Cellular and Engineered Organoids for Cardiovascular Models

Abstract

An ensemble of in vitro cardiac tissue models has been developed over the past several decades to aid our understanding of complex cardiovascular disorders using a reductionist approach. These approaches often rely on recapitulating single or multiple clinically relevant end points in a dish indicative of the cardiac pathophysiology. The possibility to generate disease-relevant and patient-specific human induced pluripotent stem cells has further leveraged the utility of the cardiac models as screening tools at a large scale. To elucidate biological mechanisms in the cardiac models, it is critical to integrate physiological cues in form of biochemical, biophysical, and electromechanical stimuli to achieve desired tissue-like maturity for a robust phenotyping. Here, we review the latest advances in the directed stem cell differentiation approaches to derive a wide gamut of cardiovascular cell types, to allow customization in cardiac model systems, and to study diseased states in multiple cell types. We also highlight the recent progress in the development of several cardiovascular models, such as cardiac organoids, microtissues, engineered heart tissues, and microphysiological systems. We further expand our discussion on defining the context of use for the selection of currently available cardiac tissue models. Last, we discuss the limitations and challenges with the current state-of-the-art cardiac models and highlight future directions.

Keywords: cardiovascular disease; heart; organoids; phenotype; pluripotent stem cells; tissue engineering.

Figure 1.. Deconstructing cardiogenesis using induced pluripotent stem cell (iPSC) technology.

An overview of cell lineage trajectory to derive cardiac cellular subtypes in vitro from pluripotent stem cells. Key factors that induce cell-specific morphogenesis and their identity is depicted in the development pathway. Most commonly, all cardiac cell subtypes originate from the mesendoderm progenitors (KDR1, MESP1) that arise from the primitive streak (PS). Further biphasic Wnt modulation leads to the generation of cardiomyocytes, endothelium. Chamber-specific myocyte cell types are obtained through bone morphogenetic protein (BMP) and retinoic acid (RA) mediated NOTCH signaling activation. Smooth muscles cells and pericytes develop from lateral plate mesoderm (LPM) and paraxial mesoderm (PM). Cardiac fibroblasts are derived from epicardial and mesodermal progenitors. Macrophages and natural killer (NK) cells that naturally reside in the cardiac tissue can be successfully derived from hemogenic precursors. Improving cellular diversity in the human cardiac model systems will offer higher resemblance to cardiac tissue composition.

Figure 2.. Embryoid body-like human cardiac models.

Summary of recent noteworthy human cardiac organoid (CO) models that mimic some features of heart development in vivo. A, The COs can be classified based on the presence or absence of “cavity” that resemble chamber-like features during early heart development. B, A brief summary of the morphogenetic factors or small molecules used to direct the formation of distinct pre-cardiac structures in vitro.

Figure 3.. Hierarchical structure of cardiomyocyte and ECM structure in the heart.

A, Cardiomyocytes shows aligned myofilament structure which is connected to ECM through costamere. Costameres contain focal adhesions (FAs) complex connecting with cytoskeletal actin filaments. B, A cardiomyocyte cultured on 1:7 aspect ratio of rectangular microcontact printed ECM pattern, showing intracellular organization using a DIC image and immunostaining images for vinculin, F-actin and sarcomeric a-actinin in (i-iv), respectively. Scale bar, 10 μm. Reprinted from Bray et al. with permission. Copyright ©2008, Wiley-Liss, Inc. C, Junction formation is occurred at the intercalated disc where the cells are connected end to end. D, Immunostained images of cardiomyocytes cell pair images showing FAs and junction formation and intracellular organization according to culture days. Scale bar, 10 μm. Reprinted from McCain et al. with permission. Copyright ©2012, NAS. E, Fibrous ECM structures supporting cardiomyocytes with their intra/extracellular organization. F, In vitro cardiac tissue organization by ECM geometry patterns. Scale bar, 20μm. Reprinted from Lee et al. with permission. Copyright ©2022, AAAS. G, Hierarchical cardiac muscle tissues are organized into the heart, inducing cyclic blood pumping with coordinated tissue contraction.

Figure 4.. Cardiac microphysiological system for electrical and mechanical functional assessment.

A, Optical mapping system from in vitro cardiac tissue showing anisotropic electrical potential propagation in aligned tissue direction and arrhythmias in disease model. B, Microelectrodes arrays measuring the local electrical potential changes, providing high spatiotemporal bioactivity of the cardiac tissues. C, Muscular thin film (MTF) system to measure contractility by measuring bending force of the laminar tissues that are formed on the cantilevers. D, Engineering heart tissues (EHT) system allowing for monitoring contractility of the cardiac tissue that causes cyclic movement of the posts. E, Biowires deformation caused by contractile cardiac tissue as the tissue is sustained by wires.

Figure 5.. The evolving paradigm of in vitro cardiovascular disease modeling.

A plethora of in vitro cardiac models have been developed over several decades. iPSC technology has fueled the development of human cardiac models to accelerate predictability of cardiovascular disorders and drug responses to complement in vivo studies. The flowchart offers a novice guide on general principles for choosing a more relevant model in the context of the disease in question, and validity of the model based on an understanding of cellular contribution toward the disease, obtaining clinically translatable phenotype, and most importantly the reliability and reproducibility for disease modeling.