Optogenetic Control of Human Induced Pluripotent Stem Cell-Derived Cardiac Tissue Models.

Background Optogenetics, using light-sensitive proteins, emerged as a unique experimental paradigm to modulate cardiac excitability. We aimed to develop high-resolution optogenetic approaches to modulate electrical activity in 2- and 3-dimensional cardiac tissue models derived from human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes. Methods and Results To establish light-controllable cardiac tissue models, opsin-carrying HEK293 cells, expressing the light-sensitive cationic-channel CoChR, were mixed with hiPSC-cardiomyocytes to generate 2-dimensional hiPSC-derived cardiac cell-sheets or 3-dimensional engineered heart tissues. Complex illumination patterns were designed with a high-resolution digital micro-mirror device. Optical mapping and force measurements were used to evaluate the tissues' electromechanical properties. The ability to optogenetically pace and shape the tissue's conduction properties was demonstrated by using single or multiple illumination stimulation sites, complex illumination patterns, or diffuse illumination. This allowed to establish in vitro models for optogenetic-based cardiac resynchronization therapy, where the electrical activation could be synchronized (hiPSC-derived cardiac cell-sheets and engineered heart tissue models) and contractile properties improved (engineered heart tissues). Next, reentrant activity (rotors) was induced in the hiPSC-derived cardiac cell-sheets and engineered heart tissue models through optogenetics programmed- or cross-field stimulations. Diffuse illumination protocols were then used to terminate arrhythmias, demonstrating the potential to study optogenetics cardioversion mechanisms and to identify optimal illumination parameters for arrhythmia termination. Conclusions By combining optogenetics and hiPSC technologies, light-controllable human cardiac tissue models could be established, in which tissue excitability can be modulated in a functional, reversible, and localized manner. This approach may bring a unique value for physiological/pathophysiological studies, for disease modeling, and for developing optogenetic-based cardiac pacing, resynchronization, and defibrillation approaches.

Using Decellularization/Recellularization Processes to Prepare Liver and Cardiac Engineered Tissues.

Tissue engineering provides unique opportunities for disease modeling, drug testing, and regenerative medicine applications. The use of cell-seeded scaffolds to promote tissue development is the hallmark of the tissue engineering. Among the different types of scaffolds (derived from either natural or synthetic polymers) used in the field, the use of decellularized tissues/organs is specifically attractive. The decellularization process involves the removal of native cells from the original tissue, allowing for the preservation of the three-dimensional (3D) macroscopic and microscopic structures of the tissue and extracellular matrix (ECM) composition. Following recellularization, the resulting scaffold provides the seeded cells with the appropriate biological signals and mechanical properties of the original tissue. Here, we describe different methods to create viable scaffolds from decellularized heart and liver as useful tools to study and exploit ECM biological key factors for the generation of engineered tissues with enhanced regenerative properties.

Human Induced Pluripotent Stem Cell-Derived Cardiac Cell Sheets Expressing Genetically Encoded Voltage Indicator for Pharmacological and Arrhythmia Studies.

Fulfilling the potential of human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes for studying conduction and arrhythmogenesis requires development of multicellular models and methods for long-term repeated tissue phenotyping. We generated confluent hiPSC-derived cardiac cell sheets (hiPSC-CCSs), expressing the genetically encoded voltage indicator ArcLight. ArcLight-based optical mapping allowed generation of activation and action-potential duration (APD) maps, which were validated by mapping the same hiPSC-CCSs with the voltage-sensitive dye, Di-4-ANBDQBS. ArcLight mapping allowed long-term assessment of electrical remodeling in the hiPSC-CCSs and evaluation of drug-induced conduction slowing (carbenoxolone, lidocaine, and quinidine) and APD prolongation (quinidine and dofetilide). The latter studies also enabled step-by-step depiction of drug-induced arrhythmogenesis ("torsades de pointes in the culture dish") and its prevention by MgSO4 and rapid pacing. Phase-mapping analysis allowed biophysical characterization of spiral waves induced in the hiPSC-CCSs and their termination by electrical cardioversion and overdrive pacing. In conclusion, ArcLight mapping of hiPSC-CCSs provides a powerful tool for drug testing and arrhythmia investigation.