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The availability of high-frequency, real-time measurements of the concentrations of specific metabolites in cell culture systems will enable a deeper understanding of cellular metabolism and facilitate the application of good laboratory practice standards in cell culture protocols. However, currently available approaches to this end either are constrained to single-time-point and single-parameter measurements or are limited in the range of detectable analytes. Electrochemical aptamer-based (EAB) biosensors have demonstrated utility in real-time monitoring of analytes in vivo in blood and tissues. Here, we characterize a pH-sensing capability of EAB sensors that is independent of the specific target analyte of the aptamer sequence. We applied this dual-purpose EAB to the continuous measurement of pH and phenylalanine in several in vitro cell culture settings. The miniature EAB sensor that we developed exhibits rapid response times, good stability, high repeatability, and biologically relevant sensitivity. We also developed and characterized a leak-free reference electrode that mitigates the potential cytotoxic effects of silver ions released from conventional reference electrodes. Using the resulting dual-purpose sensor, we performed hourly measurements of pH and phenylalanine concentrations in the medium superfusing cultured epithelial tumor cell lines (A549, MDA-MB-23) and a human fibroblast cell line (MRC-5) for periods of up to 72 h. Our scalable technology may be multiplexed for high-throughput monitoring of pH and multiple analytes in support of the broad metabolic qualification of microphysiological systems.

This research introduces a vascular phenotypic and proteomic analysis (VPT) platform designed to perform high-throughput experiments on vascular development. The VPT platform utilizes an open-channel configuration that facilitates angiogenesis by precise alignment of endothelial cells, allowing for a 3D morphological examination and protein analysis. We study the effects of antiangiogenic agents─bevacizumab, ramucirumab, cabozantinib, regorafenib, wortmannin, chloroquine, and paclitaxel─on cytoskeletal integrity and angiogenic sprouting, observing an approximately 50% reduction in sprouting at higher drug concentrations. Precise LC-MS/MS analyses reveal global protein expression changes in response to four of these drugs, providing insights into the signaling pathways related to the cell cycle, cytoskeleton, cellular senescence, and angiogenesis. Our findings emphasize the intricate relationship between cytoskeletal alterations and angiogenic responses, underlining the significance of integrating morphological and proteomic data for a comprehensive understanding of angiogenesis. The VPT platform not only advances our understanding of drug impacts on vascular biology but also offers a versatile tool for analyzing proteome and morphological features across various models beyond blood vessels.

Recent advances have made modeling human small intestines in vitro possible, but it remains a challenge to recapitulate fully their structural and functional characteristics. We suspected interstitial flow within the intestine, powered by circulating blood plasma during embryonic organogenesis, to be a vital factor. We aimed to construct an in vivo-like multilayered small intestinal tissue by incorporating interstitial flow into the system and, in turn, developed the micro-small intestine system by differentiating definitive endoderm and mesoderm cells from human pluripotent stem cells simultaneously on a microfluidic device capable of replicating interstitial flow. This approach enhanced cell maturation and led to the development of a three-dimensional small intestine-like tissue with villi-like epithelium and an aligned mesenchymal layer. Our micro-small intestine system not only overcomes the limitations of conventional intestine models but also offers a unique opportunity to gain insights into the detailed mechanisms underlying intestinal tissue development., Competing Interests: Declaration of interests K.T. and S.D. are inventors on JP patent application no. 2023-132372 “Construction of micro-small intestine systems”. K.T. receives research funds for developing micro-small intestine systems under the contract with Kanto Chemical Co., Inc. M.I. is a scientific adviser for xFOREST Therapeutics without salary., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Microphysiological systems have attracted attention because of their use in drug screening. However, it is challenging to measure cell functions in real time using a device. In this study, we developed a cell culture device using a porous membrane electrode for in situ electrochemical glucose measurements for cell analysis. First, a porous membrane electrode was fabricated and electrochemically evaluated for enzyme-free glucose measurement. Subsequently, the glucose uptake of MCF-7 spheroids was evaluated using living spheroids, fixed spheroids, supernatants, and glucose transporter inhibitor-treated spheroids. Conventionally, the direct optical measurement of glucose uptake requires fluorescence-labeled glucose derivatives. In addition, the glucose uptake can be evaluated by measuring the glucose concentration in the medium by optical or electrochemical measurements. However, glucose needs to be consumed in the entire cell culture medium, which needs a long culture time. In contrast, our system can measure glucose in approximately 5 min without any labels because of in situ electrochemical measurements. This system can be used for in situ measurements in in vitro cell culture systems, including organ-on-a-chip for drug screening.

Diabetes mellitus (DM) has substantial global implications and contributes to vascular inflammation and the onset of atherosclerotic cardiovascular diseases. However, translating the findings from animal models to humans has inherent limitations, necessitating a novel platform. Therefore, herein, an arterial model is established using a microphysiological system. This model successfully replicates the stratified characteristics of human arteries by integrating collagen, endothelial cells (ECs), and vascular smooth muscle cells (VSMCs). Perfusion via a peristaltic pump shows dynamic characteristics distinct from those of static culture models. High glucose, advanced glycation end products (AGEs), and interleukin-1 beta are employed to stimulate diabetic conditions, resulting in notable cellular changes and different levels of cytokines and nitric oxide. Additionally, the interactions between the disease models and oxidized low-density lipoproteins (LDL) are examined. Finally, the potential therapeutic effects of metformin, atorvastatin, and diphenyleneiodonium are investigated. Metformin and diphenyleneiodonium mitigate high-glucose- and AGE-associated pathological changes, whereas atorvastatin affects only the morphology of ECs. Altogether, the arterial model represents a pivotal advancement, offering a robust and insightful platform for investigating cardiovascular diseases and their corresponding drug development., (© 2024 The Authors. Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

Human organoids have the potential to revolutionize in vitro disease modeling by providing multicellular architecture and function that are similar to those in vivo. This innovative and evolving technology, however, still suffers from assay throughput and reproducibility to enable high-throughput screening (HTS) of compounds due to cumbersome organoid differentiation processes and difficulty in scale-up and quality control. Using organoids for HTS is further challenged by the lack of easy-to-use fluidic systems that are compatible with relatively large organoids. Here, these challenges are overcome by engineering "microarray three-dimensional (3D) bioprinting" technology and associated pillar and perfusion plates for human organoid culture and analysis. High-precision, high-throughput stem cell printing, and encapsulation techniques are demonstrated on a pillar plate, which is coupled with a complementary deep well plate and a perfusion well plate for static and dynamic organoid culture. Bioprinted cells and spheroids in hydrogels are differentiated into liver and intestine organoids for in situ functional assays. The pillar/perfusion plates are compatible with standard 384-well plates and HTS equipment, and thus may be easily adopted in current drug discovery efforts., (© 2023 The Authors. Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

