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Subtype Separation of iPSC-derived Cardiomyocytes May Foster Safer Therapies

Review of “An Intrinsic, Label-Free Signal for Identifying Stem Cell Derived Cardiomyocyte Subtype” from STEM CELLS by Stuart P. Atkinson

The purification of cardiomyocyte subtypes from differentiating cultures of induced pluripotent stem cells (iPSC) [1, 2] aims to provide safety and efficacy with regards to stem cell-based heart disease therapies and authenticity to cardiac cell function and disease models and drug cardiotoxicity assessments [3]. To this end, a previous study from the laboratory of James W. Chan (University of California, Davis, USA), established that the detection of second-harmonic generation signals emanating from cardiomyocyte-specific myosin rod bundles represents an efficient label‐free optical means to identify hiPSC-derived cardiomyocytes and define their maturity [4]. 

Now, the team returns with a new STEM CELLS study in which they report the separation of hiPSC-derived cardiomyocyte subtypes based on their specific second-harmonic generation intensities due to their differing contents of myosin filaments [5]. Overall, this study describes a novel strategy that may provide an effective means to isolate pure populations of cardiomyocytes from differentiating cultures of iPSCs. 

Chang et al. first determined cardiomyocyte subtype employing action potential profiling, discovering a population comprising 10% pacemaker‐like cardiomyocytes and approximately 45% for each of contractile ventricular‐like and atrial‐like cardiomyocytes. Analysis of second-harmonic generation intensities through confocal microscopy of the three subtypes found that the contractile cardiomyocytes exhibited a higher second-harmonic generation intensity when compared to the non-contractile pacemaker‐like cells (approximately 70% lower signal strength) thanks to the presence of relatively periodic, organized myosin patterns.

Therefore, the authors suggest that second-harmonic generation may be employed to “purge” any potential arrhythmia‐inducing pacemaker‐like cardiomyocytes and other non-cardiomyocyte cell types from a population of hiPSC-derived cardiomyocytes to generate a highly pure population of contractile cells. However, the study takes care to note that a proportion of contractile cardiomyocytes would be lost when following this strategy, leading to the generation of a population of cells representing 30% of the original cell population.

Overall, the authors provide evidence that the evaluation of second-harmonic generation signals may find use as a label‐free optical marker to remove any unwanted and potentially dangerous cells from a heterogeneous population of cells derived from the cardiomyocytic differentiation of iPSCs. The authors next hope to integrate second-harmonic generation analysis into high‐throughput microfluidic sorting for the analysis of individual hiPSC-derived cardiomyocytes and the creation of pure cardiomyocyte populations for disease modeling, drug screening, and stem cell-based therapies.

For more on stem cell-based cardiac therapies and advanced cell sorting technologies, stay tuned to the Stem Cells Portal!


  1. Burridge Paul W, Keller G, Gold Joseph D, et al., Production of De Novo Cardiomyocytes: Human Pluripotent Stem Cell Differentiation and Direct Reprogramming. Cell Stem Cell 2012;10:16-28.
  2. Yechikov S, Copaciu R, Gluck JM, et al., Same-Single-Cell Analysis of Pacemaker-Specific Markers in Human Induced Pluripotent Stem Cell-Derived Cardiomyocyte Subtypes Classified by Electrophysiology. STEM CELLS 2016;34:2670-2680.
  3. Oikonomopoulos A, Kitani T, and Wu JC, Pluripotent Stem Cell-Derived Cardiomyocytes as a Platform for Cell Therapy Applications: Progress and Hurdles for Clinical Translation. Molecular Therapy 2018;26:1624-1634.
  4. Awasthi S, Matthews DL, Li RA, et al., Label-free identification and characterization of human pluripotent stem cell-derived cardiomyocytes using second harmonic generation (SHG) microscopy. Journal of Biophotonics 2012;5:57-66.
  5. Chang C-W, Kao HKJ, Yechikov S, et al., An intrinsic, label-free signal for identifying stem cell-derived cardiomyocyte subtype. STEM CELLS 2020;38:390-394.