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With current plans for manned missions to Mars and beyond, the need to better understand, prevent, and counteract the harmful effects of long-duration spaceflight on the body is becoming increasingly important. In this study, an automated heart-on-a-chip platform was flown to the International Space Station on a 1-mo mission during which contractile cardiac function was monitored in real-time. Upon return to Earth, engineered human heart tissues (EHTs) were further analyzed with ultrastructural imaging and RNA sequencing to investigate the impact of prolonged microgravity on cardiomyocyte function and health. Spaceflight EHTs exhibited significantly reduced twitch forces, increased incidences of arrhythmias, and increased signs of sarcomere disruption and mitochondrial damage. Transcriptomic analyses showed an up-regulation of genes and pathways associated with metabolic disorders, heart failure, oxidative stress, and inflammation, while genes related to contractility and calcium signaling showed significant down-regulation. Finally, in silico modeling revealed a potential link between oxidative stress and mitochondrial dysfunction that corresponded with RNA sequencing results. This represents an in vitro model to faithfully reproduce the adverse effects of spaceflight on three-dimensional (3D)-engineered heart tissue. [ABSTRACT FROM AUTHOR]

A major challenge in myocardial tissue engineering is replicating the heart's highly complex three-dimensional (3D) anisotropic structure. Heart-on-a-chip (HOC) is an emerging technology for constructing myocardial tissue in vitro in recent years, but most existing HOC systems face difficulties in constructing 3D myocardial tissue aligned with multiple cell layers. Electrospun nanofibers are commonly used as scaffolds for cell growth in myocardial tissue engineering, which can structurally simulate the extracellular matrix to induce the aligned growth of myocardial cells. Here, we developed an HOC that integrates multi-layered aligned polycaprolactone (PCL) nanofiber scaffolds inside microfluidic chips, and constructed 3D thick and aligned tissue with a layered seeding approach. By culturing human-induced pluripotent stem-cell-derived cardiomyocytes (hiPSC-CMs) on chip, the myocardial tissue on the two layered nanofibers reached a thickness of ~53 μm compared with ~19 μm for single-layered nanofibers. The obtained myocardial tissue presented well-aligned structures, with densely distributed α-actinin. By the third day post seeding, the hiPSC-CMs contract highly synchronously, with a contraction frequency of 18 times/min. The HOC with multi-layered biomimetic scaffolds provided a dynamic in vitro culture environment for hiPSC-CMs. Together with the layered cell-seeding process, the designed HOC promoted the formation of thick, well-aligned myocardial tissue. [ABSTRACT FROM AUTHOR]

One of the many unresolved obstacles in the field of cardiovascular research is an uncompromising in vitro cardiac model. While primary cell sources from animal models offer both advantages and disadvantages, efforts over the past half-century have aimed to reduce their use. Additionally, obtaining a sufficient quantity of human primary cardiomyocytes faces ethical and legal challenges. As the practically unlimited source of human cardiomyocytes from induced pluripotent stem cells (hiPSC-CM) is now mostly resolved, there are great efforts to improve their quality and applicability by overcoming their intrinsic limitations. The greatest bottleneck in the field is the in vitro ageing of hiPSC-CMs to reach a maturity status that closely resembles that of the adult heart, thereby allowing for more appropriate drug developmental procedures as there is a clear correlation between ageing and developing cardiovascular diseases. Here, we review the current state-of-the-art techniques in the most realistic heart models used in disease modelling and toxicity evaluations from hiPSC-CM maturation through heart-on-a-chip platforms and in silico models to the in vitro models of certain cardiovascular diseases. [ABSTRACT FROM AUTHOR]

Abstract In recent years, research on organ-on-a-chip technology has been flourishing, particularly for drug screening and disease model development. Fibroblasts and vascular endothelial cells engage in crosstalk through paracrine signaling and direct cell–cell contact, which is essential for the normal development and function of the heart. Therefore, to faithfully recapitulate cardiac function, it is imperative to incorporate fibroblasts and vascular endothelial cells into a heart-on-a-chip model. Here, we report the development of a human heart-on-a-chip composed of induced pluripotent stem cell (iPSC)-derived cardiomyocytes, fibroblasts, and vascular endothelial cells. Vascular endothelial cells cultured on microfluidic channels responded to the flow of culture medium mimicking blood flow by orienting themselves parallel to the flow direction, akin to in vivo vascular alignment in response to blood flow. Furthermore, the flow of culture medium promoted integrity among vascular endothelial cells, as evidenced by CD31 staining and lower apparent permeability. The tri-culture condition of iPSC-derived cardiomyocytes, fibroblasts, and vascular endothelial cells resulted in higher expression of the ventricular cardiomyocyte marker IRX4 and increased contractility compared to the bi-culture condition with iPSC-derived cardiomyocytes and fibroblasts alone. Such tri-culture-derived cardiac tissues exhibited cardiac responses similar to in vivo hearts, including an increase in heart rate upon noradrenaline administration. In summary, we have achieved the development of a heart-on-a-chip composed of cardiomyocytes, fibroblasts, and vascular endothelial cells that mimics in vivo cardiac behavior.