The liver microphysiological system (MPS) model is an in-vitro culture method that mimics physiological blood flow, which enhances basal cellular functions. However, the liver MPS model has not been tested in the preclinical stage because of its obscure utility. It can overcome the major problem of conventional systems--rapid loss of mitochondrial activity in cultured hepatocytes due to limited oxygen supply--by supplying oxygen to cultured hepatocytes using a perfusion device. In this study, we developed a new perfusion culture system that can detect mitochondrial toxicity. Primary mouse hepatocytes were cultured under perfusion condition for 48 hr. The hepatocytes showed increased oxygen consumption and reduced lactate release. These results indicated that the ATP-production pathway was switched from glycolysis to mitochondrial oxidative phosphorylation in the perfusion culture system. Furthermore, ATP levels were considerably reduced in the perfusion culture system after exposure to phenformin, a mitochondrial complex I inhibitor. To summarize, the perfusion culture system could improve the mitochondrial activity in primary mouse hepatocytes, and thus, has potential implications in the detection of mitochondrial toxicity. [ABSTRACT FROM AUTHOR]

Microphysiological systems (MPSs), based on microfabrication technologies and cell culture, can faithfully recapitulate the complex physiology of various tissues. However, 3D tissues formed using MPS have limitations in size and accessibility; their use in regenerative medicine is, therefore, still challenging. Here, an MPS‐inspired scale‐up vascularized engineered tissue construct that can be used in regenerative medicine is designed. Endothelial cell‐laden hydrogels are sandwiched between two through‐hole membranes. The microhole array in the through‐hole membranes enables the molecular transport across the hydrogel layer, allowing long‐term cell culture. Furthermore, the time‐controlled delamination of through‐hole membranes enables the harvesting of cell‐cultured hydrogel constructs without damaging the capillary network. Importantly, when the tissue constructs are implanted in a mouse ischemic model, they protect against necrosis and promoted functional recovery to a greater extent than implanted cells, hydrogels, and simple gel–cell mixtures. [ABSTRACT FROM AUTHOR]

BACKGROUND/AIM: Tissue pathogenesis of aortic valve (AV) stenosis is research focus in cardiac surgery. Model limitations of conventional 2D culture of human or porcine valvular interstitial/endothelial cells (VIC/VECs) isolated from aortic valve tissues but also limited ability of (small) animal models to reflect human (patho)physiological situation in AV position raise the need to establish an in vitro setup using AV tissues. Resulting aim is to approximate (patho)physiological conditions in a dynamic pulsatile Microphysiological System (MPS) to culture human and porcine AV tissue with preservation of tissue viability but also defined ECM composition. MATERIALS/METHODS: A tissue incubation chamber (TIC) was designed to implement human or porcine tissues (3×5 mm2) in a dynamic pulsatile culture in conventional cell culture ambience in a MPS. Cell viability assays based on lactate dehydrogenase (LDH)-release or resazurin-conversion were tested for applicability in the system and applied for a culture period of 14 days with interval evaluation of tissue viability on every other day. Resazurin-assay setup was compared in static vs. dynamic culture using varying substance saturation settings (50–300μM), incubation times and tissue masses and was consequently adapted. RESULTS: Sterile dynamic culture of human and porcine AV tissue segments was established at a pulsatile flow rate range of 0.9–13.4μl/s. Implementation of tissues was realized by stitching the material in a thermoplastic polyurethane (TPU)-ring and insertion in the TIC-MPS-system. Culture volume of 2 ml caused LDH dilution not detectable in standard membrane integrity assay setup. Therefore, detection of resazurin-conversion of viable tissue was investigated. Optimal incubation time for viability conversion was determined at two hours at a saturated concentration of 300μM resazurin. Measurement in static conditions was shown to offer comparable results as dynamic condition but allowing optimal handling and TIC sterilization protocols for long term culture. Preliminary results revealed favourable porcine AV tissue viability over a 14 day period confirmed via resazurin-assay comparing statically cultured tissue counterparts. CONCLUSIONS: Human and porcine AV tissue can be dynamically cultured in a TIC-MPS with monitoring of tissue viability using an adapted resazurin-assay setup. Preliminary results reveal advantageous viability of porcine AV tissues after dynamic TIC-MPS culture compared to static control. [ABSTRACT FROM AUTHOR]

Existing first-line cancer therapies often fail to cope with the heterogeneity and complexity of cancers, so that new therapeutic approaches are urgently needed. Among novel alternative therapies, adoptive cell therapy (ACT) has emerged as a promising cancer treatment in recent years. The limited clinical applications of ACT, despite its advantages over standard-of-care therapies, can be attributed to (i) time-consuming and cost-intensive procedures to screen for potent anti-tumor immune cells and the corresponding targets, (ii) difficulties to translate in-vitro and animal-derived in-vivo efficacies to clinical efficacy in humans, and (iii) the lack of systemic methods for the safety assessment of ACT. Suitable experimental models and testing platforms have the potential to accelerate the development of ACT. Immunocompetent microphysiological systems (iMPS) are microfluidic platforms that enable complex interactions of advanced tissue models with different immune cell types, bridging the gap between in-vitro and in-vivo studies. Here, we present a proof-of-concept iMPS that supports a triple culture of three-dimensional (3D) colorectal tumor microtissues, 3D cardiac microtissues, and human-derived natural killer (NK) cells in the same microfluidic network. Different aspects of tumor-NK cell interactions were characterized using this iMPS including: (i) direct interaction and NK cell-mediated tumor killing, (ii) the development of an inflammatory milieu through enrichment of soluble pro-inflammatory chemokines and cytokines, and (iii) secondary effects on healthy cardiac microtissues. We found a specific NK cell-mediated tumor-killing activity and elevated levels of tumor- and NK cell-derived chemokines and cytokines, indicating crosstalk and development of an inflammatory milieu. While viability and morphological integrity of cardiac microtissues remained mostly unaffected, we were able to detect alterations in their beating behavior, which shows the potential of iMPS for both, efficacy and early safety testing of new candidate ACTs. [ABSTRACT FROM AUTHOR]

Existing first-line cancer therapies often fail to cope with the heterogeneity and complexity of cancers, so that new therapeutic approaches are urgently needed. Among novel alternative therapies, adoptive cell therapy (ACT) has emerged as a promising cancer treatment in recent years. The limited clinical applications of ACT, despite its advantages over standard-of-care therapies, can be attributed to (i) time-consuming and cost-intensive procedures to screen for potent anti-tumor immune cells and the corresponding targets, (ii) difficulties to translate in-vitro and animal-derived in-vivo efficacies to clinical efficacy in humans, and (iii) the lack of systemic methods for the safety assessment of ACT. Suitable experimental models and testing platforms have the potential to accelerate the development of ACT. Immunocompetent microphysiological systems (iMPS) are microfluidic platforms that enable complex interactions of advanced tissue models with different immune cell types, bridging the gap between in-vitro and in-vivo studies. Here, we present a proof-of-concept iMPS that supports a triple culture of three-dimensional (3D) colorectal tumor microtissues, 3D cardiac microtissues, and human-derived natural killer (NK) cells in the same microfluidic network. Different aspects of tumor-NK cell interactions were characterized using this iMPS including: (i) direct interaction and NK cell-mediated tumor killing, (ii) the development of an inflammatory milieu through enrichment of soluble pro-inflammatory chemokines and cytokines, and (iii) secondary effects on healthy cardiac microtissues. We found a specific NK cell-mediated tumor-killing activity and elevated levels of tumor- and NK cell-derived chemokines and cytokines, indicating crosstalk and development of an inflammatory milieu. While viability and morphological integrity of cardiac microtissues remained mostly unaffected, we were able to detect alterations in their beating behavior, which shows the potential of iMPS for both, efficacy and early safety testing of new candidate ACTs.

