Biowire: a platform for maturation of human pluripotent stem cell–derived cardiomyocytes

Abstract

Directed differentiation protocols enable derivation of cardiomyocytes from human pluripotent stem cells (hPSC) and permit engineering of human myocardium in vitro. However, hPSC-derived cardiomyocytes are reflective of very early human development, limiting their utility in the generation of in vitro models of mature myocardium. Here, we developed a new platform that combines three-dimensional cell cultivation in a microfabricated system with electrical stimulation to mature hPSC-derived cardiac tissues. We utilized quantitative structural, molecular and electrophysiological analyses to elucidate the responses of immature human myocardium to electrical stimulation and pacing. We demonstrated that the engineered platform allowed for the generation of 3-dimensional, aligned cardiac tissues (biowires) with frequent striations. Biowires submitted to electrical stimulation markedly increased myofibril ultrastructural organization, displayed elevated conduction velocity and altered both the electrophysiological and Ca2+ handling properties versus non-stimulated controls. These changes were in agreement with cardiomyocyte maturation and were dependent on the stimulation rate.

Integrating cardiomyocytes from human pluripotent stem cells in safety pharmacology: has the time come?

“…Defined culture conditions (Burridge et al , 2014; Ribeiro et al , 2015a, 2015b) can be used to improve the maturation of hPSC‐CMs to reveal hidden disease phenotypes (Birket et al , 2015) or to drive differentiation to chamber‐specific cell populations (Devalla et al , 2015), and drug testing might benefit from their standardization. Exogenous stimuli, such as adjusted pacing frequency (Chan et al , 2013), 3D‐microenvironments (Zhao et al , 1999; Nunes et al , 2013; Hirt et al , 2014; Eder et al , 2016) and heterotypic cell co‐culture (Robertson et al , 2013) may all contribute to cell maturation, increasing the expression of ion channels and improving functional properties. Recent advances in electrophysiology (Meijer van Putten et al , 2015) and cell biology (Vaidyanathan et al , 2016) allow the introduction of exogenous I K1 conductance in hiPSC‐CMs, although neither single nor combined approaches were successful in mimicking adult maturation state entirely, in particular of Ca 2 + ‐handling, signalling and compartmentalization, mainly due to their lack of T‐tubules (Kane and Terracciano, 2015).…”

Inherited heart disease – what can we expect from the second decade of human iPS cell research?

“…Ideally 3D tissue structures will be developed, where hiPSC‐CMs (subtypes) and other cells are mixed together and organised in microtissues, with or without the addition of extracellular scaffolds, and will be subjected to electrical or mechanical stimulation 77. However, it is still unclear to what extent adult CM properties can be acquired in a culture dish.…”

Human pluripotent stem cell-derived cardiomyocyte based models for cardiotoxicity and drug discovery

“…Specifically, in terms of automaticity, gap junctional protein expression, contractile apparatus, electrophysiology, and Ca 2+ handling properties, hPSC-CMs are more similar to fetal CMs than adult CMs [14]. While the optimal degree of maturation for assessing cardiac liability has not yet been established, it is generally believed that maturation would benefit these assessments because (i) the target patient population for safety screens is the adult human and (ii) maturation results in changes at the gene and protein level, through to the higher levels of structure and function [12].…”

“…The post platform both induces alignment within the 3D ECT (Figure 1(f)) due to the static stretch applied by the posts and provides a means of directly measuring the contractility of the ECT in situ. The Biowire ™ (TARA Biosystems, Inc., New York, USA) [14] ECT (Figure 1(g,h)), wherein the cardiac cell suspension self-assembled around a suture, also showed improved cell alignment (Figure 1(i)), and with improved alignment, improved cardiac phenotype compared with age-matched embryoid bodies. Electrical stimulation further enhanced the maturation effects of the Biowire culture.…”

“…Generating 3D micro ECTs of variable cell compositions, using human-derived cardiac cells, has been demonstrated as discussed. Moreover, varying degrees of hPSC-CM maturation have been demonstrated by means of prolonged culture [26], topographical cues [14], electrical stimulation [14], mechanical stretching [19], and/or chemical manipulation [27], as well as coculture with cardiac fibroblasts [28] and endothelial cells [25]. Yet, there are currently no robust methods of promoting the functional maturation of hPSC-CMs within a short time frame.…”