In recent years, research on organ-on-a-chip technology has been flourishing, particularly for drug screening and disease model development. Fibroblasts and vascular endothelial cells engage in crosstalk through paracrine signaling and direct cell–cell contact, which is essential for the normal development and function of the heart. Therefore, to faithfully recapitulate cardiac function, it is imperative to incorporate fibroblasts and vascular endothelial cells into a heart-on-a-chip model. Here, we report the development of a human heart-on-a-chip composed of induced pluripotent stem cell (iPSC)-derived cardiomyocytes, fibroblasts, and vascular endothelial cells. Vascular endothelial cells cultured on microfluidic channels responded to the flow of culture medium mimicking blood flow by orienting themselves parallel to the flow direction, akin to in vivo vascular alignment in response to blood flow. Furthermore, the flow of culture medium promoted integrity among vascular endothelial cells, as evidenced by CD31 staining and lower apparent permeability. The tri-culture condition of iPSC-derived cardiomyocytes, fibroblasts, and vascular endothelial cells resulted in higher expression of the ventricular cardiomyocyte marker IRX4 and increased contractility compared to the bi-culture condition with iPSC-derived cardiomyocytes and fibroblasts alone. Such tri-culture-derived cardiac tissues exhibited cardiac responses similar to in vivo hearts, including an increase in heart rate upon noradrenaline administration. In summary, we have achieved the development of a heart-on-a-chip composed of cardiomyocytes, fibroblasts, and vascular endothelial cells that mimics in vivo cardiac behavior. [ABSTRACT FROM AUTHOR]

Recent advances in both cardiac tissue engineering and hearts‐on‐a‐chip are grounded in new biomaterial development as well as the employment of innovative fabrication techniques that enable precise control of the mechanical, electrical, and structural properties of the cardiac tissues being modelled. The elongated structure of cardiomyocytes requires tuning of substrate properties and application of biophysical stimuli to drive its mature phenotype. Landmark advances have already been achieved with induced pluripotent stem cell‐derived cardiac patches that advanced to human testing. Heart‐on‐a‐chip platforms are now commonly used by a number of pharmaceutical and biotechnology companies. Here, we provide an overview of cardiac physiology in order to better define the requirements for functional tissue recapitulation. We then discuss the biomaterials most commonly used in both cardiac tissue engineering and heart‐on‐a‐chip, followed by the discussion of recent representative studies in both fields. We outline significant challenges common to both fields, specifically: scalable tissue fabrication and platform standardization, improving cellular fidelity through effective tissue vascularization, achieving adult tissue maturation, and ultimately developing cryopreservation protocols so that the tissues are available off the shelf. [ABSTRACT FROM AUTHOR]

The implementation of three-dimensional tissue engineering concurrently with stem cell technology holds great promise for in vitro research in pharmacology and toxicology and modeling cardiac diseases, particularly for rare genetic and pediatric diseases for which animal models, immortal cell lines, and biopsy samples are unavailable. It also allows for a rapid assessment of phenotype–genotype relationships and tissue response to pharmacological manipulation. Mutations in the TSC1 and TSC2 genes lead to dysfunctional mTOR signaling and cause tuberous sclerosis complex (TSC), a genetic disorder that affects multiple organ systems, principally the brain, heart, skin, and kidneys. Here we differentiated healthy (CC3) and tuberous sclerosis (TSP8-15) human induced pluripotent stem cells (hiPSCs) into cardiomyocytes to create engineered cardiac tissue constructs (ECTCs). We investigated and compared their mechano-elastic properties and gene expression and assessed the effects of rapamycin, a potent inhibitor of the mechanistic target of rapamycin (mTOR). The TSP8-15 ECTCs had increased chronotropy compared to healthy ECTCs. Rapamycin induced positive inotropic and chronotropic effects (i.e., increased contractility and beating frequency, respectively) in the CC3 ECTCs but did not cause significant changes in the TSP8-15 ECTCs. A differential gene expression analysis revealed 926 up- and 439 down-regulated genes in the TSP8-15 ECTCs compared to their healthy counterparts. The application of rapamycin initiated the differential expression of 101 and 31 genes in the CC3 and TSP8-15 ECTCs, respectively. A gene ontology analysis showed that in the CC3 ECTCs, the positive inotropic and chronotropic effects of rapamycin correlated with positively regulated biological processes, which were primarily related to the metabolism of lipids and fatty and amino acids, and with negatively regulated processes, which were predominantly associated with cell proliferation and muscle and tissue development. In conclusion, this study describes for the first time an in vitro TSC cardiac tissue model, illustrates the response of normal and TSC ECTCs to rapamycin, and provides new insights into the mechanisms of TSC. [ABSTRACT FROM AUTHOR]

Recent advances in heart-on-a-chip systems hold great promise to facilitate cardiac physiological, pathological, and pharmacological studies. This review focuses on the development of heart-on-a-chip systems with tissue-specific functionalities. For one thing, the strategies for developing cardiac microtissues on heart-on-a-chip systems that closely mimic the structures and behaviors of the native heart are analyzed, including the imitation of cardiac structural and functional characteristics. For another, the development of techniques for real-time monitoring of biophysical and biochemical signals from cardiac microtissues on heart-on-a-chip systems is introduced, incorporating cardiac electrophysiological signals, contractile activity, and biomarkers. Furthermore, the applications of heart-on-a-chip systems in intelligent cardiac studies are discussed regarding physiological/pathological research and pharmacological assessment. Finally, the future development of heart-on-a-chip toward a higher level of systematization, integration, and maturation is proposed.