Microfluidic devices are widely used for biomedical applications but there is still a lack of affordable, reliable and user-friendly systems for transferring microfluidic chips from an incubator to a microscope while maintaining physiological conditions when performing microscopy. The presented carrier represents a cost-effective option for sustaining environmental conditions of microfluidic chips in combination with minimizing the device manipulation required for reagent injection, media exchange or sample collection. The carrier, which has the outer dimension of a standard well plate size, contains an integrated perfusion system that can recirculate the media using piezo pumps, operated in either continuous or intermittent modes (50–1000 µl/min). Furthermore, a film resistive heater made from 37 µm-thick copper wires, including temperature feedback control, was used to maintain the microfluidic chip temperature at 37 °C when outside the incubator. The heater characterisation showed a uniform temperature distribution along the chip channel for perfusion flow rates up to 10 µl/min. To demonstrate the feasibility of our platform for long term cell culture monitoring, mouse brain endothelial cells (bEnd.3) were repeatedly monitored for a period of 10 days, demonstrating a system with both the versatility and the potential for long imaging in microphysiological system cell cultures.

Immunosurveillance of the gastrointestinal epithelium by mononuclear phagocytes (MNPs) is essential for maintaining gut health. However, studying the complex interplay between the human gastrointestinal epithelium and MNPs such as dendritic cells (DCs) is difficult, since traditional cell culture systems lack complexity, and animal models may not adequately represent human tissues. Microphysiological systems, or tissue chips, are an attractive alternative for these investigations, because they model functional features of specific tissues or organs using microscale culture platforms that recreate physiological tissue microenvironments. However, successful integration of multiple of tissue types on a tissue chip platform to reproduce physiological cell-cell interactions remains a challenge. We previously developed a tissue chip system, the gut organoid flow chip (GOFlowChip), for long term culture of 3-D pluripotent stem cell-derived human intestinal organoids. Here, we optimized the GOFlowChip platform to build a complex microphysiological immune-cell-epithelial cell co-culture model in order to study DC-epithelial interactions in human stomach. We first tested different tubing materials and chip configurations to optimize DC loading onto the GOFlowChip and demonstrated that DC culture on the GOFlowChip for up to 20 h did not impact DC activation status or viability. However, Transwell chemotaxis assays and live confocal imaging revealed that Matrigel, the extracellular matrix (ECM) material commonly used for organoid culture, prevented DC migration towards the organoids and the establishment of direct MNP-epithelial contacts. Therefore, we next evaluated DC chemotaxis through alternative ECM materials including Matrigel-collagen mixtures and synthetic hydrogels. A polysaccharide-based synthetic hydrogel, VitroGel®-ORGANOID-3 (V-ORG-3), enabled significantly increased DC chemotaxis through the matrix, supported organoid survival and growth, and did not significantly alter DC activation or viability. On the GOFlowChip, DCs that were flowed into the chip migrated rapidly through the V-ORG matrix and reached organoids embedded deep within the chip, with increased interactions between DCs and gastric organoids. The successful integration of DCs and V-ORG-3 embedded gastric organoids into the GOFlowChip platform now permits real-time imaging of MNP-epithelial interactions and other investigations of the complex interplay between gastrointestinal MNPs and epithelial cells in their response to pathogens, candidate drugs and mucosal vaccines.

Immunosurveillance of the gastrointestinal epithelium by mononuclear phagocytes (MNPs) is essential for maintaining gut health. However, studying the complex interplay between the human gastrointestinal epithelium and MNPs such as dendritic cells (DCs) is difficult, since traditional cell culture systems lack complexity, and animal models may not adequately represent human tissues. Microphysiological systems, or tissue chips, are an attractive alternative for these investigations, because they model functional features of specific tissues or organs using microscale culture platforms that recreate physiological tissue microenvironments. However, successful integration of multiple of tissue types on a tissue chip platform to reproduce physiological cell-cell interactions remains a challenge. We previously developed a tissue chip system, the gut organoid flow chip (GOFlowChip), for long term culture of 3-D pluripotent stem cell-derived human intestinal organoids. Here, we optimized the GOFlowChip platform to build a complex microphysiological immune-cell-epithelial cell co-culture model in order to study DC-epithelial interactions in human stomach. We first tested different tubing materials and chip configurations to optimize DC loading onto the GOFlowChip and demonstrated that DC culture on the GOFlowChip for up to 20 h did not impact DC activation status or viability. However, Transwell chemotaxis assays and live confocal imaging revealed that Matrigel, the extracellular matrix (ECM) material commonly used for organoid culture, prevented DC migration towards the organoids and the establishment of direct MNP-epithelial contacts. Therefore, we next evaluated DC chemotaxis through alternative ECM materials including Matrigel-collagen mixtures and synthetic hydrogels. A polysaccharide-based synthetic hydrogel, VitroGel®-ORGANOID-3 (V-ORG-3), enabled significantly increased DC chemotaxis through the matrix, supported organoid survival and growth, and did not significantly alter DC activation or viability. On the GOFlowChip, DCs that were flowed into the chip migrated rapidly through the V-ORG matrix and reached organoids embedded deep within the chip, with increased interactions between DCs and gastric organoids. The successful integration of DCs and V-ORG-3 embedded gastric organoids into the GOFlowChip platform now permits real-time imaging of MNP-epithelial interactions and other investigations of the complex interplay between gastrointestinal MNPs and epithelial cells in their response to pathogens, candidate drugs and mucosal vaccines. [ABSTRACT FROM AUTHOR]

Human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs) have found utility for conducting in vitro drug screening and disease modelling to gain crucial insights into pharmacology or disease phenotype. However, diseases such as atrial fibrillation, affecting >33 M people worldwide, demonstrate the need for cardiac subtype-specific cells. Here, we sought to investigate the base characteristics and pharmacological differences between commercially available chamber-specific atrial or ventricular hiPSC-CMs seeded onto ultra-thin, flexible PDMS membranes to simultaneously measure contractility in a 96 multi-well format. We investigated the effects of GPCR agonists (acetylcholine and carbachol), a Ca2+ channel agonist (S-Bay K8644), an HCN channel antagonist (ivabradine) and K+ channel antagonists (4-AP and vernakalant). We observed differential effects between atrial and ventricular hiPSC-CMs on contractile properties including beat rate, beat duration, contractile force and evidence of arrhythmias at a range of concentrations. As an excerpt of the compound analysis, S-Bay K8644 treatment showed an induced concentration-dependent transient increase in beat duration of atrial hiPSC-CMs, whereas ventricular cells showed a physiological increase in beat rate over time. Carbachol treatment produced marked effects on atrial cells, such as increased beat duration alongside a decrease in beat rate over time, but only minimal effects on ventricular cardiomyocytes. In the context of this chamber-specific pharmacology, we not only add to contractile characterization of hiPSC-CMs but propose a multi-well platform for medium-throughput early compound screening. Overall, these insights illustrate the key pharmacological differences between chamber-specific cardiomyocytes and their application on a multi-well contractility platform to gain insights for in vitro cardiac liability studies and disease modelling. [ABSTRACT FROM AUTHOR]

