Thyroid and Glucocorticoid Hormones Promote Functional T-tubule Development in Human-Induced Pluripotent Stem Cell Derived Cardiomyocytes
Rationale: Human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CM) are increasingly being used for modeling heart disease and are under development for regeneration of the injured heart. However, incomplete structural and functional maturation of hiPSC-CM including lack of t-tubules, immature excitation-contraction (EC) coupling, and inefficient Ca-induced Ca release (CICR) remain major limitations.
Objective: Thyroid and glucocorticoid hormones are critical for heart maturation. We hypothesized that their addition to standard protocols would promote t-tubule development and mature EC coupling of hiPSC-CM when cultured on extracellular matrix with physiological stiffness (Matrigel mattress).
Methods and Results: HiPSC-CM were generated using a standard chemical differentiation method supplemented with triiodo-L-thyronine (T3) and/or dexamethasone (Dex) during days 16-30 followed by single-cell culture for 5 days on Matrigel mattress. HiPSC-CM treated with T3+Dex, but not with either T3 or Dex alone, developed an extensive t-tubule network. Notably, Matrigel mattress was necessary for t-tubule formation. Compared to adult human ventricular CM, t-tubules in T3+Dex-treated hiPSC-CM were less organized and had more longitudinal elements. Confocal line scans demonstrated spatially and temporally uniform Ca release that is characteristic of EC coupling in the heart ventricle. T3+Dex enhanced elementary Ca release measured by Ca sparks as well as promoted ryanodine receptor (RyR2) structural organization. Simultaneous measurements of L-type Ca current and intracellular Ca release confirmed enhanced functional coupling between L-type Ca channels and RyR2 in T3+Dex cells.
Conclusions: Our results suggest a permissive role of combined thyroid and glucocorticoid hormones during the cardiac differentiation process which, when coupled with further maturation on Matrigel mattress, is sufficient for t-tubule development, enhanced CICR, and more ventricular-like EC coupling. This new hormone maturation method could advance the utility of hiPSC-CM for disease modeling and cell-based therapy.
- induced pluripotent-derived stem cells
- calcium induced calcium release
- Matrigel mattress
- excitation-contraction coupling
- Received August 29, 2017.
- Revision received September 26, 2017.
- Accepted October 2, 2017.