Animal models and traditional cell cultures are essential tools for drug development. However, these platforms can show striking discrepancies in efficacy and side effects when compared to human trials. These differences can lengthen the drug development process and even lead to drug withdrawal from the market. The establishment of preclinical drug screening platforms that have higher relevancy to physiological conditions is desirable to facilitate drug development. Here, a heart-on-a-chip platform, incorporating microgrooves and electrical pulse stimulations to recapitulate the well-aligned structure and synchronous beating of cardiomyocytes (CMs) for drug screening, is reported. Each chip is made with facile lithographic and laser-cutting processes that can be easily scaled up to high-throughput format. The maturation and phenotypic changes of CMs cultured on the heart-on-a-chip is validated and it can be treated with various drugs to evaluate cardiotoxicity and cardioprotective efficacy. The heart-on-a-chip can provide a high-throughput drug screening platform in preclinical drug development.

[Display omitted] Heart-on-chips have emerged as a powerful tool to promote the paradigm innovation in cardiac pathological research and drug development. Attempts are focused on improving microphysiological visuals, enhancing bionic characteristics, as well as expanding their biomedical applications. Herein, inspired by the bright feathers of peacock, we present a novel optical and electrical dual-responsive heart-on-a-chip based on cardiomyocytes hybrid bright MXene structural color hydrogels for hormone toxicity evaluation. Such hydrogels with inverse opal nanostructure are generated by using pregel to replicate MXene-decorated colloidal photonic crystal (PhC) array templates. The attendant MXene in the hydrogels could not only enhance the saturation of structural color, but also ensure the composite hydrogel with excellent electroconductivity to facilitate the synergetic beating of their surface cultured cardiomyocytes. In this case, the hydrogels would undergo a synchronous deformation and generate shift in corresponding photonic band gap and structural color, which could be employed as visual signal for self-reporting of the cardiomyocyte mechanics. Based on these features, we demonstrated the practical value of the optical and electrical dual-responsive structural color MXene hydrogels constructed heart-on-a-chip in hormone toxicity testing. These results indicated that the proposed heart-on-a-chip might find broad prospects in drug screening, biological research, and so on. [ABSTRACT FROM AUTHOR]

The implementation of three-dimensional tissue engineering concurrently with stem cell technology holds great promise for in vitro research in pharmacology and toxicology and modeling cardiac diseases, particularly for rare genetic and pediatric diseases for which animal models, immortal cell lines, and biopsy samples are unavailable. It also allows for a rapid assessment of phenotype–genotype relationships and tissue response to pharmacological manipulation. Mutations in the TSC1 and TSC2 genes lead to dysfunctional mTOR signaling and cause tuberous sclerosis complex (TSC), a genetic disorder that affects multiple organ systems, principally the brain, heart, skin, and kidneys. Here we differentiated healthy (CC3) and tuberous sclerosis (TSP8-15) human induced pluripotent stem cells (hiPSCs) into cardiomyocytes to create engineered cardiac tissue constructs (ECTCs). We investigated and compared their mechano-elastic properties and gene expression and assessed the effects of rapamycin, a potent inhibitor of the mechanistic target of rapamycin (mTOR). The TSP8-15 ECTCs had increased chronotropy compared to healthy ECTCs. Rapamycin induced positive inotropic and chronotropic effects (i.e., increased contractility and beating frequency, respectively) in the CC3 ECTCs but did not cause significant changes in the TSP8-15 ECTCs. A differential gene expression analysis revealed 926 up- and 439 down-regulated genes in the TSP8-15 ECTCs compared to their healthy counterparts. The application of rapamycin initiated the differential expression of 101 and 31 genes in the CC3 and TSP8-15 ECTCs, respectively. A gene ontology analysis showed that in the CC3 ECTCs, the positive inotropic and chronotropic effects of rapamycin correlated with positively regulated biological processes, which were primarily related to the metabolism of lipids and fatty and amino acids, and with negatively regulated processes, which were predominantly associated with cell proliferation and muscle and tissue development. In conclusion, this study describes for the first time an in vitro TSC cardiac tissue model, illustrates the response of normal and TSC ECTCs to rapamycin, and provides new insights into the mechanisms of TSC.

The study of human cardiomyopathies and the development and testing of new therapies has long been limited by the availability of appropriate in vitro model systems. Cardiomyocytes are highly specialized cells whose internal structure and contractile function are sensitive to the local microenvironment and the combination of mechanical and biochemical cues they receive. The complementary technologies of human induced pluripotent stem cell (hiPSC) derived cardiomyocytes (CMs) and microphysiological systems (MPS) allow for precise control of the genetics and microenvironment of human cells in in vitro contexts. These combined systems also enable quantitative measurement of mechanical function and intracellular organization. This review describes relevant factors in the myocardium microenvironment that affect CM structure and mechanical function and demonstrates the application of several engineered microphysiological systems for studying development, disease, and drug discovery.

The fabrication of complex and stable vasculature in engineered cardiac tissues represents a significant hurdle towards building physiologically relevant models of the heart. Here, we implemented a 3D model of cardiac vasculogenesis, incorporating endothelial cells (EC), stromal cells, and human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CM) in a fibrin hydrogel. The presence of CMs disrupted vessel formation in 3D tissues, resulting in the upregulation of endothelial activation markers and altered extracellular vesicle (EV) signaling in engineered tissues as determined by the proteomic analysis of culture supernatant. miRNA sequencing of CM- and EC-secreted EVs highlighted key EV-miRNAs that were postulated to play differing roles in cardiac vasculogenesis, including the let-7 family and miR-126-3p in EC-EVs. In the absence of CMs, the supplementation of CM-EVs to EC monolayers attenuated EC migration and proliferation and resulted in shorter and more discontinuous self-assembling vessels when applied to 3D vascular tissues. In contrast, supplementation of EC-EVs to the tissue culture media of 3D vascularized cardiac tissues mitigated some of the deleterious effects of CMs on vascular self-assembly, enhancing the average length and continuity of vessel tubes that formed in the presence of CMs. Direct transfection validated the effects of the key EC-EV miRNAs let-7b-5p and miR-126-3p in improving the maintenance of continuous vascular networks. EC-EV supplementation to biofabricated cardiac tissues and microfluidic devices resulted in tissue vascularization, illustrating the use of this approach in the engineering of enhanced, perfusable, microfluidic models of the myocardium., (Creative Commons Attribution license.)