Human mesenchymal stem cells (MSCs) have attracted attention as a major cellular source for regenerative medicine and preclinical studies. Three-dimensional (3D) cell culture systems can be used to construct physiologically relevant MSC spheroids with enhanced tissue regeneration capacity. However, what aspect of the MSC spheroids retains the regeneration capacity is not yet fully understood. Here, we induced osteoblast differentiation of MSC spheroids and investigated their respiratory activity using electrochemiluminescence (ECL) imaging developed in our previous study. In parallel, the ability of MSC spheroids to induce angiogenesis was investigated in a microfluidic device with a co-culture of endothelial cells. We found that the osteogenic differentiation of MSCs induced changes in their metabolic activity (respiratory activity of the spheroids decreased during differentiation) and lead to the loss of their angiogenesis-inducing ability. In other words, it is suggested that maintaining the stem cell nature with in the MSC spheroids is an important factor for tissue regeneration. In this study, we demonstrated the effective use of ECL imaging and microfluidic systems for the evaluation of MSC spheroids, providing new insights into stem cell tissue engineering. [Display omitted] [ABSTRACT FROM AUTHOR]

Currently, there is no test system, whether in vitro or in vivo , capable of examining all endpoints required for genotoxicity evaluation used in pre-clinical drug safety assessment. The objective of this study was to develop a model which could assess all the required endpoints and possesses robust human metabolic activity, that could be used in a streamlined, animal-free manner. Liver-on-chip (LOC) models have intrinsic human metabolic activity that mimics the in vivo environment, making it a preferred test system. For our assay, the LOC was assembled using primary human hepatocytes or HepaRG cells, in a MPS-T12 plate, maintained under microfluidic flow conditions using the PhysioMimix® Microphysiological System (MPS), and co-cultured with human lymphoblastoid (TK6) cells in transwells. This system allows for interaction between two compartments and for the analysis of three different genotoxic endpoints, i.e. DNA strand breaks (comet assay) in hepatocytes, chromosome loss or damage (micronucleus assay) and mutation (Duplex Sequencing) in TK6 cells. Both compartments were treated at 0, 24 and 45 h with two direct genotoxicants: methyl methanesulfonate (MMS) and ethyl methanesulfonate (EMS), and two genotoxicants requiring metabolic activation: benzo[ a ]pyrene (B[ a ]P) and cyclophosphamide (CP). Assessment of cytochrome activity, RNA expression, albumin, urea and lactate dehydrogenase production, demonstrated functional metabolic capacities. Genotoxicity responses were observed for all endpoints with MMS and EMS. Increases in the micronucleus and mutations (MF) frequencies were also observed with CP, and %Tail DNA with B[ a ]P, indicating the metabolic competency of the test system. CP did not exhibit an increase in the %Tail DNA, which is in line with in vivo data. However, B[ a ]P did not exhibit an increase in the % micronucleus and MF, which might require an optimization of the test system. In conclusion, this proof-of-principle experiment suggests that LOC-MPS technology is a promising tool for in vitro hazard identification genotoxicants. • LOC built with primary hepatocytes or HepaRG and maintained under microfluidic flow. • LOC co-cultured with TK6 cells for comet and micronucleus/mutagenicity analysis. • Statistically significant increases observed for all endpoints with MMS and EMS. • Increases in the micronuclei/mutation frequency with CP and %Tail DNA with B[ a ]P. • This study suggests that LOC-MPS is a promising tool for genotoxicity assessment. [ABSTRACT FROM AUTHOR]

Recently, we could demonstrate positive effects of microfluidic cultivation conditions on maturation of cardiomyocytes derived from human induced pluripotent stem cells (iPS-CMs) in a 2D model. However, 3D cell culture models are much closer to physiological conditions. Combined with microfluidics, 3D systems should resemble the in-vivo conditions even better than standard 2D cultivation. For 3D models, two main technical challenges arise, the tissue integration and sufficient nutrient supply for the cells. This work focuses on concepts for the tissue integration based on a modular approach and different manufacturing technologies as well as using an oxygenator in the microfluidic device to provide sufficient oxygen supply for the cells.

The liver is the most commonly involved organ in metastases from a wide variety of solid tumors. The use of biologically and cellularly complex liver tissue systems have shown that tumor cell behavior and therapeutic responses are modulated within the liver microenvironment and in ways distinct from the behaviors in the primary locations. These microphysiological systems have provided unexpected and powerful insights into the tumor cell biology of metastasis. However, neither the tumor nor the liver exist in an isolated tissue situation, having to function within a complete body and respond to systemic events as well as those in other organs. To examine the influence of one organ on the function of other tissues, microphysiological systems are being linked. Herein, we discuss extending this concept to tumor metastases by integrating complex models of the primary tumor with the liver metastatic environment. In addition, inflammatory organs and the immune system can be incorporated into these multi-organ systems to probe the effects on tumor behavior and cancer treatments. [ABSTRACT FROM AUTHOR]

Metastatic breast cancer remains a largely incurable and fatal disease with liver involvement bearing the worst prognosis. The danger is compounded by a subset of disseminated tumor cells that may lie dormant for years to decades before re-emerging as clinically detectable metastases. Pathophysiological signals can drive these tumor cells to emerge. Prior studies indicated CXCR3 ligands as being the predominant signals synergistically and significantly unregulated during inflammation in the gut-liver axis. Of the CXCR3 ligands, IP-10 (CXCL10) was the most abundant, correlated significantly with shortened survival of human breast cancer patients with metastatic disease and was highest in those with triple negative (TNBC) disease. Using a complex ex vivo all-human liver microphysiological (MPS) model of dormant-emergent metastatic progression, CXCR3 ligands were found to be elevated in actively growing populations of metastatic TNBC breast cancer cells whereas they remained similar to the tumor-free hepatic niche in those with dormant breast cancer cells. Subsequent stimulation of dormant breast cancer cells in the ex vivo metastatic liver MPS model with IP-10 triggered their emergence in a dose-dependent manner. Emergence was indicated to occur indirectly possibly via activation of the resident liver cells in the surrounding metastatic microenvironment, as stimulation of breast cancer cells with exogenous IP-10 did not significantly change their migratory, invasive or proliferative behavior. The findings reveal that IP-10 is capable of triggering the emergence of dormant breast cancer cells within the liver metastatic niche and identifies the IP-10/CXCR3 as a candidate targetable pathway for rational approaches aimed at maintaining dormancy. [ABSTRACT FROM AUTHOR]

Aortic aneurysm is a common cardiovascular disease characterised by continuous dilation of the aorta, and this disease places a heavy burden on healthcare worldwide. Few drugs have been suggested to be effective in controlling the progression of aortic aneurysms. Preclinical drug responses from traditional cell culture and animals are usually controversial. An effective in vitro model is of great demand for successful drug screening. In this study, we induced an in vitro microphysiological system to test metformin, which is a potential drug for the treatment of aortic aneurysms. Human pluripotent stem cell-derived aortic smooth muscle cells (hPSC-HASMCs) were cultured on an in vitro microphysiological system, which could replicate the cyclic stretch of the human native aortic wall. By using this system, we found that HASMCs were more likely to present a physiologically contractile phenotype compared to static cell cultures. Moreover, we used hPSC-HASMCs in our microphysiological system to perform metformin drug screening. The results showed that hPSC-HASMCs presented a more contractile phenotype via NOTCH 1 signalling while being treated with metformin. This result indicated that metformin could be utilised to rescue hPSC-HASMCs from phenotype switching during aortic aneurysm progression. This study helps to elucidate potential drug targets for the treatment of aortic aneurysms.

