Rationale
Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) are invaluable for patient-specific disease modeling and drug testing¹. However, modeling hypoxia-induced injury using iPSC-CMs remains challenging due to their immature phenotype and distinct metabolic profile compared to adult CMs2,3.
Objective
We aimed to enhance iPSC-CM maturation by combining lipid-supplemented maturation medium (MM), nanopatterned surface alignment (NP), and electrostimulation (ES) to establish a robust human hypoxia-injury model.
Methods
We systematically characterized the influence of MM, NP and ES, in comparison to B27 medium. Mitochondrial maturation was evaluated via Tom20 immunostaining and RNA-sequencing. Cell viability after hypoxia was measured using live/dead staining and lactate-dehydrogenase (LDH) activity in medium supernatant.
Results
Transcriptome analysis revealed a gradual metabolic maturation of iPSC-CMs through combination of MM with NP, and MM with NP and ES (MM+NP+ES) based on the upregulation of genes involved in the electron transport chain and mitochondrial biogenesis1. Those genes were enriched in TFAM (mitochondrial transcription factor A)- and HMCES (5-hydroxymethylcytosine binding, embryonic stem cell-specific)-related target sets, consistent with their important roles in mitochondrial DNA transcription and integrity. Tom20 staining revealed an increased mitochondrial mass in iPSC-CMs cultured under MM+NP+ES or MM+ES versus MM, identifying ES as the key driver for mitochondrial development. Cells matured under MM+ES showed substantial cell death after short-term hypoxia, while few dead cells were observed for iPSC-CMs in MM even after prolonged hypoxia (24 h).
Conclusion
Our findings demonstrate the crucial role of metabolic maturation and mitochondrial development for the generation of and iPSC-CM-based human model for myocardial infarction.