Heart‐on‐a‐chip plays an important role in revealing the biological mechanism and developing new drugs for cardiomyopathy. Tremendous efforts have been devoted to developing heart‐on‐a‐chip systems featuring simplified fabrication, accurate imitation and microphysiological visuality. In this paper, the authors present a novel electroconductive and anisotropic structural color hydrogel by simply polymerizing non‐close‐packed colloidal arrays on super aligned carbon nanotube sheets (SACNTs) for visualized and accurate heart‐on‐a‐chip construction. The generated anisotropic hydrogel consists of a colloidal array‐locked hydrogel layer with brilliant structural color on one surface and a conductive methacrylated gelatin (GelMA)/SACNTs film on the other surface. It is demonstrated that the anisotropic morphology of the SACNTs could effectively induce the alignment of cardiomyocytes, and the conductivity of SACNTs could contribute to the synchronous beating of cardiomyocytes. Such consistent beating rhythm caused the deformation of the hydrogel substrates and dynamic shifts in structural color and reflection spectra of the whole hybrid hydrogels. More attractively, with the integration of such cardiomyocyte‐driven living structural color hydrogels and microfluidics, a visualized heart‐on‐a‐chip system with more consistent beating frequency has been established for dynamic cardiomyocyte sensing and drug screening. The results indicate that the electroconductive and anisotropic structural color hydrogels are potential for various biomedical applications. [ABSTRACT FROM AUTHOR]

In vitro studies of cardiac physiology and drug response have traditionally been performed on individual isolated cardiomyocytes or isotropic monolayers of cells that may not mimic desired physiological traits of the laminar adult myocardium. Recent studies have reported a number of advances to Heart-on-a-Chip platforms for the fabrication of more sophisticated engineered myocardium, but cardiomyocyte immaturity remains a challenge. In the anisotropic musculature of the heart, interactions between cardiac myocytes, the extracellular matrix (ECM), and neighboring cells give rise to changes in cell shape and tissue architecture that have been implicated in both development and disease. We hypothesized that engineered myocardium fabricated from cardiac myocytes cultured in vitro could mimic the physiological characteristics and gene expression profile of adult heart muscle. To test this hypothesis, we fabricated engineered myocardium comprised of neonatal rat ventricular myocytes with laminar architectures reminiscent of that observed in the mature heart and compared their sarcomere organization, contractile performance characteristics, and cardiac gene expression profile to that of isolated adult rat ventricular muscle strips. We found that anisotropic engineered myocardium demonstrated a similar degree of global sarcomere alignment, contractile stress output, and inotropic concentration-response to the β-adrenergic agonist isoproterenol. Moreover, the anisotropic engineered myocardium exhibited comparable myofibril related gene expression to muscle strips isolated from adult rat ventricular tissue. These results suggest that tissue architecture serves an important developmental cue for building in vitro model systems of the myocardium that could potentially recapitulate the physiological characteristics of the adult heart. Impact statement With the recent focus on developing in vitro Organ-on-Chip platforms that recapitulate tissue and organ-level physiology using immature cells derived from stem cell sources, there is a strong need to assess the ability of these engineered tissues to adopt a mature phenotype. In the present study, we compared and contrasted engineered tissues fabricated from neonatal rat ventricular myocytes in a Heart-on-a-Chip platform to ventricular muscle strips isolated from adult rats. The results of this study support the notion that engineered tissues fabricated from immature cells have the potential to mimic mature tissues in an Organ-on-Chip platform.

Human induced pluripotent stem cells (hiPSCs) can be used to generate various cell types in the human body. Hence, hiPSC-derived cardiomyocytes (hiPSC-CMs) represent a significant cell source for disease modeling, drug testing, and regenerative medicine. The immaturity of hiPSC-CMs in two-dimensional (2D) culture limit their applications. Cardiac tissue engineering provides a new promise for both basic and clinical research. Advanced bioengineered cardiac in vitro models can create contractile structures that serve as exquisite in vitro heart microtissues for drug testing and disease modeling, thereby promoting the identification of better treatments for cardiovascular disorders. In this review, we will introduce recent advances of bioengineering technologies to produce in vitro cardiac tissues derived from hiPSCs. [ABSTRACT FROM AUTHOR]