We guide the use of organ-on-chip technology in tissue engineering applications. Organ-on-chip technology is a form of microengineered cell culture platform that elaborates the in-vivo like organ or tissue microenvironments. The organ-on-chip platform consists of microfluidic channels, cell culture chambers, and stimulus sources that emulate the in-vivo microenvironment. These platforms are typically engraved into an oxygen-permeable transparent material. Fabrication of these materials requires the use of microfabrication strategies, including soft lithography, 3D printing, and injection molding. Here we provide an overview of what is an organ-on-chip platform, where it can be used, what it is composed of, how it can be fabricated, and how it can be operated. In connection with this topic, we also introduce an overview of the recent applications, where different organs are modeled on the microscale using this technology.

[Display omitted] Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global pandemic. Clinical evidence suggests that the intestine is another high-risk organ for SARS-CoV-2 infection besides the lungs. However, a model that can accurately reflect the response of the human intestine to the virus is still lacking. Here, we created an intestinal infection model on a chip that allows the recapitulation of human relevant intestinal pathophysiology induced by SARS-CoV-2 at organ level. This microengineered gut-on-chip reconstitutes the key features of the intestinal epithelium-vascular endothelium barrier through the three-dimensional (3D) co-culture of human intestinal epithelial, mucin-secreting, and vascular endothelial cells under physiological fluid flow. The intestinal epithelium showed permissiveness for viral infection and obvious morphological changes with injury of intestinal villi, dispersed distribution of mucus-secreting cells, and reduced expression of tight junction (E-cadherin), indicating the destruction of the intestinal barrier integrity caused by virus. Moreover, the vascular endothelium exhibited abnormal cell morphology, with disrupted adherent junctions. Transcriptional analysis revealed abnormal RNA and protein metabolism, as well as activated immune responses in both epithelial and endothelial cells after viral infection (e.g., upregulated cytokine genes), which may contribute to the injury of the intestinal barrier associated with gastrointestinal symptoms. This human organ system can partially mirror intestinal barrier injury and the human response to viral infection, which is not possible in existing in vitro culture models. It provides a unique and rapid platform to accelerate COVID-19 research and develop novel therapies. [ABSTRACT FROM AUTHOR]

Introduction: Interstitial hypertension, a rise in interstitial fluid pressure, is a common feature of many solid tumors as they progress to an invasive state. It is currently unclear whether this elevated pressure alters the probability that tumor cells eventually escape into a neighboring blood or lymphatic vessel. Methods: In this study, we analyze the escape of MDA-MB-231 human breast tumor cells from a ~3-mm-long preformed aggregate into a 120-μm-diameter empty cavity in a micromolded type I collagen gel. The "micro-tumors" were located within ~300 μm of one or two cavities. Pressures of ~0.65 cm H2O were applied only to the tumor ("interstitial hypertension") or to its adjacent cavity. Results: This work shows that interstitial hypertension suppresses escape into the adjacent cavity, but not because tumor cells respond directly to the pressure profile. Instead, hypertension alters the chemical microenvironment at the tumor margin to one that hampers escape. Administration of tumor interstitial fluid phenocopies the effects of hypertension. Conclusions: This work uncovers a link between tumor pressure, interstitial flow, and tumor cell escape in MDA-MB-231 cells, and suggests that interstitial hypertension serves to hinder further progression to metastatic escape. [ABSTRACT FROM AUTHOR]

The pharmaceutical industry has been desperately searching for efficient drug discovery methods. Organ-on-a-Chip, a cutting-edge technology that can emulate the physiological environment and functionality of human organs on a chip for disease modeling and drug testing, shows great potential for revolutionizing the drug development pipeline. However, successful translation of this novel engineering platform into routine pharmacological and medical scenarios remains to be realized. In this review, we discuss how the Organ-on-a-Chip technology can have critical roles in different preclinical stages of drug development and highlight the current challenges in translation and commercialization of this technology for the pharmacological and medical end-users. Moreover, this review sheds light on the future developmental trends and need for a next-generation Organ-on-a-Chip platform to bridge the gap between animal studies and clinical trials for the pharmaceutical industry. Organ-on-a-Chip is a promising interdisciplinary technique emulating in vivo physiology and pathology for in vitro disease modeling, drug screening, and precision medicine. The Organ-on-a-Chip technology can be organically incorporated into the drug development pipeline from early drug discovery to preclinical screening, testing, and translation of new drugs, which bridges the gap between animal studies and clinical trials involving human subjects. The future development of personalized Organ-on-a-Chip and continuous integration of novel engineering tools (e.g., automation handling, 3D printing. and in situ multisensors) and biological concepts (e.g., patient-specific induced pluripotent stem cells and organoids) into Organ-on-a-Chip platform will unprecedentedly promote its biomedical applications. [ABSTRACT FROM AUTHOR]

Tissues on a chip are sophisticated three-dimensional (3D) in vitro microphysiological systems designed to replicate human tissue conditions within dynamic physicochemical environments. However, the current fabrication methods for tissue spheroids on a chip require multiple parts and manual processing steps, including the deposition of spheroids onto prefabricated "chips." These challenges also lead to limitations regarding scalability and reproducibility. To overcome these challenges, we employed 3D printing techniques to automate the fabrication process of tissue spheroids on a chip. This allowed the simultaneous high-throughput printing of human liver spheroids and their surrounding polymeric flow chamber "chips" containing inner channels in a single step. The fabricated liver tissue spheroids on a liver-on-a-chip (LOC) were subsequently subjected to dynamic culturing by a peristaltic pump, enabling assessment of cell viability and metabolic activities. The 3D printed liver spheroids within the printed chips demonstrated high cell viability (>80%), increased spheroid size, and consistent adenosine triphosphate (ATP) activity and albumin production for up to 14 days. Furthermore, we conducted a study on the effects of acetaminophen (APAP), a nonsteroidal anti-inflammatory drug, on the LOC. Comparative analysis revealed a substantial decline in cell viability (<40%), diminished ATP activity, and reduced spheroid size after 7 days of culture within the APAP-treated LOC group, compared to the nontreated groups. These results underscore the potential of 3D bioprinted tissue chips as an advanced in vitro model that holds promise for accurately studying in vivo biological processes, including the assessment of tissue response to administered drugs, in a high-throughput manner.

