Mandy B. Esch

The Role of Body-on-a-Chip Devices in Drug and Toxicity Studies

High-quality, in vitro screening tools are essential in identifying promising compounds during drug development. Tests with currently used cell-based assays provide an indication of a compounds potential therapeutic benefits to the target tissue, but not to the whole body. Data obtained with animal models often cannot be extrapolated to humans. Multicompartment microfluidic-based devices, particularly those that are physical representations of physiologically based pharmacokinetic (PBPK) models, may contribute to improving the drug development process. These scaled-down devices, termed micro cell culture analogs (μCCAs) or body-on-a-chip devices, can simulate multitissue interactions under near-physiological fluid flow conditions and with realistic tissue-to-tissue size ratios. Because the device can be used with both animal and human cells, it can facilitate cross-species extrapolation. Used in conjunction with PBPK models, the devices permit an estimation of effective concentrations that can be used for studies with animal models or predict the human response. The devices also provide a means for relatively high-throughput screening of drug combinations and, when utilized with a patients tissue sample, an opportunity for individualized medicine. Here we review efforts made toward the development of microfabricated cell culture systems and give examples that demonstrate their potential use in drug development, such as identifying synergistic drug interactions as well as simulating multiorgan metabolic interactions. In addition to their use in drug development, the devices also can be used to estimate the toxicity of chemicals as occupational hazards and environmental contaminants.

TEER measurement techniques for in vitro barrier model systems

Transepithelial/transendothelial electrical resistance (TEER) is a widely accepted quantitative technique to measure the integrity of tight junction dynamics in cell culture models of endothelial and epithelial monolayers. TEER values are strong indicators of the integrity of the cellular barriers before they are evaluated for transport of drugs or chemicals. TEER measurements can be performed in real time without cell damage and generally are based on measuring ohmic resistance or measuring impedance across a wide spectrum of frequencies. The measurements for various cell types have been reported with commercially available measurement systems and also with custom-built microfluidic implementations. Some of the barrier models that have been widely characterized using TEER include the blood–brain barrier (BBB), gastrointestinal (GI) tract, and pulmonary models. Variations in these values can arise due to factors such as temperature, medium formulation, and passage number of cells. The aim of this article is to review the different TEER measurement techniques and analyze their strengths and weaknesses, determine the significance of TEER in drug toxicity studies, examine the various in vitro models and microfluidic organs-on-chips implementations using TEER measurements in some widely studied barrier models (BBB, GI tract, and pulmonary), and discuss the various factors that can affect TEER measurements.

Characterization of a gastrointestinal tract microscale cell culture analog used to predict drug toxicity

The lining of the gastrointestinal (GI) tract is the largest surface exposed to the external environment in the human body. One of the main functions of the small intestine is absorption, and intestinal absorption is a route used by essential nutrients, chemicals, and pharmaceuticals to enter the systemic circulation. Understanding the effects of digestion on a drug or chemical, how compounds interact with and are absorbed through the small intestinal epithelium, and how these compounds affect the rest of the body is critical for toxicological evaluation. Our goal is to create physiologically realistic in vitro models of the human GI tract that provide rapid, inexpensive, and accurate predictions of the bodys response to orally delivered drugs and chemicals. Our group has developed an in vitro microscale cell culture analog (µCCA) of the GI tract that includes digestion, a mucus layer, and physiologically realistic cell populations. The GI tract µCCA, coupled with a multi‐chamber silicon µCCA representing the systemic circulation, is described and challenged with acetaminophen. Proof of concept experiments showed that acetaminophen passes through and is metabolized by the in vitro intestinal epithelium and is further metabolized by liver cells, resulting in liver cell toxicity in a dose‐dependent manner. The µCCA response is also consistent with in vivo measurements in mice. The system should be broadly useful for studies on orally delivered drugs or ingestion of chemicals with potential toxicity. Biotechnol. Bioeng. 2009; 104: 193–205

Surface topography induces 3D self-orientation of cells and extracellular matrix resulting in improved tissue function

