Matrigel Mattress

Abstract

Rationale The lack of measurable single cell contractility of human induced pluripotent stem cell-derived cardiac myocytes (hiPSC-CMs) currently limits the utility of hiPSC-CMs for evaluating contractile performance for both basic research and drug discovery. Objective To develop a culture method that rapidly generates contracting single hiPSC-CMs and allows quantification of cell shortening with standard equipment used for studying adult cardiac myocytes (CMs). Methods and Results Single hiPSC-CMs were cultured for 5 – 7 days on a 0.4 – 0.8 mm thick mattress of undiluted Matrigel (“mattress hiPSC-CM”) and compared to hiPSC-CMs maintained on control substrate (<0.1 mm thick 1:60 diluted matrigel, “control hiPSC-CM”). Compared to control hiPSC-CM, mattress hiPSC-CMs had more rod-shape morphology and significantly increased sarcomere length. Contractile parameters of mattress hiPSC-CMs measured with video-based edge detection was comparable to that of freshly isolated adult rabbit ventricular CMs. Morphological and contractile properties of mattress hiPSC-CM were consistent across cryopreserved hiPSC-CMs generated independently at another institution. Unlike control hiPSC-CM, mattress hiPSC-CMs display robust contractile responses to positive inotropic agents such as myofilament calcium sensitizers. Mattress hiPSC-CMs exhibit molecular changes that include increased expression of the maturation marker cardiac troponin I and significantly increased action potential upstroke velocity due to a 2-fold increase in sodium current (INa). Conclusions The Matrigel mattress method enables the rapid generation of robustly contracting hiPSC-CMs and enhances maturation. This new method allows quantification of contractile performance at the single cell level, which should be valuable to disease modeling, drug discovery and preclinical cardiotoxicity testing.

Myofilament Calcium-Buffering Dependent Action Potential Triangulation in Human-Induced Pluripotent Stem Cell Model of Hypertrophic Cardiomyopathy

“…The other two lines were named population controls. We then used the matrigel mattress method (3) to generate single rod-shaped CMs for each iPSC line and studied TnT-I79N protein levels, cytosolic Ca buffering and electrophysiology.…”

A Recipe for T-Tubules in Human iPS Cell–Derived Cardiomyocytes

“…6 In addition to soluble signaling molecules, the extracellular matrix composition and associated substrate stiffness have been found to be important variables in promoting maturation. 7, 8 However, these previous studies and multiple other related studies have not observed formation of t-tubules in hPSC-CMs or they have not looked specifically for this feature. Two recent studies did provide evidence that strategies for maturation could promote some t-tubule formation using either prolonged culture on nanopatterned surfaces or engineered substrates of optimized shape and stiffness.…”

“…11 The authors found that combining the Knollman lab’s previously published Matrigel mattress technique 7 with T3 and Dex resulted in hPSC-CMs exhibiting abundant T-tubules with largely synchronized Ca 2+ release throughout the myocytes similar to adult cardiomyocytes. This contrasted the untreated hPSC-CM that exhibited a delayed Ca 2+ release present in the center of the cells.…”

Circulation Research “In This Issue” Anthology

“…They discovered that, unlike control animals, sympathectomized mice showed no apical regeneration at all, but only scarring, thus identifying that reinnervation is essential for cardiac regeneration in newborn mice. 62 Feaster et al have figured out an easy way to get human iPSC-derived cardiomyocytes into better shape for contraction.…”