Control of oxygen tension recapitulates zone-specific functions in human liver microphysiology systems

Abstract

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.

Pre-clinical and clinical investigations of metabolic zonation in liver diseases: The potential of microphysiology systems

“…74 Various zone-specific physiological measurements including acetaminophen toxicity, glycolysis, oxidative phosphorylation, and protein secretions were consistent with measurements in animals ( Table 1). In particular, the human-based LAMPS models can be used to exploit the role of canonical Wnt/b-catenin signaling in liver zonation to control zone 3 hepatocyte gene expression to generate human-based models of the normal liver and to study the importance of zonation on liver disease (Figures 1 and 3).…”

“…Several platform technologies have demonstrated zonation, 67,68,70,71,75,69,[72][73][74]78 and three have demonstrated zone-specific toxicity. 68,71,74 Further investigations are needed to understand how these models can best be used to interrogate the role zonation may have on druginduced liver injury and in disease initiation and progression.…”

“…In particular, the human-based LAMPS models can be used to exploit the role of canonical Wnt/b-catenin signaling in liver zonation to control zone 3 hepatocyte gene expression to generate human-based models of the normal liver and to study the importance of zonation on liver disease (Figures 1 and 3). 74 …”

3D Primary Hepatocyte Culture Systems for Analyses of Liver Diseases, Drug Metabolism, and Toxicity: Emerging Culture Paradigms and Applications

“…In an attempt to mimic these conditions, devices have been developed that permit perfusion of 2D hepatocyte cultures and such experimental setups have been reported to improve hepatic functions, such as urea and albumin secretion . Furthermore, perfusion allows the establishment of oxygen gradients and hepatic zonation, resulting in graded CYP expression and metabolism, as well as the recapitulation of zone‐specific patterns of drug‐induced liver injury (DILI) …”

Tissue Engineering in Liver Regenerative Medicine: Insights into Novel Translational Technologies

“…Substantial advances are developing in metabolic liver zonation to study hepatocyte functions and zone-specific toxicity [115]. The zonation studies can be performed in microtiter plate in static cultures [116], bioreactors [117] and microfluidic systems [118].…”