Conclusion
We found MYBPC3 +/- mutations cause iPSC-μHM to exhibit hypercontractility, and also a lower tolerance for mechanical stiffness. Understanding how genetics work in combination with mechanical stiffness to trigger and/or exacerbate pathophysiology may lead to more effective therapies for HCM.
Methods
We differentiated isogenic control (WTC) and MYBPC3 +/- iPSC into cardiomyocytes using small molecule manipulation of Wnt signaling, and then purified them using lactate media. The purified cardiomyocytes were seeded into "dog bone" shaped stencil molds to form micro-heart muscle arrays (μHM). To mimic changes in myocardial stiffness stemming from pressure overload, we varied the rigidity of the substrates μHM contract against. Stiffness levels ranged from those corresponding to fetal (5 kPa), healthy (15 kPa), pre-fibrotic (30 kPa) to fibrotic (65 kPa) myocardium. Substrates were embedded with a thin layer of fluorescent beads to track contractile force, and parent iPSC were engineered to express the genetic calcium indicator, GCaMP6f. High speed video microscopy and image analysis were used to quantify calcium handling and contractility of μHM.
Results
Substrate rigidity triggered physiological adaptation for both genotypes. However, MYBPC3 +/- μHM showed a lower tolerance to substrate stiffness with the peak traction on 15 kPa, while WTC μHM had peak traction on 30 kPa. MYBPC3 +/- μHM exhibited hypercontractility, which was exaggerated by substrate rigidity. MYBPC3 +/- μHM hypercontractility was associated with longer rise times for calcium uptake and force development, along with higher overall Ca2+ intake.
Supplementary Information
The online version contains supplementary material available at (10.1007/s12195-021-00684-x).