Abstract Heart‐on‐a‐chip plays an important role in revealing the biological mechanism and developing new drugs for cardiomyopathy. Tremendous efforts have been devoted to developing heart‐on‐a‐chip systems featuring simplified fabrication, accurate imitation and microphysiological visuality. In this paper, the authors present a novel electroconductive and anisotropic structural color hydrogel by simply polymerizing non‐close‐packed colloidal arrays on super aligned carbon nanotube sheets (SACNTs) for visualized and accurate heart‐on‐a‐chip construction. The generated anisotropic hydrogel consists of a colloidal array‐locked hydrogel layer with brilliant structural color on one surface and a conductive methacrylated gelatin (GelMA)/SACNTs film on the other surface. It is demonstrated that the anisotropic morphology of the SACNTs could effectively induce the alignment of cardiomyocytes, and the conductivity of SACNTs could contribute to the synchronous beating of cardiomyocytes. Such consistent beating rhythm caused the deformation of the hydrogel substrates and dynamic shifts in structural color and reflection spectra of the whole hybrid hydrogels. More attractively, with the integration of such cardiomyocyte‐driven living structural color hydrogels and microfluidics, a visualized heart‐on‐a‐chip system with more consistent beating frequency has been established for dynamic cardiomyocyte sensing and drug screening. The results indicate that the electroconductive and anisotropic structural color hydrogels are potential for various biomedical applications.

[Display omitted] Cellular mechanics plays an important role in physiological processes such as cell growth, differentiation, apoptosis and gene expression. Herein, we present a nano-imprinted structural color graphene film with anisotropic microgroove and two-dimensional (2D) photonic crystal structures for cardiomyocytes dynamic displaying. The anisotropic structural color graphene film was generated by a "sandwich" mold in one step with the assistance of two designed templates. Because of the microgroove structure and biocompatibility of the film, the cardiomyocytes could be induced into a highly ordered arrangement with recoverable autonomous beating. Meanwhile, the opposite surface of the film was imparted with defect-free 2D photonic crystal structures, giving it a vivid structural color and photonic band gap (PBG). We demonstrated that the flexible film would undergo synchronous volume or shape changes with the cardiomyocytes' elongation and contraction during the beating process, showing cyclic changes of the structural color, which in turn could be converted into force. In addition, by integrating this anisotropic structural color graphene film with microfluidic channels, a novel and accurate heart-on-a-chip platform was established for real-time cell monitoring and drug screening. These characteristics of the present anisotropic structural color graphene films make them extremely valuable in the field of biomedicine. [ABSTRACT FROM AUTHOR]

New drug development is currently very expensive and time-consuming. In addition, some drugs that are approved after animal and clinical trials have their approval revoked because of adverse effects. About 30% of such drugs have heart side effects. Conventional cell-based drug toxicity tests are performed under conditions entirely different from the in vivo environment, and animal testing for drug evaluation has limitations because of differences between species. Therefore, researchers are increasingly focusing on developing models that can overcome these limitations to enable accurate drug toxicity tests. This review outlines biomimetic in vitro heart platforms, such as heart organoids, 3-dimensional bioprinting, and heart-on-a-chip devices, and describes their advantages, limitations, future perspectives. The development and use of effective cardiac biomimetic models could contribute to the development of alternatives to animal testing by providing more specific information on drug metabolism and reducing the rate of failure in later stages of drug development.

Epicardial cells (EPIs) form the outer layer of the heart and play an important role in development and disease. Current heart-on-a-chip platforms still do not fully mimic the native cardiac environment due to the absence of relevant cell types, such as EPIs. Here, using the Biowire II platform, engineered cardiac tissues with an epicardial outer layer and inner myocardial structure are constructed, and an image analysis approach is developed to track the EPI cell migration in a beating myocardial environment. Functional properties of EPI cardiac tissues improve over two weeks in culture. In conditions mimicking ischemia reperfusion injury (IRI), the EPI cardiac tissues experience less cell death and a lower impact on functional properties. EPI cell coverage is significantly reduced and more diffuse under normoxic conditions compared to the post-IRI conditions. Upon IRI, migration of EPI cells into the cardiac tissue interior is observed, with contributions to alpha smooth muscle actin positive cell population. Altogether, a novel heart-on-a-chip model is designed to incorporate EPIs through a formation process that mimics cardiac development, and this work demonstrates that EPI cardiac tissues respond to injury differently than epicardium-free controls, highlighting the importance of including EPIs in heart-on-a-chip constructs that aim to accurately mimic the cardiac environment., (© 2024 The Authors. Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

Development of new drugs is of high interest for the field of cardiac and cardiovascular diseases, which are a dominant cause of death worldwide. Before being allowed to be used and distributed, every new potentially therapeutic compound must be strictly validated during preclinical and clinical trials. The preclinical studies usually involve the in vitro and in vivo evaluation. Due to the increasing reporting of discrepancy in drug effects in animal and humans and the requirement to reduce the number of animals used in research, improvement of in vitro models based on human cells is indispensable. Primary cardiac cells are difficult to access and maintain in cell culture for extensive experiments; therefore, the human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) became an excellent alternative. This technology enables a production of high number of patient- and disease-specific cardiomyocytes and other cardiac cell types for a large-scale research. The drug effects can be extensively evaluated in the context of electrophysiological responses with a use of well-established tools, such as multielectrode array (MEA), patch clamp, or calcium ion oscillation measurements. Cardiotoxicity, which is a common reason for withdrawing drugs from marketing or rejection at final stages of clinical trials, can be easily verified with a use of hiPSC-CM model providing a prediction of human-specific responses and higher safety of clinical trials involving patient cohort. Abovementioned studies can be performed using two-dimensional cell culture providing a high-throughput and relatively lower costs. On the other hand, more complex structures, such as engineered heart tissue, organoids, or spheroids, frequently applied as co-culture systems, represent more physiological conditions and higher maturation rate of hiPSC-derived cells. Furthermore, heart-on-a-chip technology has recently become an increasingly popular tool, as it implements controllable culture conditions, application of various stimulations and continuous parameters read-out. This paper is an overview of possible use of cardiomyocytes and other cardiac cell types derived from hiPSC as in vitro models of heart in drug research area prepared on the basis of latest scientific reports and providing thorough discussion regarding their advantages and limitations. [ABSTRACT FROM AUTHOR]