Chimeric antigen receptor (CAR) cells are genetically engineered immune cells that specifically target tumor-associated antigens and have revolutionized cancer treatment, particularly in hematological malignancies, with ongoing investigations into their potential applications in solid tumors. This review provides a comprehensive overview of the current status and challenges in drug metabolism and pharmacokinetics (DMPK) for CAR cell therapy, specifically emphasizing on quantitative modeling and simulation (M&S). Furthermore, the recent advances in quantitative model analysis have been reviewed, ranging from clinical data characterization to mechanism-based modeling that connects in vitro and in vivo nonclinical and clinical study data. Additionally, the future perspectives and areas for improvement in CAR cell therapy translation have been reviewed. This includes using formulation quality considerations, characterization of appropriate animal models, refinement of in vitro models for bottom-up approaches, and enhancement of quantitative bioanalytical methodology. Addressing these challenges within a DMPK framework is pivotal in facilitating the translation of CAR cell therapy, ultimately enhancing the patients' lives through efficient CAR cell therapies., Competing Interests: Declaration of competing interest AG, YM, MN, SI, and SY are employees of Takeda Pharmaceutical Company Limited., (Copyright © 2024 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.)

Aim: Recently, a GuMI gut microphysiological system has been established to coculture oxygen-intolerant Faecalibacterium prausnitzii ( F. prausnitzii ) A2-165 with organoids-derived primary human colonic epithelium. This study aims to test if this GuMI system applies to different donors with different healthy states and uses metabolomics to reveal the role of gut microbes in modulating host- and diet-derived molecules in the gut lumen. Methods: Organoids-derived colonic monolayers were generated from an uninflamed region of diverticulitis, ulcerative colitis, and Crohn's disease patients and then integrated into the GuMI system to coculture with F. prausnitzii A2-165 for 2 to 4 days. Apical media was collected for metabolomic analysis. Targeted metabolomics was performed to profile 169 polar chemicals under three conditions: conventional static culture without bacteria, GuMI without bacteria, and GuMI with F. prausnitzii . The barrier function of monolayers was measured using transepithelial resistance. Results: GuMI successfully cocultured patient-derived monolayers and F. prausnitzii for up to 4 days, with active bacterial growth. Introducing flow and oxygen gradient significantly increases the barrier function, while exposure to F. prausnitzii slightly increases the barrier function. Targeted metabolomics screened 169 compounds and detected 76 metabolites, of which 70 significantly differed between at least two conditions. F. prausnitzii significantly modulates the levels of nucleosides, nucleobases, and amino acids on the apical side. Further analysis suggests that F. prausnitzii changes the mRNA level of 260 transcription factor genes in colonic epithelial cells. Conclusion: The GuMI physiomimetic system can maintain the coculture of F. prausnitzii and colonic epithelium from different donors. Together with metabolomics, we identified the modulation of F. prausnitzii in extracellular chemicals and colonic epithelial cell transcription in coculture with human colonic epithelium, which may reflect its function in gut lumen in vivo ., Competing Interests: All authors declared that there are no conflicts of interest., (© The Author(s) 2024.)

Spheroids and organoids have attracted significant attention as innovative models for disease modeling and drug screening. By employing diverse types of spheroids or organoids, it is feasible to establish microphysiological systems that enhance the precision of disease modeling and offer more dependable and comprehensive drug screening. High-throughput microphysiological systems that support optional, parallel testing of multiple drugs have promising applications in personalized medical treatment and drug research. However, establishing such a system is highly challenging and requires a multidisciplinary approach. This study introduces a dynamic Microphysiological System Chip Platform (MSCP) with multiple functional microstructures that encompass the mentioned advantages. We developed a high-throughput lung cancer spheroids model and an intestine-liver-heart-lung cancer microphysiological system for conducting parallel testing on four anti-lung cancer drugs, demonstrating the feasibility of the MSCP. This microphysiological system combines microscale and macroscale biomimetics to enable a comprehensive assessment of drug efficacy and side effects. Moreover, the microphysiological system enables evaluation of the real pharmacological effect of drug molecules reaching the target lesion after absorption by normal organs through fluid-based physiological communication. The MSCP could serves as a valuable platform for microphysiological system research, making significant contributions to disease modeling, drug development, and personalized medical treatment., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Fetal membrane(amniochorion), the innermost lining of the intrauterine cavity, surround the fetus and enclose amniotic fluid. Unlike unidirectional blood flow, amniotic fluid subtly rocks back and forth, and thus, the innermost amnion epithelial cells are continuously exposed to low levels of shear stress from fluid undulation. Here, we tested the impact of fluid motion on amnion epithelial cells (AECs) as a bearer of force impact and their potential vulnerability to cytopathologic changes that can destabilize fetal membrane functions. An amnion membrane (AM) organ-on-chip (OOC) was utilized to culture human fetal amnion membrane cells. The applied flow was modulated to perfuse culture media back and forth for 48 hours flow culture to mimic fluid motion. Static culture condition was used as a negative control, and oxidative stress (OS) condition was used as a positive control for pathophysiological changes. The impacts of fluidic motion were evaluated by measuring cell viability, cellular transition, and inflammation. Additionally, scanning electron microscopy (SEM) imaging was performed to observe microvilli formation. The results show that regardless of the applied flow rate, AECs and AMCs maintained their viability, morphology, innate meta-state, and low production of pro-inflammatory cytokines. E-cadherin expression and microvilli formation in the AECs were upregulated in a flow rate-dependent fashion; however, this did not impact cellular morphology or cellular transition or inflammation. OS treatment induced a mesenchymal morphology, significantly higher vimentin to CK-18 ratio, and pro-inflammatory cytokine production in AECs, whereas AMCs did not respond in any significant manner. Fluid motion and shear stress, if any, did not impact AEC cell function and did not cause inflammation. Thus, when using an amnion membrane OOC model, the inclusion of a flow culture environment is not necessary to mimic any in utero physiologic cellular conditions of fetal membrane-derived cells., Competing Interests: Conflict of interest The authors declare no conflict of interest.

Static three-dimensional (3D) cell culture has been demonstrated in ultralow attachment well plates, hanging droplet plates, and microtiter well plates with hydrogels or magnetic nanoparticles. Although it is simple, reproducible, and relatively inexpensive, thus potentially used for high-throughput screening, statically cultured 3D cells often suffer from a necrotic core due to limited nutrient and oxygen diffusion and waste removal and have a limited in vivo -like tissue structure. Here, we overcome these challenges by developing a pillar/perfusion plate platform and demonstrating high-throughput, dynamic 3D cell culture. Cell spheroids were loaded on the pillar plate with hydrogel by simple sandwiching and encapsulation and cultured dynamically in the perfusion plate on a digital rocker. Unlike traditional microfluidic devices, fast flow velocity was maintained within perfusion wells and the pillar plate was separated from the perfusion plate for cell-based assays. It was compatible with common lab equipment and allowed cell culture, testing, staining, and imaging in situ . The pillar/perfusion plate enhanced cell growth by rapid diffusion, reproducibility, assay throughput, and user friendliness in a dynamic 3D cell culture.

