Organs-on-a-chip最新文献

Development of three dimensional (3D) in vitro models to realistically recapitulate tumor microenvironment has the potential to improve translatability of anti-cancer drugs at the preclinical stage. To capture the in vivo complexity, these in vitro models should minimally incorporate the 3D interactions between multiple cell types, cellular structures such as vasculature and extracellular matrices. Here, we utilised microfluidic platforms to study the effect of various natural hydrogels (fibrin, collagen, Matrigel) and presence of tumor spheroids on the 3D vascularisation morphology. Various extracellular matrix (ECM) compositions impacted the vessel morphology while near the tumor spheroids the vessel diameter was considerably smaller for all different ECM compositions. Strikingly, cancer cells could enter the microvessel lumens (i.e. intravasate) only when the ECM was comprised of all the three types of hydrogels which increased the physical contact between the microvessels and the tumour spheroids. Our findings highlight the role of ECM composition in modulating the intravasation capacity of tumours.

3-dimensional (3D) cell cultures better mimic natural environment of cells than 2-dimensional (2D) cell cultures to obtain in vivo like inter and intracellular responses. However, third dimension brings complexity to cell culture. Therefore, high-resolution/high-content screening in 3D is one of the most important challenges with this type of cell cultures. Although optical monitoring techniques, well-established in 2D area, are enhanced to monitor 3D cell cultures, they are generally endpoint, static, time inefficient, and labor intensive. Alternatively, electrical sensing can become a solution to achieve dynamic, real-time, and label-free monitoring of cells in both 2D and 3D cell cultures. Developments in electrical monitoring of cell culture have led to novel approaches, proposed by adapting fundamentals of 2D electrical techniques to 3D to obtain high spatiotemporal systems. In this review, we classified these approaches into five main groups: (i) 3D impedance measurement approach (ii) electrical impedance tomography, (iii) 3D microelectrode array approach, (iv) 3D nanoelectronics scaffold approach, and (v) microphysiometry. We also defined the challenges in the adaptation of electrical monitoring techniques to 3D cultures and explained possible solutions in terms of specific applications and technical point of views, including methods particular to our group. In conclusion, 3D electrical monitoring in cell cultures is considerably challenging but highly accelerated recently by significant advances of microfabrication technology, bioengineering, and material science. Novel approaches reviewed here have a lot of potential and offer opportunities for further developments to find solutions, fit to serve the (bio)medical needs.

Transition to extrauterine life results in a surge of catecholamines necessary for increased cardiovascular, respiratory, and metabolic activity. Mechanisms mediating adrenomedullary catecholamine release are poorly understood. Important mechanistic insight is provided by newborns delivered by cesarean section or subjected to prenatal nicotine or opioid exposure, demonstrating impaired release of adrenomedullary catecholamines. To investigate mechanisms regulating adrenomedullary innervation, we developed compartmentalized 3D microphysiological systems (MPS) by exploiting GelPins, capillary pressure barriers between cell-laden hydrogels. The MPS comprises discrete cultures of adrenal chromaffin cells and preganglionic sympathetic neurons within a contiguous bioengineered microtissue. Using this model, we demonstrate that adrenal chromaffin innervation plays a critical role in hypoxia-mediated catecholamine release. Opioids and nicotine were shown to affect adrenal chromaffin cell response to a reduced oxygen environment, but neurogenic control mechanisms remained intact. GelPin containing MPS represent an inexpensive and highly adaptable approach to study innervated organ systems and improve drug screening platforms.