The organization of cells and extracellular matrix (ECM) in native tissues plays a crucial role in their functionality. However, in tissue engineering, cells and ECM are randomly distributed within a scaffold. Thus, the production of engineered-tissue with complex 3D organization remains a challenge. In the present study, we used contact guidance to control the interactions between the material topography, the cells and the ECM for three different tissues, namely vascular media, corneal stroma and dermal tissue. Using a specific surface topography on an elastomeric material, we observed the orientation of a first cell layer along the patterns in the material. Orientation of the first cell layer translates into a physical cue that induces the second cell layer to follow a physiologically consistent orientation mimicking the structure of the native tissue. Furthermore, secreted ECM followed cell orientation in every layer, resulting in an oriented self-assembled tissue sheet. These self-assembled tissue sheets were then used to create 3 different structured engineered-tissue: cornea, vascular media and dermis. We showed that functionality of such structured engineered-tissue was increased when compared to the same non-structured tissue. Dermal tissues were used as a negative control in response to surface topography since native dermal fibroblasts are not preferentially oriented in vivo. Non-structured surfaces were also used to produce randomly oriented tissue sheets to evaluate the impact of tissue orientation on functional output. This novel approach for the production of more complex 3D tissues would be useful for clinical purposes and for in vitro physiological tissue model to better understand long standing questions in biology.

Multi-Organ toxicity demonstration in a functional human in vitro system composed of four organs

We report on a functional human model to evaluate multi-organ toxicity in a 4-organ system under continuous flow conditions in a serum-free defined medium utilizing a pumpless platform for 14 days. Computer simulations of the platform established flow rates and resultant shear stress within accepted ranges. Viability of the system was demonstrated for 14 days as well as functional activity of cardiac, muscle, neuronal and liver modules. The pharmacological relevance of the integrated modules were evaluated for their response at 7 days to 5 drugs with known side effects after a 48 hour drug treatment regime. The results of all drug treatments were in general agreement with published toxicity results from human and animal data. The presented phenotypic culture model exhibits a multi-organ toxicity response, representing the next generation of in vitro systems, and constitutes a step towards an in vitro “human-on-a-chip” assay for systemic toxicity screening.

Using physiologically-based pharmacokinetic-guided ''body-on-a-chip'' systems to predict mammalian response to drug and chemical exposure

The continued development of in vitro systems that accurately emulate human response to drugs or chemical agents will impact drug development, our understanding of chemical toxicity, and enhance our ability to respond to threats from chemical or biological agents. A promising technology is to build microscale replicas of humans that capture essential elements of physiology, pharmacology, and/or toxicology (microphysiological systems). Here, we review progress on systems for microscale models of mammalian systems that include two or more integrated cellular components. These systems are described as a “body-on-a-chip”, and utilize the concept of physiologically-based pharmacokinetic (PBPK) modeling in the design. These microscale systems can also be used as model systems to predict whole-body responses to drugs as well as study the mechanism of action of drugs using PBPK analysis. In this review, we provide examples of various approaches to construct such systems with a focus on their physiological usefulness and various approaches to measure responses (e.g. chemical, electrical, or mechanical force and cellular viability and morphology). While the goal is to predict human response, other mammalian cell types can be utilized with the same principle to predict animal response. These systems will be evaluated on their potential to be physiologically accurate, to provide effective and efficient platform for analytics with accessibility to a wide range of users, for ease of incorporation of analytics, functional for weeks to months, and the ability to replicate previously observed human responses.

Integration of in silico and in vitro platforms for pharmacokinetic–pharmacodynamic modeling

Importance of the field: Pharmacokinetic–pharmacodynamic (PK-PD) modeling enables quantitative prediction of the dose–response relationship. Recent advances in microscale technology enabled researchers to create in vitro systems that mimic biological systems more closely. Combination of mathematical modeling and microscale technology offers the possibility of faster, cheaper and more accurate prediction of the drugs effect with a reduced need for animal or human subjects. Areas covered in this review: This article discusses combining in vitro microscale systems and PK-PD models for improved prediction of drugs efficacy and toxicity. First, we describe the concept of PK-PD modeling and its applications. Different classes of PK-PD models are described. Microscale technology offers an opportunity for building physical systems that mimic PK-PD models. Recent progress in this approach during the last decade is summarized. What the reader will gain: This article is intended to review how microscale technology combined with cell cultures, also known as ‘cells-on-a-chip’, can confer a novel aspect to current PK-PD modeling. Readers will gain a comprehensive knowledge of PK-PD modeling and ‘cells-on-a-chip’ technology, with the prospect of how they may be combined for synergistic effect. Take home message: The combination of microscale technology and PK-PD modeling should contribute to the development of a novel in vitro/in silico platform for more physiologically-realistic drug screening.

