Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) generally have an immature fetal-like phenotype when directly compared to isolated CMs from human hearts, despite significant advance in differentiation of human pluripotent stem cells (hPSCs) to multiple cardiac lineages. Therefore, hPSC-CMs may not accurately mimic all facets of healthy and diseased human adult CMs. During embryonic development, the cardiac extracellular matrix (ECM) experiences a gradual assembly of matrix proteins that transits along the maturation of CMs. Mimicking these dynamic stages may contribute to hPSC-CMs maturation in vitro. Thus, in this review, we describe the progressive build-up of the cardiac ECM during embryonic development, the ECM of the adult human heart and the application of natural and synthetic biomaterials for cardiac tissue engineering with hPSC-CMs.

Advanced in vitro models that recapitulate the structural organization and function of the human heart are highly needed for accurate disease modeling, more predictable drug screening, and safety pharmacology. Conventional 3D Engineered Heart Tissues (EHTs) lack heterotypic cell complexity and culture under flow, whereas microfluidic Heart‐on‐Chip (HoC) models in general lack the 3D configuration and accurate contractile readouts. In this study, an innovative and user‐friendly HoC model is developed to overcome these limitations, by culturing human pluripotent stem cell (hPSC)‐derived cardiomyocytes (CMs), endothelial (ECs)‐ and smooth muscle cells (SMCs), together with human cardiac fibroblasts (FBs), underflow, leading to self‐organized miniaturized micro‐EHTs (µEHTs) with a CM‐EC interface reminiscent of the physiological capillary lining. µEHTs cultured under flow display enhanced contractile performance and conduction velocity. In addition, the presence of the EC layer altered drug responses in µEHT contraction. This observation suggests a potential barrier‐like function of ECs, which may affect the availability of drugs to the CMs. These cardiac models with increased physiological complexity, will pave the way to screen for therapeutic targets and predict drug efficacy.

Cardiomyocytes derived from human pluripotent stem cells (hPSC-CMs) hold a great potential as human in vitro models for studying heart disease and for drug safety screening. Nevertheless, their associated immaturity relative to the adult myocardium limits their utility in cardiac research. In this study, we describe the development of a platform for generating three-dimensional engineered heart tissues (EHTs) from hPSC-CMs for the measurement of force while under mechanical and electrical stimulation. The modular and versatile EHT platform presented here allows for the formation of three tissues per well in a 12-well plate format, resulting in 36 tissues per plate. We compared the functional performance of EHTs and their histology in three different media and demonstrated that tissues cultured and maintained in maturation medium, containing triiodothyronine (T3), dexamethasone, and insulin-like growth factor-1 (TDI), resulted in a higher force of contraction, sarcomeric organization and alignment, and a higher and lower inotropic response to isoproterenol and nifedipine, respectively. Moreover, in this study, we highlight the importance of integrating a serum-free maturation medium in the EHT platform, making it a suitable tool for cardiovascular research, disease modeling, and preclinical drug testing.

Cardiovascular diseases caused by loss of functional cardiomyocytes (CMs) are a major cause of mortality and morbidity worldwide due in part to the low regenerative capacity of the adult human heart. Human pluripotent stem cell (hPSC)-derived cardiovascular progenitor cells (CPCs) are a potential cell source for cardiac repair. The aim of this study was to examine the impact of extensive remuscularization and coincident revascularization on cardiac remodelling and function in a mouse model of myocardial infarction (MI) by transplanting doxycycline (DOX)-inducible (Tet-On-MYC) hPSC-derived CPCs in vivo and inducing proliferation and cardiovascular differentiation in a drug-regulated manner. CPCs were injected firstly at a non-cardiac site in Matrigel suspension under the skin of immunocompromised mice to assess their commitment to the cardiovascular lineage and ability to self-renew or differentiate in vivo when instructed by systemically delivered factors including DOX and basic fibroblast growth factor (bFGF). CPCs in Matrigel were then injected intra-myocardially in mice subjected to MI to assess whether expandable CPCs could mediate cardiac repair. Transplanted CPCs expanded robustly both subcutis and in the myocardium using the same DOX/growth factor inducing regime. Upon withdrawal of these cell-renewal factors, CPCs differentiated with high efficiency at both sites into the major cardiac lineages including CMs, endothelial cells, and smooth muscle cells. After MI, engraftment of CPCs in the heart significantly reduced fibrosis in the infarcted area and prevented left ventricular remodelling, although cardiac function determined by magnetic resonance imaging was unaltered. Replacement of large areas of muscle may be required to regenerate the heart of patients following MI. Our human/mouse model demonstrated that proliferating hPSC-CPCs could reduce infarct size and fibrosis resulting in formation of large grafts. Importantly, the results suggested that expanding transplanted cells in situ at the progenitor stage maybe be an effective alternative causing less tissue damage than injection of very large numbers of CMs.

In order to fabricate functional organoids and microtissues, a high cell density is generally required. As such, the placement of cell suspensions in molds or microwells to allow for cell concentration by sedimentation is the current standard for the production of organoids and microtissues. Even though molds offer some level of control over the shape of the resulting microtissue, this control is limited as microtissues tend to compact towards a sphere after sedimentation of the cells. 3D bioprinting on the other hand offers complete control over the shape of the resulting structure. Even though the printing of dense cell suspensions in the ink has been reported, extruding dense cellular suspensions is challenging and generally results in high shear stresses on the cells and a poor shape fidelity of the print. As such, additional materials such as hydrogels are added in the bioink to limit shear stresses, and to improve shape fidelity and resolution. The maximum cell concentration that can be incorporated in a hydrogel-based ink before the ink’s rheological properties are compromised, is significantly lower than the concentration in a tissue equivalent. Additionally, the hydrogel components often interfere with cellular self-assembly processes. To circumvent these limitations, we report a simple and inexpensive xanthan bath based embedded printing method to 3D print dense functional linear tissues using dilute particle suspensions consisting of cells, spheroids, hydrogel beads, or combinations thereof. Using this method, we demonstrated the self-organization of functional cardiac tissue fibers with a layer of epicardial cells surrounding a body of cardiomyocytes.

Cardiotoxicity remains a major cause of drug withdrawal, partially due to lacking predictability of animal models. Additionally, risk of cardiotoxicity following treatment of cancer patients is treatment limiting. It is unclear which patients will develop heart failure following therapy. Human pluripotent stem cell (hPSC)-derived cardiomyocytes present an unlimited cell source and may offer individualized solutions to this problem. We developed a platform to predict molecular and functional aspects of cardiotoxicity. Our platform can discriminate between the different cardiotoxic mechanisms of existing and novel anthracyclines Doxorubicin, Aclarubicin, and Amrubicin. Doxorubicin and Aclarubicin unlike Amrubicin substantially affected the transcriptome, mitochondrial membrane integrity, contractile force and transcription factor availability. Cardiomyocytes recovered fully within two or three weeks, corresponding to the intermittent clinical treatment regimen. Our system permits the study of hPSC-cardiomyocyte recovery and the effects of accumulated dose after multiple dosing, allowing individualized cardiotoxicity evaluation, which effects millions of cancer patients treated annually.