Generating Self-Assembling Human Heart Organoids Derived from Pluripotent Stem Cells

Lewis-Israeli, Yonatan R.; Volmert, Brett D.; Huang, Amanda R.; Aguirre, Aitor

doi:10.3791/63097

Cited by 8 publications

(20 citation statements)

References 36 publications

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“…We also reported a high level of reproducibility and control over the differentiation of the hHOs. Lastly, using a transgenic cell line with a fluorescent marker for apoptosis, we observed the absence of a necrotic core, and using doxorubicin as a positive control, we showed apoptotic responses and demonstrated the capacity of this hHO platform to model cardiotoxicity (Lewis‐Israeli et al., 2021).…”

Section: Commentarymentioning

confidence: 68%

“…Although organoid models have been described for a broad range of tissues, progress in the cardiovascular field has been limited. The field of engineered heart tissues has traditionally relied on direct assembly approaches, where different cardiac cell types (obtained from animal tissues or, more recently, differentiated from PSCs) are co‐cultured into a 3D aggregate using various approaches such as layering, surrounding pillars, embedding in hydrogels, or seeding in patterned molds, to name a few (Lewis‐Israeli et al., 2021). More recently, self‐assembling heart organoid protocols have emerged.…”

Section: Commentarymentioning

confidence: 99%

“…They can also be produced in large quantities in a systematic manner for high‐throughput analyses or high‐content screenings (Kim, Koo, & Knoblich, 2020). Heart organoids have been used for a variety of studies, including drug screenings (Mills et al., 2019; Skardal, Shupe, & Atala, 2016), disease development (Lewis‐Israeli et al., 2021; Miyamoto, Nam, Kannan, & Kwon, 2021), and toxicity research (Hoang et al., 2021; Richards et al., 2020). Indeed, PSC‐derived heart organoids provide an ideal platform to study the cardiotoxicity of therapeutic agents.…”

Section: Commentarymentioning

confidence: 99%

“…In recent years, novel stem cell–based technologies have enabled the creation of engineered, highly complex human organ–like 3D tissues in vitro , with properties that recapitulate the physiological setting to a significant extent (Kretzschmar & Clevers, 2016; Paşca, 2019; Rossi et al., 2020; Sasai, 2013; Weeber, Ooft, Dijkstra, & Voest, 2017). These organoids (i.e., “resembling an organ”) are particularly useful to study unapproachable disease states in humans (e.g., early disease progression when symptoms are not still present), or states for which animal models are not well‐suited (Bredenoord, Clevers, & Knoblich, 2017; Lewis‐Israeli et al., 2021; Tuveson & Clevers, 2019). Organoids can also be powerful tools to verify, in human models, what we have learned in animal studies, thus complementing them.…”

Section: Introductionmentioning

confidence: 99%

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Modeling the Effects of Maternal Diabetes on the Developing Human Heart Using Pluripotent Stem Cell–Derived Heart Organoids

et al. 2022

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Congenital heart defects (CHD) constitute the most common type of birth defect in humans. Maternal diabetes during the first trimester of pregnancy (pregestational diabetes, or PGD) is one of the most prominent factors contributing to CHD, and is present in a significant population of female patients with diabetes in reproductive age. PGD is challenging to manage clinically due to the extreme sensitivity of the developing embryo to glucose oscillations, and constitutes a critical health problem for the mother and the fetus. The prevalence of PGD-induced CHD is increasing due to the ongoing diabetes epidemic. While studies using animal models and cells in culture have demonstrated that PGD alters critical cellular and developmental processes, the mechanisms remain obscure, and it is unclear to what extent these models recapitulate PGD-induced CHD in humans. Clinical practice precludes direct studies in developing human embryos, further highlighting the need for physiologically relevant models. To bypass many of these technical and ethical limitations, we describe here a human pluripotent stem cell (hPSC)-based method to generate developmentally relevant self-organizing human heart organoids. By using glucose and insulin to mimic the diabetic environment that the embryo faces in PGD, this system allows modeling critical features of PGD in a human system with relevant physiology, structure, and cell types. The protocol starts with the generation of hPSC-derived embryoid bodies in a 96-well plate, followed by a small molecule-based three-step Wnt activation/inhibition/activation strategy. Organoids are then differentiated under healthy (normal insulin and glucose) and diabetic conditions (high insulin and glucose) over time, allowing for the study of the effects of pregestational diabetes on the developing human heart. We also provide an immunofluorescence protocol for comparing, characterizing, and analyzing the differences between the healthy and diabetic organoids, and comment on additional steps for preparing the organoids for analysis by other techniques after differentiation.

