From Pluripotency to Function: The Journey, Challenges, and Future of iPSC‐Based Heart Organoids

Cardiovascular disease (CVD) remains a leading cause of global mortality. The Global Burden of Cardiovascular Diseases Report indicated that CVD was responsible for approximately 19.8 million deaths worldwide in 2022 [1]. In response to this formidable challenge, basic and translational research in CVD has intensified. However, the field continues to grapple with significant limitations, primarily because traditional two-dimensional cell cultures and animal models often fail to fully recapitulate the complex physiology of the human cardiovascular system. For instance, rodent models (mice and rats) lack key pathological features of human atherosclerotic plaque rupture [2]. To overcome these persistent obstacles, technologies involving induced pluripotent stem cells (iPSCs) and organoids have rapidly emerged as promising new platforms for CVD research, offering more human-relevant models for investigation. IPSCs were first pioneered by Takahashi and Yamanaka in 2006 [3], who reprogrammed adult somatic cells into an embryonic stem cell-like state by introducing key transcription factors, namely Oct4, Sox2, Klf4, and c-Myc. A defining characteristic of iPSCs is their capacity to differentiate into various cell types derived from the three primary germ layers under appropriate conditions [4]. This ability allows them to be directed into functional cells that retain the donor patient's specific disease genetics, making them exceptionally valuable for in vitro disease modeling and drug screening applications [5]. Despite this potential, iPSC technology still confronts challenges related to differentiation efficiency, genomic stability, and the need for further optimization of culture conditions and differentiation protocols. The generation of heart organoids from iPSCs is a multi-step process. It typically begins by directing iPSCs toward a mesodermal lineage. Subsequently, specific inducing factors such as bone morphogenetic protein 4, Activin A, and vascular endothelial growth factor are used to steer differentiation toward cardiac lineages (e.g., cardiomyocytes) or vascular lineages (e.g., endothelial and smooth muscle cells). The next critical phase involves co-culturing these diverse cardiovascular cells—including cardiomyocytes, endothelial cells, and fibroblasts—within a three-dimensional (3D) environment, such as a suspension system or a structured scaffold. Current methodologies to achieve this include the embryoid body formation technique, the use of microfluidic chips [6] to create controlled microenvironments, and advanced 3D bioprinting. IPSC-derived heart organoids are now extensively utilized as in vitro models for both inherited and acquired heart diseases. They are particularly impactful for studying hereditary cardiomyopathies, such as dilated cardiomyopathy and hypertrophic cardiomyopathy, as well as various arrhythmias. By using iPSCs derived from patients with these conditions, researchers can generate organoids that manifest key disease-specific phenotypes, providing a powerful platform for mechanistic studies and therapeutic discovery. Notwithstanding their considerable promise, iPSC and organoid technologies face several substantial hurdles. A primary limitation is the relative immaturity of most iPSC-derived cardiomyocytes, which often exhibit fetal-like characteristics rather than fully adult phenotypes. This immaturity is reflected in their action potentials, ion channel electrophysiology, and contractile properties, limiting their ability to completely model adult heart disease [7]. Furthermore, heterogeneity is a significant concern, arising from variations in the epigenetic memory of donor-derived iPSCs, which can lead to biased differentiation outcomes. Technical variables, including initial cell density, passage number, and batch-to-batch inconsistencies in extracellular matrices like Matrigel, also contribute to variability in organoid morphology and gene expression profiles. Future efforts will focus on leveraging bioengineering strategies to enhance the fidelity and utility of cardiovascular organoids. Optimizing culture conditions through the application of electrical stimulation, precise 3D bioprinting, and controlled growth factor gradients is expected to drive the maturation of iPSC-derived cardiac and vascular cells toward more adult-like states. The integration of microfluidics and nanotechnology will enable the creation of more sophisticated and physiologically relevant culture systems, potentially paving the way for automated and standardized production protocols. Nanomaterials and advanced delivery systems, such as nanofiber scaffolds, can be employed to provide better structural support and biochemical cues [8]. A major frontier lies in the application of iPSC organoids for tissue regeneration and transplantation. Combining these organoids with gene-editing tools like CRISPR/Cas9 offers the potential to correct disease-causing mutations in patient-specific iPSCs, restoring normal cellular function and bringing the prospect of curing certain hereditary heart conditions closer to reality. In summary, iPSC-induced heart and vascular organoids represent a transformative advance as emerging 3D in vitro models for CVD research. They offer a superior ability to mimic the structural and functional complexity of human heart tissues compared to traditional two-dimensional cultures and animal models, thereby enabling more accurate disease modeling, drug screening, and toxicity assessments. Research utilizing these models has already yielded significant insights into hereditary cardiomyopathies, ischemic heart disease, drug-induced cardiotoxicity, and regenerative medicine, with further refinements achieved through multi-lineage co-culture and gene editing. Nevertheless, challenges pertaining to cellular maturity, functional heterogeneity, and a lack of standardization remain and must be addressed through continued optimization of culture protocols and the development of robust regulatory frameworks. Looking forward, the convergence of nanotechnology, bioengineering, and artificial intelligence with organoid technology is poised to accelerate its transition from a research tool to clinical applications and novel therapeutic strategies for cardiovascular diseases. Yuqi Liu: writing – original draft, writing – review and editing. The author has nothing to report. The author has nothing to report. The author has nothing to report. The author declares no conflicts of interest. The author has nothing to report.