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Human Engineered Heart Tissue: Analysis of Contractile Force.

ABSTRACT: Analyzing contractile force, the most important and best understood function of cardiomyocytes in vivo is not established in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM). This study describes the generation of 3D, strip-format, force-generating engineered heart tissues (EHT) from hiPSC-CM and their physiological and pharmacological properties. CM were differentiated from hiPSC by a growth factor-based three-stage protocol. EHTs were generated and analyzed histologically and functionally. HiPSC-CM in EHTs showed well-developed sarcomeric organization and alignment, and frequent mitochondria. Systematic contractility analysis (26 concentration-response curves) reveals that EHTs replicated canonical response to physiological and pharmacological regulators of inotropy, membrane- and calcium-clock mediators of pacemaking, modulators of ion-channel currents, and proarrhythmic compounds with unprecedented precision. The analysis demonstrates a high degree of similarity between hiPSC-CM in EHT format and native human heart tissue, indicating that human EHTs are useful for preclinical drug testing and disease modeling.

SUBMITTER: Mannhardt I

PROVIDER: S-EPMC4944531 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Human Engineered Heart Tissue: Analysis of Contractile Force.

Mannhardt Ingra I Breckwoldt Kaja K Letuffe-Brenière David D Schaaf Sebastian S Schulz Herbert H Neuber Christiane C Benzin Anika A Werner Tessa T Eder Alexandra A Schulze Thomas T Klampe Birgit B Christ Torsten T Hirt Marc N MN Huebner Norbert N Moretti Alessandra A Eschenhagen Thomas T Hansen Arne A

Stem cell reports 20160519 1

Analyzing contractile force, the most important and best understood function of cardiomyocytes in vivo is not established in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM). This study describes the generation of 3D, strip-format, force-generating engineered heart tissues (EHT) from hiPSC-CM and their physiological and pharmacological properties. CM were differentiated from hiPSC by a growth factor-based three-stage protocol. EHTs were generated and analyzed histologically ...[more]

PMID: 27211213

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Project description:BackgroundOne third of heart failure patients exhibit dyssynchronized electromechanical activity of the heart (evidenced by a broad QRS-complex). Cardiac resynchronization therapy (CRT) in the form of biventricular pacing improves cardiac output and clinical outcome of responding patients. Technically demanding and laborious large animal models have been developed to better predict responders of CRT and to investigate molecular mechanisms of dyssynchrony and CRT. The aim of this study was to establish a first humanized in vitro model of dyssynchrony and CRT.MethodsCardiomyocytes were differentiated from human induced pluripotent stem cells and cast into a fibrin matrix to produce engineered heart tissue (EHT). EHTs were either field stimulated in their entirety (symmetrically) or excited locally from one end (asymmetrically) or they were allowed to beat spontaneously.ResultsAsymmetrical pacing led to a depolarization wave from one end to the other end, which was visualized in human EHT transduced with a fast genetic Ca2+-sensor (GCaMP6f) arguing for dyssynchronous excitation. Symmetrical pacing in contrast led to an instantaneous (synchronized) Ca2+-signal throughout the EHT. To investigate acute and long-term functional effects, spontaneously beating human EHTs (0.5-0.8 Hz) were divided into a non-paced control group, a symmetrically and an asymmetrically paced group, each stimulated at 1 Hz. Symmetrical pacing was clearly superior to asymmetrical pacing or no pacing regarding contractile force both acutely and even more pronounced after weeks of continuous stimulation. Contractile dysfunction that can be evoked by an increased afterload was aggravated in the asymmetrically paced group. Consistent with reports from paced dogs, p38MAPK and CaMKII-abundance was higher under asymmetrical than under symmetrical pacing while pAKT was considerably lower.ConclusionsThis model allows for long-term pacing experiments mimicking electrical dyssynchrony vs. synchrony in vitro. Combined with force measurement and afterload stimulus manipulation, it provides a robust new tool to gain insight into the biology of dyssynchrony and CRT.

Project description:Engineered heart tissues (EHTs) have emerged as a robust in vitro model to study cardiac physiology. Although biomimetic culture environments have been developed to better approximate in vivo conditions, currently available methods do not permit full recapitulation of the four phases of the cardiac cycle. We have developed a bioreactor which allows EHTs to undergo cyclic loading sequences that mimic in vivo work loops. EHTs cultured under these working conditions exhibited enhanced concentric contractions but similar isometric contractions compared with EHTs cultured isometrically. EHTs that were allowed to shorten cyclically in culture had increased capacity for contractile work when tested acutely. Increased work production was correlated with higher levels of mitochondrial proteins and mitochondrial biogenesis; this effect was eliminated when tissues were cyclically shortened in the presence of a myosin ATPase inhibitor. Leveraging our novel in vitro method to precisely apply mechanical loads in culture, we grew EHTs under two loading regimes prescribing the same work output but with different associated afterloads. These groups showed no difference in mitochondrial protein expression. In loading regimes with the same afterload but different work output, tissues subjected to higher work demand exhibited elevated levels of mitochondrial protein. Our findings suggest that regulation of mitochondrial mass in cultured human EHTs is potently modulated by the mechanical work the tissue is permitted to perform in culture, presumably communicated through ATP demand. Precise application of mechanical loads to engineered heart tissues in culture represents a novel in vitro method for studying physiological and pathological cardiac adaptation.NEW & NOTEWORTHY In this work, we present a novel bioreactor that allows for active length control of engineered heart tissues during extended tissue culture. Specific length transients were designed so that engineered heart tissues generated complete cardiac work loops. Chronic culture with various work loops suggests that mitochondrial mass and biogenesis are directly regulated by work output.

Dataset Information

Human Engineered Heart Tissue: Analysis of Contractile Force.

Publications

Human Engineered Heart Tissue: Analysis of Contractile Force.

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