ABSTRACT

Currently, human induced pluripotent stem cell (hiPSC) technology (a Nobel Prize winning technology that can turn abundant human cells, such as skin cells and fat cells, into stem cells that can give rise to every other cell type in the body) has made it possible to model human heart diseases in cell culture as a "disease-in-a-dish." However, the grand challenge in current hiPSC disease modeling is that these models have tended to simplify the diseases as being a result of a single defective gene without taking into account the many other influences of genetic-environmental interactions. Specifically, hypertrophic cardiomyopathy (HCM), a condition in which the heart cells enlarge causing the heart wall to thicken, is the leading cause of sudden cardiac death among young adults and athletes, which indicates that physical stress increases the risk of developing heart failure in the patients already at risk due to genetic factors. Therefore, to develop useful "HCM-in-a dish" model systems, it is necessary to precisely control the environmental stress exerted on hiPS-derived cardiac tissues. This project focuses on the MYBPC3 gene, which is one of the most frequently mutated HCM genes. Though the relationship has been established, the mechanisms by which MYBPC3 mutations lead to HCM are not known. Thus, the primary goal of this project is to investigate the correlation between HCM characteristics and reduced MYBPC3 expression, and how this could be influenced by the increase of environmental stress to the cardiac tissues. Key to the success of this effort is creating a functional/beating 3D cardiac tissue model of HCM, which offers better understanding of how the genetic defects combine with the cellular and tissue environment to initiate and advance the disease. More broadly, the strategies developed in this project could be applied to studying other cardiac diseases and potentially lead to new therapies for disease management and treatment. This new approach (which requires hiPSC technology, cardiac tissue engineering, advanced 3D bioprinting, and materials processing and characterization) will provide significant and presently unavailable opportunities for high school, undergraduate and graduate students to have exciting research experiences and state-of-the-art training in biomedical engineering and nanotechnology. This will be accomplished with coordinated, structured instruction and assessments in the form of coursework, seminars, and workshops, as well as with participation in the research laboratory environment.

The primary goal of this project is to establish an isogenic, human induced pluripotent stem cell (hiPSs) based tissue model of hypertrophic cardiomyopathy (HCM), for studying how genetic defects interplay with the cellular and tissue environment to initiate and progress the disease. HCM is the leading cause of sudden cardiac death among young adults and athletes, which indicates that physical stress increases the risk of developing heart failure in patients with HCM-related genetic predispositions. This project focuses on the MYBPC3 gene, one of the most frequent mutated HCM genes, though molecular mechanisms by which MYBPC3 mutations lead to HCM remain elusive. The central hypothesis of this project is that the severity of HCM phenotype would be dose-dependent on the reduction of MYBPC3 gene expression and protein content (haploinsufficiency), which could be exacerbated by the increase of environmental stress to cardiac microtissues derived from hiPSCs (hiPS-microCTs). The microtissue model will be established by integrating: 1) hiPSC technology for understanding human-specific HCM disease mechanisms associated with MYBPC3 mutations, 2) laser-based bioprinting method for the creation of three-dimensional (3D) hiPS-microCTs on the filamentous matrices with controllable biomechanical stress, and 3) gene-editing approach for the generation of MYBPC3 loss-of-function mutations with identical genetic background (isogenic) as wild type (WT) and dose-dependent reduction of MYBPC3 gene expression. The research plan is organized under three objectives: 1) To correlate the biomechanical stress presented to the MYBPC3 deficient isogenic hiPS-microCTs with the HCM disease severity based on the primary phenotypic metrics; 2) To correlate the haploinsufficiency level in the MYBPC3 deficient isogenic hiPS-microCTs with the HCM disease severity under different biomechanical stress; and 3) To elucidate the molecular mechanisms involved in the stress-induced disease progression of MYBPC3-associated HCM. The combination of hiPSC technology, 3D bioprinting, gene editing method and tissue engineering approaches provides great potential in the development of next generation hiPSC-based disease-specific in vitro preclinical tissue models. This model will be a significant advancement for investigating genotype-phenotype correlation associated with the clinical heterogeneity, elucidating the disease progression in human cardiomyopathies, and developing new therapeutic strategies for disease management and treatment.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

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