Spatiotemporal imaging documented the maturation of the cardiomyocytes from human induced pluripotent stem cells.

OBJECTIVES

Cardiomyocytes derived from human induced pluripotent stem cells are a promising source of cells for regenerative medicine. However, contractions in such derived cardiomyocytes are often irregular and asynchronous, especially at early stages of differentiation. This study aimed to determine the differentiation stage of initiation of synchronized and regular contractions, using spatiotemporal imaging and physiological and genetic analyses.

METHODS

Knock-in human induced pluripotent stem cell lines were established with clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats-associated protein 9 to analyze cardiac and pacemaker cell maturation. Time-frequency analysis and Ca imaging were performed, and the expression of related proteins and specific cardiac/pacemaker mRNAs in contracting embryoid bodies was analyzed at various differentiation stages.

RESULTS

Time-frequency analysis and Ca imaging revealed irregular, asynchronous contractions at the early stage of differentiation with altered electrophysiological properties upon differentiation. Genes associated with electrophysiological properties were upregulated after 70 days of culturing in differentiation media, whereas pacemaker genes were initially upregulated during the early stage and downregulated at the later stage.

CONCLUSIONS

A differentiation period >70 days is required for adequate development of cardiac elements including ion channels and gap junctions and for sarcomere maturation.