Microphysiological systems, also known as organs-on-chips, are microfluidic devices designed to model human physiology in vitro. Polydimethylsiloxane (PDMS) is the most widely used material for organs-on-chips due to established microfabrication methods, and properties that make it suitable for biological applications such as low cytotoxicity, optical transparency, gas permeability. However, absorption of small molecules and leaching of uncrosslinked oligomers might hinder the adoption of PDMS-based organs-on-chips for drug discovery assays. Here, we have engineered a modular, PDMS-free microphysiological system that is capable of recapitulating biologic barrier functions commonly demonstrated in PDMS-based devices. Our microphysiological system is comprised of a microfluidic chip to house cell cultures and pneumatic microfluidic pumps to drive flow with programmable pressure and shear stress. The modular architecture and programmable pumps enabled us to model multiple in vivo microenvironments. First, we demonstrate the ability to generate cyclic strain on the culture membrane and establish a model of the alveolar air-liquid interface. Next, we utilized three-dimensional finite element analysis modeling to characterize the fluid dynamics within the device and develop a model of the pressure-driven filtration that occurs at the glomerular filtration barrier. Finally, we demonstrate that our model can be used to recapitulate sphingolipid induced kidney injury. Together, our results demonstrate that a multifunctional and modular microphysiological system can be deployed without the use of PDMS. Further, the bio-inert plastic used in our microfluidic device is amenable to various established, high-throughput manufacturing techniques, such as injection molding. As a result, the development plastic organs-on-chips provides an avenue to meet the increasing demand for organ-on-chip technology.

Organoids are adept at preserving the inherent complexity of a given cellular environment and when integrated with engineered micro-physiological systems (MPS) present distinct advantages for simulating a precisely controlled geometrical, physical, and biochemical micro-environment. This then allows for real-time monitoring of cell-cell interactions. As a result, the two aforementioned technologies hold significant promise and potential in studying ocular physiology and diseases by replicating specific eye tissue microstructures in vitro. This miniaturized review begins with defining the science behind organoids/MPS and subsequently introducing methods for generating organoids and engineering MPS. Furthermore, we will discuss the current state of organoids and MPS models in retina, cornea surrogates, and other ocular tissue, in regards to physiological/disease conditions. Finally, future prospective on organoid/MPS will be covered here. Organoids and MPS technologies closely recapture the in vivo microenvironment and thusly will continue to provide new understandings in organ functions and novel approaches to drug development.

While conventional in vitro culture systems and animal models have been used to study the pathogenesis of viral infections and to facilitate development of vaccines and therapeutics for viral diseases, models that can accurately recapitulate human responses to infection are still lacking. Human organ-on-a-chip (Organ Chip) microfluidic culture devices that recapitulate tissue–tissue interfaces, fluid flows, mechanical cues, and organ-level physiology have been developed to narrow the gap between in vitro experimental models and human pathophysiology. Here, we describe how recent developments in Organ Chips have enabled re-creation of complex pathophysiological features of human viral infections in vitro. Microfluidic Organ Chip culture devices are emerging alternatives to conventional in vitro and animal models due to their ability to replicate many structural and functional features of human physiology and disease states. Recent innovations demonstrate that Organ Chip technology is a promising strategy for virology studies where there have been successes in reproducing various viral disease phenotypes. Organ Chips have enabled investigation of many aspects of viral infection, including virus–host interactions, viral therapy-resistance evolution, and development of new antiviral therapeutics, as well as underlying pathogenesis. As Organ Chip-based assays provide accessibility to study virus-induced diseases in real time and at high resolution, they can open new avenues to uncover viral pathogenesis in a human-relevant environment and may eventually enable development of novel therapeutics and vaccines. [ABSTRACT FROM AUTHOR]

High-throughput in vitro models lack human-relevant complexity, which undermines their ability to accurately mimic in vivo biologic and pathologic responses. The emergence of microphysiological systems (MPS) presents an opportunity to revolutionize in vitro modeling for both basic biomedical research and applied drug discovery. The MPS platform has been an area of interdisciplinary collaboration to develop new, predictive, and reliable in vitro methods for regulatory acceptance. The current MPS models have been developed to recapitulate an organ or tissue on a smaller scale. However, the complexity of these models (ie, including all cell types present in the in vivo tissue) with appropriate structural, functional, and biochemical attributes are often not fully characterized. Here, we provide an overview of the capabilities and limitations of the microfluidic MPS model (aka organs-on-chips) within the scope of drug development. We recommend the engagement of pathologists early in the MPS design, characterization, and validation phases, because this will enable development of more robust and comprehensive MPS models that can accurately replicate normal biology and pathophysiology and hence be more predictive of human responses. [ABSTRACT FROM AUTHOR]

Organoids are adept at preserving the inherent complexity of a given cellular environment and when integrated with engineered micro-physiological systems (MPS) present distinct advantages for simulating a precisely controlled geometrical, physical, and biochemical micro-environment. This then allows for real-time monitoring of cell-cell interactions. As a result, the two aforementioned technologies hold significant promise and potential in studying ocular physiology and diseases by replicating specific eye tissue microstructures in vitro. This miniaturized review begins with defining the science behind organoids/MPS and subsequently introducing methods for generating organoids and engineering MPS. Furthermore, we will discuss the current state of organoids and MPS models in retina, cornea surrogates, and other ocular tissue, in regards to physiological/disease conditions. Finally, future prospective on organoid/MPS will be covered here. Organoids and MPS technologies closely recapture the in vivo microenvironment and thusly will continue to provide new understandings in organ functions and novel approaches to drug development. [ABSTRACT FROM AUTHOR]

Abstract Background Our understanding of the multiple roles exosomes play during tumor progression is still very poor and the contribution of the normal tissue derived exosomes in distant seeding and tumor outgrowth has also not been widely appreciated. Methods Using our all-human liver microphysiological system (MPS) platform as a model to closely recapitulate the early metastatic events, we isolated exosomes from both tumor cells and liver microenvironment. Results We observed that while priming of the hepatic niche (HepN) with MDA-231 breast cancer derived exosomes facilitated seeding of the cancer cells in the liver, subsequent tumor outgrowth was diminished; this was consistent with increased entry into dormancy. We found that hepatic niche (HepN) derived exosomes contribute significantly to the exosome pool and are distinguished from cancer derived exosomes based on their size, protein and miRNA content. By Ingenuity Pathway Analysis (IPA) of the miRNA content of the HepN, MDA-231/HepN and MDA-231 cells we showed that the HepN derived exosomes affect the breast cancer cells by suppressing pathways involved in cancer cell proliferation and invasion. More importantly exposure of MDA-231 and MDA-468 cells to purified normal HepN derived exosomes, induced changes in the cells consistent with a Mesenchymal to Epithelial reverting Transition (MErT). miRNA arrays performed on MDA-231 treated with Hum Hep/NPC derived exosomes showed significant changes in the levels of a select number of miRNAs involved in epithelial cell differentiation and miRNAs, such as miR186, miR23a and miR205, from our top and bottom bins have previously been reported to regulate E-cadherin transcription and MErT induction in various cancer types. Consistently HepN derived exosome treatment of breast and prostate cancer lines lead to a transient induction of E-cadherin and ZO-1 at the protein level and a more epithelial-like morphology of the cells. Conclusions Collectively our data revealed a novel mechanism of regulation of the metastatic cascade, showing a well-orchestrated, timely controlled crosstalk between the cancer cells and the HepN and implicating for the first time the normal tissue/HepN derived exosomes in enabling seeding and entry into dormancy of the cancer cells at the metastatic site.