Tumour cell proliferation, metabolism and treatment response depend on the dynamic interaction of the tumour cells with other cellular components and physicochemical gradients present in the tumour microenvironment. Traditional experimental approaches used to investigate the dynamic tumour tissue face a number of limitations, such as lack of biological relevance for the tumour microenvironment and the difficulty to precisely control fluctuating internal conditions, for example in oxygen and nutrients. The arrival of advanced in vitro models represents an alternative approach for modelling the tumour microenvironment using cutting-edge technologies, such as microfabrication. Advanced model systems provide a promising platform for modelling the physiochemical conditions of the tumour microenvironment in a well-controlled manner. Amongst others, advanced in vitro models aim to recreate gradients of oxygen, nutrients and endogenous chemokines, and cell proliferation. Furthermore, the establishment of mechanical cues within such models, e.g., flow and extracellular matrix properties that influence cellular behaviour, are active research areas. These model systems aim to maintain tumour cells in an environment that resembles in vivo conditions. A prominent example of such a system is the microfluidic tumour-on-chip model, which aims to precisely control the local chemical and physical environment that surrounds the tumour cells. In addition, these models also have the potential to recapitulate environmental conditions in isolation or in combination. This enables the analysis of the dynamic interactions between different conditions and their potentially synergistic effects on tumour cells. In this review, we will discuss the various gradients present within the tumour microenvironment and the effects they exert on tumour cells. We will further highlight the challenges and limitations of traditional experimental models in modelling these gradients. We will outline recent achievements in advanced in vitro models with a particular focus on tumour-on-chip systems. We will also discuss the future of these models in cancer research and their contribution to developing more biologically relevant models for cancer research.

With cardiac disease a reigning problem in the world, the need for accurate and high-throughput drug testing is paramount. 3D cardiac tissues are promising models, as they can recapitulate the cell-cell, cell-matrix, and cell-tissue interactions that impact response to a drug. Using an in-house developed micro-continuous optical printing system, we created a cardiac micro-tissue in mere seconds with microscale alignment cues in a hydrogel scaffold that is small enough to fit in a 96-well plate. The 3D printed, asymmetric, cantilever-based tissue scaffold allows one to directly measure the deformation produced by the beating micro-tissue. After 7 days, the micro-tissue exhibited a high level of sarcomere organization and a significant increase in maturity marker expression. The cardiac micro-tissues were validated against two representative drugs, isoproterenol and verapamil at various doses, showing corresponding and measurable changes in beating frequency and displacement. Such rapidly bioprinted cardiac micro-tissues in a multi-well plate offer a promising solution for high-throughput screening in drug discovery.

Nonalcoholic fatty liver disease (NAFLD) is one of the most common chronic liver conditions, and its treatment involves curing the patients without liver transplantation. Understanding the mechanism of NAFLD initiation and progression would enable the development of new diagnostic tools and drugs; however, until now, the underlying mechanisms of this condition remain largely unknown owing to the lack of experimental settings that can simplify the complicated NAFLD process in vitro. Microphysiological systems (MPSs) have long been used to recapture human pathophysiological conditions in vitro for applications in drug discovery. However, polydimethylsiloxane (PDMS) is used in most of these MPSs as the structural material; it absorbs hydrophobic molecules, such as free fatty acids (FFAs), which are the key components that initiate NAFLD. Therefore, the current PDMS-based MPSs cannot be directly applied to in vitro NAFLD modeling. In this work, we present an in vitro NAFLD model with an MPS made of cyclo-olefin polymer (COP), namely COP-MPS, to prevent absorption of FFAs. We demonstrated the induction of NAFLD-like phenotype in HepaRG hepatocyte-like cells cultured in the COP-MPS by treatment with FFAs. The FFAs induced lipid accumulation in the HepaRG cells, resulting in inactivation of the apoptotic cells. We believe that the proposed COP-MPS can contribute toward the investigation of NAFLD mechanisms and identification of new drugs to prevent the progression of liver disease and thus avoid liver transplantation.

