A human liver microphysiology platform for investigating physiology, drug safety, and disease models.
A human liver microphysiology platform for investigating physiology, drug safety, and disease models.
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DOI：
10.1177/1535370215592121
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发表时间：
2016-01
期刊：
影响因子：
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通讯作者：
Taylor DL
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文献类型：
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作者：
Vernetti LA;Senutovitch N;Boltz R;DeBiasio R;Shun TY;Gough A;Taylor DL
本文描述了一个微生理学平台的开发和表征，该平台用于药物在肝脏疾病模型中的安全性和有效性，包括人类，3D，微流体，四细胞，顺序分层，自组装肝脏模型（SQL-SAL）;荧光蛋白生物传感器用于机械读数;以及微生理学系统数据库（MPS-Db）用于管理，分析和建模数据。我们方法的目标是在细胞、基质材料和微流控设备参数方面创建最简单的设计，这些设计将支持生理相关的肝脏模型，该模型对于独立肝脏研究和与其他器官芯片的微流控集成而言至少28天是稳健的和可再现的。当前的SQL-SAL使用原代人肝细胞沿着以及生理比例的人内皮细胞（EA.hy926）、免疫细胞（U937）和星状细胞（LX-2），并且在连续流动下可存活至少28天。大约20%的原代肝细胞和/或星状细胞含有荧光蛋白生物传感器（称为哨兵细胞），以通过高含量分析（HCA）测量细胞凋亡、活性氧（ROS）和/或细胞定位。此外，在外排培养基中监测药物、药物代谢产物、白蛋白、尿素和乳酸脱氢酶（LDH）。暴露于180μM曲格列酮或210μM尼美舒利在2-4天内产生急性毒性，而28μM曲格列酮在21天内产生逐渐和延迟的毒性反应，与已知的毒性机制一致，而600μM咖啡因没有影响。证明了曲伐氟沙星与脂多糖（LPS）的免疫介导的毒性，但左氧氟沙星与LPS的免疫介导的毒性。SQL-SAL在30 nM甲氨蝶呤作用下表现出早期纤维化活化，表现为星状细胞迁移增加、α-平滑肌肌动蛋白和胶原蛋白1型α 2表达增加。从体外模型收集的数据可以整合到数据库中，可以访问从外部数据库上传的相关化学、生物活性、临床前和临床信息，以构建预测模型。
This paper describes the development and characterization of a microphysiology platform for drug safety and efficacy in liver models of disease that includes a human, 3D, microfluidic, four-cell, sequentially layered, self-assembly liver model (SQL-SAL); fluorescent protein biosensors for mechanistic readouts; as well as a microphysiology system database (MPS-Db) to manage, analyze, and model data. The goal of our approach is to create the simplest design in terms of cells, matrix materials, and microfluidic device parameters that will support a physiologically relevant liver model that is robust and reproducible for at least 28 days for stand-alone liver studies and microfluidic integration with other organs-on-chips. The current SQL-SAL uses primary human hepatocytes along with human endothelial (EA.hy926), immune (U937) and stellate (LX-2) cells in physiological ratios and is viable for at least 28 days under continuous flow. Approximately, 20% of primary hepatocytes and/or stellate cells contain fluorescent protein biosensors (called sentinel cells) to measure apoptosis, reactive oxygen species (ROS) and/or cell location by high content analysis (HCA). In addition, drugs, drug metabolites, albumin, urea and lactate dehydrogenase (LDH) are monitored in the efflux media. Exposure to 180μM troglitazone or 210μM nimesulide produced acute toxicity within 2–4 days, whereas 28μM troglitazone produced a gradual and much delayed toxic response over 21 days, concordant with known mechanisms of toxicity, while 600μM caffeine had no effect. Immune-mediated toxicity was demonstrated with trovafloxacin with lipopolysaccharide (LPS), but not levofloxacin with LPS. The SQL-SAL exhibited early fibrotic activation in response to 30nM methotrexate, indicated by increased stellate cell migration, expression of alpha-smooth muscle actin and collagen, type 1, alpha 2. Data collected from the in vitro model can be integrated into a database with access to related chemical, bioactivity, preclinical and clinical information uploaded from external databases for constructing predictive models.