Characterization of In Vitro Endothelial Linings Grown Within Microfluidic Channels

In vivo, endothelial cells grow on the inner surface of blood vessels and are shaped to conform to the vessels geometry. In the smallest vessels this shape entails substantial bending within each cell. Microfabricated channels can replicate these small-scale geometries, but endothelial cells grown within them have not been fully characterized. In particular, the presence of focal adhesions and adherens junctions in endothelial cells grown in microchannels with corners has not been confirmed. We have fabricated square microfluidic channels (50 μm wide, 50 μm deep) and semicircular microfluidic channels (60 μm wide, 45 μm deep) in polydimethylsiloxane and cultured human umbilical vein endothelial cells (HUVEC) within them. Immunofluorescent staining and three-dimensional reconstruction of image stacks taken with confocal microscopy confirmed that HUVEC are capable of forming adherens junctions on all channel walls in both channel geometries, including the sidewalls of square profile channels. The presence of shear stress is critical for the cells to form focal adhesions within both channel geometries. Shear stress is also responsible for the conforming of HUVEC to the channel walls and produces a square cross-sectional geometry of in vitro endothelial linings within square profile channels. Thus, geometry and applied shear stress are important design criteria for the development of in vitro endothelial linings of microvessels.

Dielectric properties of alginate beads and bound water relaxation studied by electrorotation.

The electrical and dielectric properties of Ba2+ and Ca2+ cross-linked alginate hydrogel beads were studied by means of single-particle electrorotation. The use of microstructured electrodes allowed the measurements to be performed over a wide range of medium conductivity from about 5 mS/m to 1 S/m. Within a conductivity range, the beads exhibited measurable electrorotation response at frequencies above 0.2 MHz with two well-resolved co- and antifield peaks. With increasing medium conductivity, both peaks shifted toward higher frequency and their magnitudes decreased greatly. The results were analyzed using various dielectric models that consider the beads as homogeneous spheres with conductive loss and allow the complex rotational behavior of beads to be explained in terms of conductivity and permittivity of the hydrogel. The rotation spectra could be fitted very accurately by assuming (a) a linear relationship between the internal hydrogel conductivity and the medium conductivity, and (b) a broad internal dispersion of the hydrogel centered between 20 and 40 MHz. We attribute this dispersion to the relaxation of water bound to the polysaccharide matrix of the beads. The dielectric characterization of alginate hydrogels is of enormous interest for biotechnology and medicine, where alginate beads are widely used for immobilization of cells and enzymes, for drug delivery, and as microcarriers for cell cultivation.

Design optimization of liquid-phase flow patterns for microfabricated lung on a chip.

Microreactors experience significant deviations from plug flow due to the no-slip boundary condition at the walls of the chamber. The development of stagnation zones leads to widening of the residence time distribution at the outlet of the reactor. A hybrid design optimization process that combines modeling and experiments has been utilized to minimize the width of the residence time distribution in a microreactor. The process was used to optimize the design of a microfluidic system for an in vitro model of the lung alveolus. Circular chambers to accommodate commercial membrane supported cell constructs are a particularly challenging geometry in which to achieve a uniform residence time distribution. Iterative computational fluid dynamics (CFD) simulations were performed to optimize the microfluidic structures for two different types of chambers. The residence time distributions of the optimized chambers were significantly narrower than those of non-optimized chambers, indicating that the final chambers better approximate plug flow. Qualitative and quantitative visualization experiments with dye indicators demonstrated that the CFD results accurately predicted the residence time distributions within the bioreactors. The results demonstrate that such a hybrid optimization process can be used to design microreactors that approximate plug flow for in vitro tissue engineered systems. This technique has broad application for optimization of microfluidic body-on-a-chip systems for drug and toxin studies.