以前的方法。在为期 2 周的培养中，只有 3D 肝模拟物表现出与体内观察到的结合胆酸与鹅脱氧胆酸的比例。这种比例是 BA 体内平衡的一个重要标志物，在稳定的胶原夹层培养物中显著更高，表明与生理行为有明显偏差。牛磺酸-β-鼠胆酸，一种主要的大鼠 BA，在工程化的肝模拟物中仅增加了一倍，而在其他系统中则减少了。这些趋势表明，3D 肝模拟物提供了一个独特的平台来研究肝脏代谢。
