A versatile high-throughput assay based on 3D ring-shaped cardiac tissues generated from human induced pluripotent stem cell-derived cardiomyocytes.

We developed a 96-well plate assay which allows fast, reproducible, and high-throughput generation of 3D cardiac rings around a deformable optically transparent hydrogel (polyethylene glycol [PEG]) pillar of known stiffness. Human induced pluripotent stem cell-derived cardiomyocytes, mixed with normal human adult dermal fibroblasts in an optimized 3:1 ratio, self-organized to form ring-shaped cardiac constructs. Immunostaining showed that the fibroblasts form a basal layer in contact with the glass, stabilizing the muscular fiber above. Tissues started contracting around the pillar at D1 and their fractional shortening increased until D7, reaching a plateau at 25±1%, that was maintained up to 14 days. The average stress, calculated from the compaction of the central pillar during contractions, was 1.4±0.4 mN/mm2. The cardiac constructs recapitulated expected inotropic responses to calcium and various drugs (isoproterenol, verapamil) as well as the arrhythmogenic effects of dofetilide. This versatile high-throughput assay allows multiple in situ mechanical and structural readouts.

hsa-miR-548v controls the viscoelastic properties of human cardiomyocytes and improves their relaxation rates.

The impairment of left ventricular (LV) diastolic function with an inadequate increase in myocardial relaxation velocity directly results in lower LV compliance, increased LV filling pressures, and heart failure symptoms. The development of agents facilitating the relaxation of human cardiomyocytes requires a better understanding of the underlying regulatory mechanisms. We performed a high-content microscopy-based screening in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) using a library of 2,565 human miRNA mimics and measured relaxation kinetics via high-computing analyses of motion movies. We identified hsa-miR-548v, a primate-specific miRNA, as the miRNA producing the largest increase in relaxation velocities. This positive lusitropic effect was reproduced in engineered cardiac tissues generated with healthy and BRAF T599R mutant hiPSC-CMs and was independent of changes in calcium transients. Consistent with improvements in viscoelastic responses to mechanical stretch, RNA-Seq showed that hsa-miR-548v downregulated multiple targets, especially components of the mechanosensing machinery. The exogenous administration of hsa-miR-548v in hiPSC-CMs notably resulted in a significant reduction of ANKRD1/CARP1 expression and localization at the sarcomeric I-band. This study suggests that the sarcomere I-band is a critical control center regulating the ability of cardiomyocytes to relax and is a target for improving relaxation and diastolic dysfunction.

Cardiac Organoids to Model and Heal Heart Failure and Cardiomyopathies.

Cardiac tissue engineering aims at creating contractile structures that can optimally reproduce the features of human cardiac tissue. These constructs are becoming valuable tools to model some of the cardiac functions, to set preclinical platforms for drug testing, or to alternatively be used as therapies for cardiac repair approaches. Most of the recent developments in cardiac tissue engineering have been made possible by important advances regarding the efficient generation of cardiac cells from pluripotent stem cells and the use of novel biomaterials and microfabrication methods. Different combinations of cells, biomaterials, scaffolds, and geometries are however possible, which results in different types of structures with gradual complexities and abilities to mimic the native cardiac tissue. Here, we intend to cover key aspects of tissue engineering applied to cardiology and the consequent development of cardiac organoids. This review presents various facets of the construction of human cardiac 3D constructs, from the choice of the components to their patterning, the final geometry of generated tissues, and the subsequent readouts and applications to model and treat cardiac diseases.