Human induced pluripotent stem cell-derived cardiomyocytes and their use in a cardiac organ-on-a-chip to assay electrophysiology, calcium and contractility.

Abstract

Cardiac organs-on-a-chip (OoCs) or microphysiological systems have the potential to predict cardiac effects of new drug candidates, including unanticipated cardiac outcomes, which are among the main causes for drug attrition. This protocol describes how to prepare and use a cardiac OoC containing cardiomyocytes differentiated from human induced pluripotent stem cells (hiPS cells). The use of cells derived from hiPS cells as reliable sources of human cells from diverse genetic backgrounds also holds great potential, especially when cultured in OoCs that are physiologically relevant culture platforms. To promote the broad adoption of hiPS cell-derived cardiac OoCs in the drug development field, there is a need to first ensure reproducibility in their preparation and use. This protocol aims to provide key information on how to reduce sources of variability during hiPS cell maintenance, differentiation, loading and maturation in OoCs. Variability in these procedures can lead to inconsistent purity after differentiation and variable function between batches of microtissues formed in OoCs. This protocol also focuses on describing the handling and functional assessment of cardiac microtissues using live-cell microscopy approaches to quantify parameters of cellular electrophysiology, calcium transients and contractility. The protocol consists of five stages: (1) thaw and maintain hiPS cells, (2) differentiate hiPS cell cardiomyocytes, (3) load differentiated cells into OoCs, (4) maintain and characterize loaded cells, and (5) evaluate and utilize cardiac OoCs. Execution of the entire protocol takes ~40 days. The required skills to carry out the protocol are experience with sterile techniques, mammalian cell culture and maintaining hiPS cells in a pluripotent state.