Heart disease is one of the leading causes of death in the world. There is a growing demand for in vitro cardiac models that can recapitulate the complex physiology of the cardiac tissue. These cardiac models can provide a platform to better understand the underlying mechanisms of cardiac development and disease and aid in developing novel treatment alternatives and platforms toward personalized medicine. Herein, a summary of engineered cardiac platforms is presented. Basic design considerations for replicating the heart's microenvironment are discussed considering the anatomy of the heart. This is followed by a detailed summary of the currently available biomaterial platforms for modeling the heart tissue in vitro. These in vitro models include 2D surface modified structures, 3D molded structures, porous scaffolds, electrospun scaffolds, bioprinted structures, and heart‐on‐a‐chip devices. The challenges faced by current models and the future directions of in vitro cardiac models are also discussed. Engineered in vitro tissue models utilizing patients’ own cells can potentially revolutionize the way we develop treatment and diagnostic alternatives.

Accumulating evidence indicates that air pollution contributes to serious and fatal damage to the cardiovascular system, yet the mechanisms that drive air pollution associated cardiovascular disease and dysfunction remain unclear. In an effort to create a more predictive in vitro model, a 3D platform, known as integrated vasculature for assessing dynamic events is used, that supports the combination of dense human induced pluripotent stem cell derived cardiac tissue and vascular interface, to unravel the impact of nanoscale air pollution on endothelial cells and cardiac tissue. Air pollution relevant nanoparticles (CuO, SiO2) and a control (Au) are used to predict the toxic effects on the cardiovascular system under perfusion. It is demonstrated that CuO nanoparticles are highly toxic, as they are able to translocate into the cardiac tissue and induce electrical and contractile dysfunction through generation of reactive oxygen species and subsequently lead to disruption of cardiac troponin T and secretion of biomarkers associated with cardiac injury (B‐type natriuretic peptide, N‐terminated pro‐hormone BNP, and Troponin I). SiO2, on the other hand, causes the secretion of pro‐inflammatory cytokines, and modulates the intracellular Ca2+ handling. This microengineering approach may offer new opportunities to more accurately model cardiovascular responses to nm‐sized air pollution. [ABSTRACT FROM AUTHOR]

Modeling of human organs has long been a task for scientists in order to lower the costs of therapeutic development and understand the pathological onset of human disease. For decades, despite marked differences in genetics and etiology, animal models remained the norm for drug discovery and disease modeling. Innovative biofabrication techniques have facilitated the development of organ-on-a-chip technology that has great potential to complement conventional animal models. However, human organ as a whole, more specifically the human heart, is difficult to regenerate in vitro , in terms of its chamber specific orientation and its electrical functional complexity. Recent progress with the development of induced pluripotent stem cell differentiation protocols, made recapitulating the complexity of the human heart possible through the generation of cells representative of atrial & ventricular tissue, the sinoatrial node, atrioventricular node and Purkinje fibers. Current heart-on-a-chip approaches incorporate biological, electrical, mechanical, and topographical cues to facilitate tissue maturation, therefore improving the predictive power for the chamber-specific therapeutic effects targeting adult human. In this review, we will give a summary of current advances in heart-on-a-chip technology and provide a comprehensive outlook on the challenges involved in the development of human physiologically relevant heart-on-a-chip. Unlabelled Image [ABSTRACT FROM AUTHOR]

l A heart-on-a-chip platform with online monitoring of contractile behavior for 3D cardiac tissue constructs was developed via an image processing system and a piezoelectric sensing system at the same time. l The image processing system measured the deformation of micro-pillar array during contraction and offered in situ multi-site detection of contractile behavior. l The piezoelectric sensing system provided the contractile behavior of the entire cardiac tissues by measuring the stresses exerted on the PVDF film due to the deformation of pillars. l The results demonstrated that this heart-on-a-chip platform had potential use in cardio-related drug screening application. Heart-on-a-chip devices have recently emerged as a viable and promising model for drug screening applications, owing to its capability of capturing important biological and physiological parameters of cardiac tissue. However, most heart-on-a-chips are not developed for online and continuous monitoring of contractile behavior, which are the main functional characteristics of cardiac tissue. In this study, we designed and investigated on a heart-on-a-chip platform that provides online monitoring of contractile behavior of a 3D cardiac tissue construct. The contractile behavior include contraction force, frequency, and synchronization. They can be evaluated by an image processing system and a piezoelectric sensing system simultaneously. Based on the deformation of a micro-pillar array embedded within the 3D cardiac tissue upon subjected to cardiac contraction, the image processing system provides in situ multi-site detection of the contractile behavior. At the same time, the piezoelectric sensing system measures the contractile behavior of the entire cardiac tissue construct. A 3D cardiac tissue construct was successfully fabricated. Then the heart-on-a-chip platform was validated by applying various motion patterns on the micro-pillars, which mimicked the contraction patterns of the 3D cardiac tissue. The drug reactivity of the 3D cardiac tissue construct after a treatment of isoproterenol and doxorubicin was evaluated by measuring the contractile behavior via the image processing and the piezoelectric sensing systems. The results from the drug reactivity provided by both these measurement systems were consistent with previous reports, demonstrating the reliability of the heart-on-a-chip platform and its potential for use in cardio-related drug screening applications. [ABSTRACT FROM AUTHOR]