Microphysiological systems play a pivotal role in progressing toward a global paradigm shift in drug development. Here, we designed a four-organ-chip interconnecting miniaturized human intestine, liver, brain and kidney equivalents. All four organ models were predifferentiated from induced pluripotent stem cells from the same healthy donor and integrated into the microphysiological system. The coculture of the four autologous tissue models in one common medium deprived of tissue specific growth factors was successful over 14-days. Although there were no added growth factors present in the coculture medium, the intestine, liver and neuronal model maintained defined marker expression. Only the renal model was overgrown by coexisting cells and did not further differentiate. This model platform will pave the way for autologous coculture cross-talk assays, disease induction and subsequent drug testing.

Organotypic and microphysiological systems (MPS) that can emulate the molecular phenotype and function of human tissues, such as liver, are increasingly used in preclinical drug development. However, despite their improved predictivity, drug development success rates have remained low with most compounds failing in clinical phases despite promising preclinical data. Here, it is tested whether absorption of small molecules to polymers commonly used for MPS fabrication can impact preclinical pharmacological and toxicological assessments and contribute to the high clinical failure rates. To this end, identical devices are fabricated from eight different MPS polymers and absorption of prototypic compounds with different physicochemical properties are analyzed. It is found that overall absorption is primarily driven by compound hydrophobicity and the number of rotatable bonds. However, absorption can differ by >1000-fold between polymers with polydimethyl siloxane (PDMS) being most absorptive, whereas polytetrafluoroethylene (PTFE) and thiol-ene epoxy (TEE) absorbed the least. Strikingly, organotypic primary human liver cultures successfully flagged hydrophobic hepatotoxins in lowly absorbing TEE devices at therapeutically relevant concentrations, whereas isogenic cultures in PDMS devices are resistant, resulting in false negative safety signals. Combined, these results can guide the selection of MPS materials and facilitate the development of preclinical assays with improved translatability., (© 2023 The Authors. Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

Patient-derived microphysiological systems (P-MPS) have emerged as powerful tools in precision medicine that provide valuable insight into individual patient characteristics. This review discusses the development of P-MPS as an integration of patient-derived samples, including patient-derived cells, organoids, and induced pluripotent stem cells, into well-defined MPSs. Emphasizing the necessity of P-MPS development, its significance as a nonclinical assessment approach that bridges the gap between traditional in vitro models and clinical outcomes is highlighted. Additionally, guidance is provided for engineering approaches to develop microfluidic devices and high-content analysis for P-MPSs, enabling high biological relevance and high-throughput experimentation. The practical implications of the P-MPS are further examined by exploring the clinically relevant outcomes obtained from various types of patient-derived samples. The construction and analysis of these diverse samples within the P-MPS have resulted in physiologically relevant data, paving the way for the development of personalized treatment strategies. This study describes the significance of the P-MPS in precision medicine, as well as its unique capacity to offer valuable insights into individual patient characteristics., (© 2023 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.)

Despite recent advances in the field of microphysiological systems (MPSs), availability of models capable of mimicking the interactions between the nervous system and innervated tissues is still limited. This represents a significant challenge in identifying the underlying processes of various pathological conditions, including neuropathic, cardiovascular and metabolic disorders. In this novel study, we introduce a compartmentalized three-dimensional (3D) coculture system that enables physiologically relevant tissue innervation while recording neuronal excitability. By integrating custom microelectrode arrays into tailored glass chips microfabricated via selective laser-etching, we developed an entirely novel class of innervation MPSs (INV-MPS). This INV-MPS allows for manipulation, visualization, and electrophysiological analysis of individual axons innervating complex 3D tissues. Here, we focused on sensory innervation of 3D tumor tissue as a model case study since cancer-induced pain represents a major unmet medical need. The system was compared with existing nociception models and successfully replicated axonal chemoattraction mediated by nerve growth factor (NGF). Remarkably, in the absence of NGF, 3D cancer spheroids cocultured in the adjacent compartment induced sensory neurons to consistently cross the separating barrier and establish fine innervation. Moreover, we observed that crossing sensory fibers could be chemically excited by distal application of known pain-inducing agonists only when cocultured with cancer cells. To our knowledge, this is the first system showcasing morphological and electrophysiological analysis of 3D-innervated tumor tissue in vitro , paving the way for a plethora of studies into innervation-related diseases and improving our understanding of underlying pathophysiology., (Creative Commons Attribution license.)

Human adipose-derived stromal/stem cells (hASC) are widely used for in vitro modeling of physiologically relevant human adipose tissue. These models are useful for the development of tissue constructs for soft tissue regeneration and 3-dimensional (3D) microphysiological systems (MPS) for drug discovery. In this systematic review, we report on the current state of hASC culture and assessment methods for adipose tissue engineering using 3D MPS. Our search efforts resulted in the identification of 184 independent records, of which 27 were determined to be most relevant to the goals of the present review. Our results demonstrate a lack of consensus on methods for hASC culture and assessment for the production of physiologically relevant in vitro models of human adipose tissue. Few studies have assessed the impact of different 3D culture conditions on hASC adipogenesis. Additionally, there has been a limited use of assays for characterizing the functionality of adipose tissue in vitro. Results from this study suggest the need for more standardized culture methods and further analysis on in vitro tissue functionality. These will be necessary to validate the utility of 3D MPS as an in vitro model to reduce, refine, and replace in vivo experiments in the drug discovery regulatory process.

The blood-retinal barrier (BRB) protects the retina by maintaining an adequate microenvironment for neuronal function. Alterations of the junctional complex of the BRB and consequent BRB breakdown in disease contribute to a loss of neuronal signaling and vision loss. As new therapeutics are being developed to prevent or restore barrier function, it is critical to implement physiologically relevant in vitro models that recapitulate the important features of barrier biology to improve disease modeling, target validation, and toxicity assessment. New directions in organ-on-a-chip technology are enabling more sophisticated 3-dimensional models with flow, multicellularity, and control over microenvironmental properties. By capturing additional biological complexity, organs-on-chip can help approach actual tissue organization and function and offer additional tools to model and study disease compared with traditional 2-dimensional cell culture. This review describes the current state of barrier biology and barrier function in ocular diseases, describes recent advances in organ-on-a-chip design for modeling the BRB, and discusses the potential of such models for ophthalmic drug discovery and development. [ABSTRACT FROM AUTHOR]