Comparisons of 3D printed materials for biomedical imaging applications.

In biomedical imaging, it is desirable that custom-made accessories for restraint, anesthesia, and monitoring can be easily cleaned and not interfere with the imaging quality or analyses. With the rise of 3D printing as a form of rapid prototyping or manufacturing for imaging tools and accessories, it is important to understand which printable materials are durable and not likely to interfere with imaging applications. Here, 15 3D printable materials were evaluated for radiodensity, optical properties, simulated wear, and capacity for repeated cleaning and disinfection. Materials that were durable, easily cleaned, and not expected to interfere with CT, PET, or optical imaging applications were identified.

A guide for selecting 3D printed materials for custom research tools through characterization of their merits and limitations in biomedical imaging.

ER stress and lipid imbalance drive diabetic embryonic cardiomyopathy in an organoid model of human heart development.

Congenital heart defects are the most prevalent human birth defects, and their incidence is exacerbated by maternal health conditions, such as diabetes during the first trimester (pregestational diabetes). Our understanding of the pathology of these disorders is hindered by a lack of human models and the inaccessibility of embryonic tissue. Using an advanced human heart organoid system, we simulated embryonic heart development under pregestational diabetes-like conditions. These organoids developed pathophysiological features observed in mouse and human studies before, including ROS-mediated stress and cardiomyocyte hypertrophy. scRNA-seq revealed cardiac cell-type-specific dysfunction affecting epicardial and cardiomyocyte populations and alterations in the endoplasmic reticulum and very-long-chain fatty acid lipid metabolism. Imaging and lipidomics confirmed these findings and showed that dyslipidemia was linked to fatty acid desaturase 2 mRNA decay dependent on IRE1-RIDD signaling. Targeting IRE1 or restoring lipid levels partially reversed the effects of pregestational diabetes, offering potential preventive and therapeutic strategies in humans.

ER stress and lipid imbalance drive embryonic cardiomyopathy in a human heart organoid model of pregestational diabetes.

Congenital heart defects constitute the most common birth defect in humans, affecting approximately 1% of all live births. The incidence of congenital heart defects is exacerbated by maternal conditions, such as diabetes during the first trimester. Our ability to mechanistically understand these disorders is severely limited by the lack of human models and the inaccessibility to human tissue at relevant stages. Here, we used an advanced human heart organoid model that recapitulates complex aspects of heart development during the first trimester to model the effects of pregestational diabetes in the human embryonic heart. We observed that heart organoids in diabetic conditions develop pathophysiological hallmarks like those previously reported in mouse and human studies, including ROS-mediated stress and cardiomyocyte hypertrophy, among others. Single cell RNA-seq revealed cardiac cell type specific-dysfunction affecting epicardial and cardiomyocyte populations, and suggested alterations in endoplasmic reticulum function and very long chain fatty acid lipid metabolism. Confocal imaging and LC-MS lipidomics confirmed our observations and showed that dyslipidemia was mediated by fatty acid desaturase 2 (FADS2) mRNA decay dependent on IRE1-RIDD signaling. We also found that the effects of pregestational diabetes could be reversed to a significant extent using drug interventions targeting either IRE1 or restoring healthy lipid levels within organoids, opening the door to new preventative and therapeutic strategies in humans.

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

The ability to study human cardiac development in health and disease is highly limited by the capacity to model the complexity of the human heart in vitro. Developing more efficient organ-like platforms that can model complex in vivo phenotypes, such as organoids and organs-on-a-chip, will enhance the ability to study human heart development and disease. This paper describes a protocol to generate highly complex human heart organoids (hHOs) by self-organization using human pluripotent stem cells and stepwise developmental pathway activation using small molecule inhibitors. Embryoid bodies (EBs) are generated in a 96-well plate with round-bottom, ultra-low attachment wells, facilitating suspension culture of individualized constructs. The EBs undergo differentiation into hHOs by a three-step Wnt signaling modulation strategy, which involves an initial Wnt pathway activation to induce cardiac mesoderm fate, a second step of Wnt inhibition to create definitive cardiac lineages, and a third Wnt activation step to induce proepicardial organ tissues. These steps, carried out in a 96-well format, are highly efficient, reproducible, and produce large amounts of organoids per run. Analysis by immunofluorescence imaging from day 3 to day 11 of differentiation reveals first and second heart field specifications and highly complex tissues inside hHOs at day 15, including myocardial tissue with regions of atrial and ventricular cardiomyocytes, as well as internal chambers lined with endocardial tissue. The organoids also exhibit an intricate vascular network throughout the structure and an external lining of epicardial tissue. From a functional standpoint, hHOs beat robustly and present normal calcium activity as determined by Fluo-4 live imaging. Overall, this protocol constitutes a solid platform for in vitro studies in human organ-like cardiac tissues.

Self-assembling human heart organoids for the modeling of cardiac development and congenital heart disease.

Congenital heart defects constitute the most common human birth defect, however understanding of how these disorders originate is limited by our ability to model the human heart accurately in vitro. Here we report a method to generate developmentally relevant human heart organoids by self-assembly using human pluripotent stem cells. Our procedure is fully defined, efficient, reproducible, and compatible with high-content approaches. Organoids are generated through a three-step Wnt signaling modulation strategy using chemical inhibitors and growth factors. Heart organoids are comparable to age-matched human fetal cardiac tissues at the transcriptomic, structural, and cellular level. They develop sophisticated internal chambers with well-organized multi-lineage cardiac cell types, recapitulate heart field formation and atrioventricular specification, develop a complex vasculature, and exhibit robust functional activity. We also show that our organoid platform can recreate complex metabolic disorders associated with congenital heart defects, as demonstrated by an in vitro model of pregestational diabetes-induced congenital heart defects.