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Section: Commentarymentioning

confidence: 68%

Section: Commentarymentioning

confidence: 99%

Section: Commentarymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Modeling the Effects of Maternal Diabetes on the Developing Human Heart Using Pluripotent Stem Cell–Derived Heart Organoids

et al. 2022

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“…Moreover, the inhibition of WNT signaling at the cardiac mesoderm stage seems crucial for CM specification but is not necessary for cardioid self-organization. Recent work uses a protocol based on three sequential modulation steps of the WNT pathway (activation/inhibition/activation) at specific time points on suspension EBs to drive the production of significant heart-like structures in terms of organization, functionality, cardiac cell type complexity, ECM composition, and vascularization [175]. The gene expression profile of cardioids showed upregulation of cardiac-specific genes and downregulation of WNT signaling at day 20 differentiation and significantly higher gene expression of transforming growth factor β (TGF-β) signaling (such as TGFβ1, TGFβ2, TGFβ3, TGFβR1, and TGFβR2) and cardiac-specific genes (MYL4, MYH7, and NKX2.5 [176]).…”

Section: Cardioidsmentioning

confidence: 99%

From Spheroids to Organoids: The Next Generation of Model Systems of Human Cardiac Regeneration in a Dish

Scalise

Marino

Salerno

et al. 2021

IJMS

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Organoids are tiny, self-organized, three-dimensional tissue cultures that are derived from the differentiation of stem cells. The growing interest in the use of organoids arises from their ability to mimic the biology and physiology of specific tissue structures in vitro. Organoids indeed represent promising systems for the in vitro modeling of tissue morphogenesis and organogenesis, regenerative medicine and tissue engineering, drug therapy testing, toxicology screening, and disease modeling. Although 2D cell cultures have been used for more than 50 years, even for their simplicity and low-cost maintenance, recent years have witnessed a steep rise in the availability of organoid model systems. Exploiting the ability of cells to re-aggregate and reconstruct the original architecture of an organ makes it possible to overcome many limitations of 2D cell culture systems. In vitro replication of the cellular micro-environment of a specific tissue leads to reproducing the molecular, biochemical, and biomechanical mechanisms that directly influence cell behavior and fate within that specific tissue. Lineage-specific self-organizing organoids have now been generated for many organs. Currently, growing cardiac organoid (cardioids) from pluripotent stem cells and cardiac stem/progenitor cells remains an open challenge due to the complexity of the spreading, differentiation, and migration of cardiac muscle and vascular layers. Here, we summarize the evolution of biological model systems from the generation of 2D spheroids to 3D organoids by focusing on the generation of cardioids based on the currently available laboratory technologies and outline their high potential for cardiovascular research.

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Self-organizing human heart assembloids with autologous and developmentally relevant cardiac neural crest-derived tissues

Kostina,

Kiselev,

Huang

et al. 2024

Preprint

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Neural crest cells (NCCs) are a multipotent embryonic cell population of ectodermal origin that extensively migrate during early development and contribute to the formation of multiple tissues. Cardiac NCCs play a critical role in heart development by orchestrating outflow tract septation, valve formation, aortic arch artery patterning, parasympathetic innervation, and maturation of the cardiac conduction system. Abnormal migration, proliferation, or differentiation of cardiac NCCs can lead to severe congenital cardiovascular malformations. However, the complexity and timing of early embryonic heart development pose significant challenges to studying the molecular mechanisms underlying NCC-related cardiac pathologies. Here, we present a sophisticated functional model of human heart assembloids derived from induced pluripotent stem cells, which, for the first time, recapitulates cardiac NCC integration into the human embryonic heartin vitro. NCCs successfully integrated at developmentally relevant stages into heart organoids, and followed developmental trajectories known to occur in the human heart. They demonstrated extensive migration, differentiated into cholinergic neurons capable of generating nerve impulses, and formed mature glial cells. Additionally, they contributed to the mesenchymal populations of the developing outflow tract. Through transcriptomic analysis, we revealed that NCCs acquire molecular features of their cardiac derivatives as heart assembloids develop. NCC-derived parasympathetic neurons formed functional connections with cardiomyocytes, promoting the maturation of the cardiac conduction system. Leveraging this model’s cellular complexity and functional maturity, we uncovered that early exposure of NCCs to antidepressants harms the development of NCC derivatives in the context of the developing heart. The commonly prescribed antidepressant Paroxetine disrupted the expression of a critical early neuronal transcription factor, resulting in impaired parasympathetic innervation and functional deficits in cardiac tissue. This advanced heart assembloid model holds great promise for high-throughput drug screening and unraveling the molecular mechanisms underlying NCC-related cardiac formation and congenital heart defects.IN BRIEFHuman neural crest heart assembloids resembling the major directions of neural crest differentiation in the human embryonic heart, including parasympathetic innervation and the mesenchymal component of the outflow tract, provide a human-relevant embryonic platform for studying congenital heart defects and drug safety.

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Generating Self-Assembling Human Heart Organoids Derived from Pluripotent Stem Cells

Cited by 8 publications

References 36 publications

Modeling the Effects of Maternal Diabetes on the Developing Human Heart Using Pluripotent Stem Cell–Derived Heart Organoids

Modeling the Effects of Maternal Diabetes on the Developing Human Heart Using Pluripotent Stem Cell–Derived Heart Organoids

From Spheroids to Organoids: The Next Generation of Model Systems of Human Cardiac Regeneration in a Dish

Self-organizing human heart assembloids with autologous and developmentally relevant cardiac neural crest-derived tissues

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