Cardiac fibrosis acts as a serious worldwide health issue due to its prevalence in numerous forms of cardiac disease and its essential link to cardiac failure. Considering the efficiency of stem cell therapy for cardiac fibrosis, great efforts have been dedicated to developing accurate models for investigating their underlying therapeutic mechanisms. Herein we present an elaborate biomimetic cardiac fibrosis-on-a-chip based on Janus structural color film (SCF) to provide microphysiological visuals for stem cell therapeutic studies. By coculturing cardiomyocytes (CMs) and cardiac fibroblasts (FBs) on Janus SCF with fibrosis induction, the chip can recreate physiological intercellular crosstalk within the fibrotic microenvironment, elucidating the physiological alterations of fibrotic hearts. In particular, the Janus structural color film possesses superior perceptual capabilities for capturing and responding to a weak cardiac force, demonstrating synchronized structural color shifts. Based on these features, we have not only explored the dynamic relationship between color mapping and the evaluated disease phenotype but also demonstrated the self-reporting capacity of the cardiac fibrosis-on-a-chip for the assessment of mesenchymal stem cell-derived exosome therapy. These features suggest that such a chip can potentially facilitate the evolution of precision medicine strategies and create a protocol for preclinical cardiac drug screening.

Cardiovascular disease is a major cause of mortality worldwide, and current preclinical models including traditional animal models and 2D cell culture models have limitations in replicating human native heart physiology and response to drugs. Heart-on-a-chip (HoC) technology offers a promising solution by combining the advantages of cardiac tissue engineering and microfluidics to create in vitro 3D cardiac models, which can mimic key aspects of human microphysiological systems and provide controllable microenvironments. Herein, recent advances in HoC technologies are introduced, including engineered cardiac microtissue construction in vitro, microfluidic chip fabrication, microenvironmental stimulation, and real-time feedback systems. The development of cardiac tissue engineering methods is focused for 3D microtissue preparation, advanced strategies for HoC fabrication, and current applications of these platforms. Major challenges in HoC fabrication are discussed and the perspective on the potential for these platforms is provided to advance research and clinical applications., (© 2023 Wiley-VCH GmbH.)

Recent advances in heart-on-a-chip systems hold great promise to facilitate cardiac physiological, pathological, and pharmacological studies. This review focuses on the development of heart-on-a-chip systems with tissue-specific functionalities. For one thing, the strategies for developing cardiac microtissues on heart-on-a-chip systems that closely mimic the structures and behaviors of the native heart are analyzed, including the imitation of cardiac structural and functional characteristics. For another, the development of techniques for real-time monitoring of biophysical and biochemical signals from cardiac microtissues on heart-on-a-chip systems is introduced, incorporating cardiac electrophysiological signals, contractile activity, and biomarkers. Furthermore, the applications of heart-on-a-chip systems in intelligent cardiac studies are discussed regarding physiological/pathological research and pharmacological assessment. Finally, the future development of heart-on-a-chip toward a higher level of systematization, integration, and maturation is proposed., 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., (© 2023 The Authors.)

Regenerative medicine and stem cells could prove to be an effective solution to the problem of treating heart failure caused by ischemic heart disease. However, further studies on the understanding of the processes which occur during the regeneration of damaged tissue are needed. Microfluidic systems, which provide conditions similar to in vivo, could be useful tools for the development of new therapies using stem cells. We investigated how mesenchymal stem cells (MSCs) affect the metabolic activity of cardiac cells (rat cardiomyoblasts and human cardiomyocytes) incubated with a potent uncoupler of mitochondrial oxidative phosphorylation under microfluidic conditions. A cyanide p-trifluoromethoxyphenylhydrazone (FCCP) was used to mimic disfunctions of mitochondria of cardiac cells. The study was performed in a microfluidic system integrated with nanofiber mats made of poly-l-lactid acid (PLLA) or polyurethane (PU). The microsystem geometry allows four different cell cultures to be conducted under different conditions (which we called: normal, abnormal—as both a mono- and co-culture). Metabolic activity of the cells, based on the bioluminescence assay, was assessed in the culture’s performed in the microsystem. It was proved that stem cells increased metabolic activity of cardiac cells maintained with FCCP.

Human cardiac slices have emerged as a promising model of the human heart for scientific research and drug testing. Retaining the normal tissue architecture, a multi-cell type environment, and the native extracellular matrix, human cardiac slices faithfully replicate organ-level adult cardiac physiology. Previously, we demonstrated that human cardiac tissue slices cultured for 24 h maintained normal electrophysiology. In this project, we further optimized the organotypic culture condition to maintain normal electrophysiology of the human cardiac slices for 4 days. The prolonged culture of human cardiac tissue slices demonstrated here enables the study of chronic drug effects, gene therapies, and gene editing. To achieve greater control of the culture environment, we have also developed an automated, self-contained heart-on-a-chip system. The culture system supports media circulation, oxygenation, temperature control, electrical stimulation, and static mechanical loading. The culture parameters can be individually adjusted to establish the optimal culture condition to achieve long-term culture and to minimize tissue dedifferentiation. The development of the heart-on-a-chip technology presented here further encourages the use of organotypic human cardiac slices as a platform for pre-clinical drug testing and research in human cardiac physiology. [ABSTRACT FROM AUTHOR]

