Lee-Montiel Felipe T, George Subin M, Gough Albert H, Sharma Anup D, Wu Juanfang, DeBiasio Richard, Vernetti Lawrence A, Taylor D Lansing
1 Drug Discovery Institute, University of Pittsburgh, Pittsburgh, PA 15260, USA.
2 Department of Computational and Systems Biology, University of Pittsburgh, Pittsburgh, PA 15260,USA.
Exp Biol Med (Maywood). 2017 Oct;242(16):1617-1632. doi: 10.1177/1535370217703978. Epub 2017 Apr 14.
This article describes our next generation human Liver Acinus MicroPhysiology System (LAMPS). The key demonstration of this study was that Zone 1 and Zone 3 microenvironments can be established by controlling the oxygen tension in individual devices over the range of ca. 3 to 13%. The oxygen tension was computationally modeled using input on the microfluidic device dimensions, numbers of cells, oxygen consumption rates of hepatocytes, the diffusion coefficients of oxygen in different materials and the flow rate of media in the MicroPhysiology System (MPS). In addition, the oxygen tension was measured using a ratiometric imaging method with the oxygen sensitive dye, Tris(2,2'-bipyridyl) dichlororuthenium(II) hexahydrate (RTDP) and the oxygen insensitive dye, Alexa 488. The Zone 1 biased functions of oxidative phosphorylation, albumin and urea secretion and Zone 3 biased functions of glycolysis, α1AT secretion, Cyp2E1 expression and acetaminophen toxicity were demonstrated in the respective Zone 1 and Zone 3 MicroPhysiology System. Further improvements in the Liver Acinus MicroPhysiology System included improved performance of selected nonparenchymal cells, the inclusion of a porcine liver extracellular matrix to model the Space of Disse, as well as an improved media to support both hepatocytes and non-parenchymal cells. In its current form, the Liver Acinus MicroPhysiology System is most amenable to low to medium throughput, acute through chronic studies, including liver disease models, prioritizing compounds for preclinical studies, optimizing chemistry in structure activity relationship (SAR) projects, as well as in rising dose studies for initial dose ranging. Impact statement Oxygen zonation is a critical aspect of liver functions. A human microphysiology system is needed to investigate the impact of zonation on a wide range of liver functions that can be experimentally manipulated. Because oxygen zonation has such diverse physiological effects in the liver, we developed and present a method for computationally modeling and measuring oxygen that can easily be implemented in all MPS models. We have applied this method in a liver MPS in which we are then able to control oxygenation in separate devices and demonstrate that zonation-dependent hepatocyte functions in the MPS recapitulate what is known about in vivo liver physiology. We believe that this advance allows a deep experimental investigation on the role of zonation in liver metabolism and disease. In addition, modeling and measuring oxygen tension will be required as investigators migrate from PDMS to plastic and glass devices.
本文介绍了我们的下一代人肝腺泡微生理系统（LAMPS）。本研究的关键证明是，通过在各个装置中控制氧气张力在约3%至13%的范围内，可以建立1区和3区微环境。利用微流控装置尺寸、细胞数量、肝细胞耗氧率、氧气在不同材料中的扩散系数以及微生理系统（MPS）中培养基流速等输入信息，对氧气张力进行了计算建模。此外，使用对氧气敏感的染料三水合三（2,2'-联吡啶）二氯钌（II）（RTDP）和对氧气不敏感的染料Alexa 488的比率成像方法测量氧气张力。在各自的1区和3区微生理系统中，证明了氧化磷酸化、白蛋白和尿素分泌的1区偏向性功能以及糖酵解、α1抗胰蛋白酶分泌、Cyp2E1表达和对乙酰氨基酚毒性的3区偏向性功能。肝腺泡微生理系统的进一步改进包括所选非实质细胞性能的改善、包含猪肝脏细胞外基质以模拟狄氏间隙，以及改进培养基以支持肝细胞和非实质细胞。以其当前形式，肝腺泡微生理系统最适合低至中等通量的急性到慢性研究，包括肝病模型、为临床前研究确定化合物优先级、在结构活性关系（SAR）项目中优化化学结构，以及在初始剂量范围的递增剂量研究中。影响声明氧气分区是肝功能的一个关键方面。需要一个人微生理系统来研究分区对一系列可通过实验操作的肝功能的影响。由于氧气分区在肝脏中具有如此多样的生理效应，我们开发并提出了一种计算建模和测量氧气的方法，该方法可以很容易地在所有MPS模型中实施。我们已将此方法应用于肝脏MPS中，在其中我们能够控制各个装置中的氧合作用，并证明MPS中依赖分区的肝细胞功能概括了体内肝脏生理学中已知的情况。我们相信这一进展允许对分区在肝脏代谢和疾病中的作用进行深入的实验研究。此外，随着研究人员从聚二甲基硅氧烷（PDMS）转向塑料和玻璃装置，将需要对氧气张力进行建模和测量。
