Assembly of Hepatocyte Spheroids Using Magnetic 3D Cell Culture for CYP450 Inhibition/Induction.
Assembly of Hepatocyte Spheroids Using Magnetic 3D Cell Culture for CYP450 Inhibition/Induction.
复制标题
DOI：
10.3390/ijms18051085
复制
发表时间：
2017-05-18
影响因子：
5.6
通讯作者：
Souza GR
中科院分区：
文献类型：
--
作者：
Desai PK;Tseng H;Souza GR
对于研究药物诱导的肝损伤的体外方法存在显著需求，所述方法对于现有的高通量系统是快速的、可再现的和可扩展的。然而，传统的单层和悬浮培养的肝细胞是难以处理和风险的表型损失。通常，三维（3D）细胞培养平台有助于重现天然肝组织表型，但受到高通量筛选的技术限制，包括可扩展性、速度和处理。在这里，我们开发了一种新的细胞色素P450（CYP 450）的诱导/抑制测定使用磁性三维细胞培养，克服了其他平台的限制，通过聚集磁化细胞与磁力。有了这个平台，球状体可以快速组装和容易处理，同时复制天然肝功能。我们在384孔板中组装了原代人肝细胞的球状体，并将该培养物维持5天以上，包括使用已知CYP 450诱导剂/抑制剂的72小时诱导期。对球状体中的CYP 450活性和活力进行了评估，并与单层平行进行了比较。如预期，在球状体中诱导/抑制CYP 450活性，与任何毒性反应无关。球形细胞显示出显著高于单层细胞的CYP 450活性和诱导的基线水平。球状体中白蛋白和多药耐药相关蛋白（MRP 2）的阳性染色表明球状体中保留了肝细胞功能。该研究提出了使用磁性3D细胞培养组装和处理新型肝组织模型的概念验证。
There is a significant need for in vitro methods to study drug-induced liver injury that are rapid, reproducible, and scalable for existing high-throughput systems. However, traditional monolayer and suspension cultures of hepatocytes are difficult to handle and risk the loss of phenotype. Generally, three-dimensional (3D) cell culture platforms help recapitulate native liver tissue phenotype, but suffer from technical limitations for high-throughput screening, including scalability, speed, and handling. Here, we developed a novel assay for cytochrome P450 (CYP450) induction/inhibition using magnetic 3D cell culture that overcomes the limitations of other platforms by aggregating magnetized cells with magnetic forces. With this platform, spheroids can be rapidly assembled and easily handled, while replicating native liver function. We assembled spheroids of primary human hepatocytes in a 384-well format and maintained this culture over five days, including a 72 h induction period with known CYP450 inducers/inhibitors. CYP450 activity and viability in the spheroids were assessed and compared in parallel with monolayers. CYP450 activity was induced/inhibited in spheroids as expected, separate from any toxic response. Spheroids showed a significantly higher baseline level of CYP450 activity and induction over monolayers. Positive staining in spheroids for albumin and multidrug resistance-associated protein (MRP2) indicates the preservation of hepatocyte function within spheroids. The study presents a proof-of-concept for the use of magnetic 3D cell culture for the assembly and handling of novel hepatic tissue models.