Drug discovery faces challenges due to the lack of proper preclinical tests, including conventional cell cultures and animal studies. Organ-on-a-chip devices can mimic the whole-body response to therapeutics by fluidically connecting microscale cell cultures and generating a realistic model of human organs of interest. Here, we describe a pumpless heart/liver-on-a-chip (HLC) using the HepG2 hepatocellular carcinoma cells and H9c2 rat cardiomyocytes to reproduce the cardiotoxicity induced by doxorubicin (DOX) in vitro. Cell studies confirmed the high viability of both cells up to 5 days of culture in HLC. The developed device demonstrated more significant damage to heart cells within the HLC than conventional static 3D culture in the case of DOX treatment, which is because of exposure of cells to both the parent drug and its cardiotoxic metabolite, Doxorubicinol (DOXOL). Our designed HLC device represents a unique approach to assess the off-target toxicity of drugs and their metabolites, which will eventually improve current preclinical studies.

This report demonstrates the ability of a microfluidic device to maintain human Graves' disease tissue enabling the isolation and characterisation of Graves' disease specific exosomes. Graves' disease (n = 7) and non-Graves’ disease (Hashimoto's thyroiditis, n = 3; follicular adenoma, n = 1) human tissue was incubated in a microfluidic device for 6 days ± dexamethasone or methimazole and effluent was analysed for the size and concentration of extracellular vesicles (EV) using nanoparticle tracking analysis. Exosomes were isolated by centrifugation and characterised using Western blotting and qRT-PCR for miRNA-146a and miRNA-155, previously reported to be immunomodulatory. EV were detected in all effluent samples. No difference in concentration was observed in the EV released from Graves' compared to non-Graves’ disease tissue and although the size of EV from Graves' disease tissue was smaller compared to those from non-Graves’ disease tissue, the difference was not consistently significant. No effect of treatment was observed on the size or concentration of EV released. The exosome markers CD63 and CD81 were detectable in 2/5 Graves' disease tissue exosomes and CD63 was also evident in exosomes from a single non-Graves’ sample. miRNA-146a and miRNA-155 were detectable in all samples with no difference between tissue cohorts. Treatment did not influence miRNA expression in exosomes isolated from Graves' disease tissue. Although miRNA-146a and miRNA-155 were both elevated following treatment of non-Graves’ disease tissue with dexamethasone and methimazole, the increase was not significant. This study provides a proof of concept that incubation of tissue on a microfluidic device allows the detection, isolation and characterisation of extracellular vesicles from human tissue biopsies.

This review article describes microfabrication techniques to define chemical, mechanical and structural patterns in hydrogels and how these can be used to prepare in vivo like, i.e. biomimetic, cell culture scaffolds. Hydrogels are attractive materials for 3D cell cultures as they provide ideal culture conditions and they are becoming more prominently used. Single material gels without any modifications do however have their limitation in use and much can be gained by in improving the in vivo resemblance of simple hydrogel cell culture scaffolds. This review article discusses the most commonly used cross-linking strategies used for hydrogel-based culture scaffolds and gives a brief introduction to microfabrication methods that can be used to define chemical, mechanical and structural patterns in hydrogels with micrometre resolution. The review article also describes a selection of literature references using these microfabrication techniques to prepare organ and disease models with controlled cell adhesion, proliferation and migration. It is intended to serve as an introduction to microfabrication of hydrogels and an inspiration for novel interdisciplinary research projects.

Controlled flow within a lab-on-a-chip is a critical element of successfully implementing culture protocols for differentiation and maintenance of stem cell derived neurons in microfluidic devices. There have been a multitude of passive pumping technologies that have been successfully used to control the flow within a lab-on-a-chip. However, most of which were only able to generate flow for very few minutes, while the most successful ones were able to achieve around an hour of flow. This is not convenient for culture protocols requiring constant flow, as hourly media changes will have to be conducted. Herein, we present a design technique adapted for the OrganoPlate, a cell culture plate fully compatible with laboratory automation, which allows its redimension to achieve over 24 h of flow. This technique uses a similarity model of a target cell type and a simple fluid flow mathematical prediction model to iterate to the optimum dimensions within some manufacturing constraints. This technique has the potential to be applied to many cell types to generate optimum design for their culture. We applied this technique to design a 3D microfluidic device, dynamically optimised for neuronal cell culture.