Abstract The evolution of micro and nanofabrication approaches significantly spurred the advancements of cardiac tissue engineering over the last decades. Engineering in the micro and nanoscale allows for the rebuilding of heart tissues using cardiomyocytes. The breakthrough of human induced pluripotent stem cells expanded this field rendering the development of human tissues from adult cells possible, thus avoiding the ethical issues of the usage of embryonic stem cells but also creating patient-specific human engineered tissues. In the case of the heart, the combination of cardiomyocytes derived from human induced pluripotent stem cells and micro/nano engineering devices gave rise to new therapeutic approaches of cardiac diseases. In this review, we survey the micro and nanofabrication methods used for cardiac tissue engineering, ranging from clean room-based patterning (such as photolithography and plasma etching) to electrospinning and additive manufacturing. Subsequently, we report on the main approaches of microfluidics for cardiac culture systems, the so-called "Heart on a Chip", and we assess their efficacy for future development of cardiac disease modeling and drug screening platforms. Highlights • Overview of micro/nanofabrication methods for cardiac tissue engineering. • Clean room-based patterning, electrospinning and additive manufacturing. • Microfluidic technologies for creating disease models and drug screening systems. • Electrical and mechanical stimulation of cardiac cells on chip. • Micro-engineered "Human Heart on a Chip". [ABSTRACT FROM AUTHOR]

• Maturation of hiPSC-CMs is key as pluripotent stem cell-derived cardiomyocytes initially express a foetal phenotype. • A biomimetic environment is needed with multiple stimuli to achieve higher maturation of hiPSC-CMs. • Electrospun scaffolds could recapitulate the natural 3D environment for tissue engineering. • Electrically conductive polymers are essential for cardiac tissue engineering. • 3D printing and bioreactors support the long-term cell culture of hiPSC-CMs. Cardiovascular diseases are the leading cause of death worldwide. Several strategies – small molecules, gene therapy, surgeries, cardiac rehabilitation – have been developed to treat patients with cardiac disorders or to prevent the disease. A novel promising tool is the application of induced pluripotent stem cell-derived human cardiomyocytes; however, their clinical application is hampered by their fundamentally immature characteristics. Although several approaches were recently proven to support certain aspects of cardiac maturation, it has been discovered that any kind of stimulus itself is insufficient to induce proper maturation of these cardiomyocytes. Therefore, researchers have developed multifactorial approaches linking biotechnology, life sciences and material sciences toolsets. In this review, we summarise the current state-of-the-art methodologies in cardiac differentiation and maturation with the application of biocompatible materials and automated cell culture systems combined with various electrical, mechanical, topological, and biochemical stimuli. [ABSTRACT FROM AUTHOR]

The successful translation of organ-on-a-chip devices requires the development of an automated workflow for device fabrication, which is challenged by the need for precise deposition of multiple classes of materials in micro-meter scaled configurations. Many current heart-on-a-chip devices are produced manually, requiring the expertise and dexterity of skilled operators. Here, we devised an automated and scalable fabrication method to engineer a Biowire II multiwell platform to generate human iPSC-derived cardiac tissues. This high-throughput heart-on-a-chip platform incorporated fluorescent nanocomposite microwires as force sensors, produced from quantum dots and thermoplastic elastomer, and 3D printed on top of a polystyrene tissue culture base patterned by hot embossing. An array of built-in carbon electrodes was embedded in a single step into the base, flanking the microwells on both sides. The facile and rapid 3D printing approach efficiently and seamlessly scaled up the Biowire II system from an 8-well chip to a 24-well and a 96-well format, resulting in an increase of platform fabrication efficiency by 17,5000-69,000% per well. The device's compatibility with long-term electrical stimulation in each well facilitated the targeted generation of mature human iPSC-derived cardiac tissues, evident through a positive force-frequency relationship, post-rest potentiation, and well-aligned sarcomeric apparatus. This system's ease of use and its capacity to gauge drug responses in matured cardiac tissue make it a powerful and reliable platform for rapid preclinical drug screening and development., (© 2023 The Authors.)

The heart is a complex organ composed of distinct cell types, such as cardiomyocytes, cardiac fibroblasts, endothelial cells, smooth muscle cells, neuronal cells and immune cells. All these cell types contribute to the structural, electrical and mechanical properties of the heart. Genetic manipulation and lineage tracing studies in mice have been instrumental in gaining critical insights into the networks regulating cardiac cell lineage specification, cell fate and plasticity. Such knowledge has been of fundamental importance for the development of efficient protocols for the directed differentiation of pluripotent stem cells (PSCs) in highly specialized cardiac cell types. In this review, we summarize the evolution and current advances in protocols for cardiac subtype specification, maturation, and assembly in cardiac microtissues and organoids., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Engineering design of microbioreactors (MBRs) and organ-on-chip (OoC) devices can take advantage of established design science theory, in which systematic evaluation of functional concepts and user requirements are analyzed. This is commonly referred to as a conceptual design. This review article compares how common conceptual design principles are applicable to MBR and OoC devices. The complexity of this design, which is exemplified byMBRs for scaled-down cell cultures in bioprocess development and drug testing in OoCs for heart and eye, is discussed and compared with previous design solutions of MBRs and OoCs, from the perspective of how similarities in understanding design from functionality and user purpose perspectives can more efficiently be exploited. The review can serve as a guideline and help the future design of MBR and OoC devices for cell culture studies. [ABSTRACT FROM AUTHOR]