Musclemotion

Sala, Luca; Meer, Berend J. van; Tertoolen, Leon G.J.; Bakkers, Jeroen; Bellin, Milena; Davis, Richard P.; Denning, Chris; Dieben, Michel A.E.; Eschenhagen, Thomas; Giacomelli, Elisa; Grandela, Catarina; Hansen, Arne; Holman, Eduard R.; Jongbloed, Monique R.M.; Kamel, Sarah M.; Koopman, Charlotte D.; Lachaud, Quentin; Mannhardt, Ingra; Mol, Mervyn P.H.; Mosqueira, Diogo; Orlova, Valeria V.; Passier, Robert; Ribeiro, Marcelo C.; Saleem, Umber; Smith, Godfrey L.; Burton, Francis L.; Mummery, Christine L.

doi:10.1161/circresaha.117.312067

Cited by 267 publications

(125 citation statements)

References 35 publications

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“…Videos of beating single cells were acquired for 20 s with a frame rate of 33 fps using Zeiss ELYRA LSM 780 (Zeiss). Cardiac contraction was measured with the Image J Musclemotion plugin [32]. Mean contraction was calculated for each individual cell.…”

Section: Quantitative Evaluation Of the Sarcomere Network And Cardiacmentioning

confidence: 99%

Quantitative Evaluation of the Sarcomere Network of Human hiPSC-Derived Cardiomyocytes Using Single-Molecule Localization Microscopy

Lemcke

Skorska

Lang

et al. 2020

IJMS

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The maturation of iPSC-derived cardiomyocytes is still a critical point for their application in cardiovascular research as well as for their clinical use. Although multiple differentiation protocols have been established, researchers failed to generate fully mature cardiomyocytes in vitro possessing identical phenotype-related and functional properties as their native adult counterparts. Besides electrophysiological and metabolic changes, the establishment of a well structured sarcomere network is important for the development of a mature cardiac phenotype. Here, we present a super resolution-based approach to quantitatively evaluate the structural maturation of iPSC-derived cardiomyocytes. Fluorescence labelling of the α-actinin cytoskeleton and subsequent visualization by photoactivated localization microscopy allows the acquisition of highly resolved images for measuring sarcomere length and z-disc thickness. Our image analysis revealed that iPSC and neonatal cardiomyocyte share high similarity with respect to their sarcomere organization, however, contraction capacity was inferior in iPSC-derived cardiac cells, indicating an early maturation level. Moreover, we demonstrate that this imaging approach can be used as a tool to monitor cardiomyocyte integrity, helping to optimize iPSC differentiation as well as somatic cell direct-reprogramming strategies.

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Section: Quantitative Evaluation Of the Sarcomere Network And Cardiacmentioning

confidence: 99%

Quantitative Evaluation of the Sarcomere Network of Human hiPSC-Derived Cardiomyocytes Using Single-Molecule Localization Microscopy

Lemcke

Skorska

Lang

et al. 2020

IJMS

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“…At days 14, 21, 30, and 50 of cardiac differentiation, we acquired the images of beating hAFSC-iPSC-CMs and hESC-CMs for analysis. We used the automated non-profit software, MUSCLEMOTION, to quantitate the changes of cardiomyocytes in terms of contractility and relaxation [19]. It was found that there were no significant differences in contraction duration, velocity, and amplitude between hAFSC-iPSC-CMs and hESC-CMs ( Figure 2).…”

Section: Contractility Of Hpsc-cmsmentioning

confidence: 99%

“…At days 15, 21, 30, and 50 of cardiac differentiation, we used a camera steadily mounted on an inverted microscope with slow-motion features (120 frames/second) to record a video of the beating hAFSC-iPSC-CMs and hESC-CMs for offline analysis. All the acquired files were analyzed using an automated nonprofit software, MUSCLEMOTION [19]. We used MUSCLEMOTION to read and convert the videos to the uncompressed AVI files, and measure the contractility profiles (i.e., contraction amplitude and velocity) of the hAFSC-iPSC-CMs and hESC-CMs [19].…”

Section: Contractility Measurements Of Hafsc-ipsc-cms and Hesc-cmsmentioning

confidence: 99%

“…All the acquired files were analyzed using an automated nonprofit software, MUSCLEMOTION [19]. We used MUSCLEMOTION to read and convert the videos to the uncompressed AVI files, and measure the contractility profiles (i.e., contraction amplitude and velocity) of the hAFSC-iPSC-CMs and hESC-CMs [19]. The reference point of relative cell relaxation was obtained according to the change of color in each frame.…”

Section: Contractility Measurements Of Hafsc-ipsc-cms and Hesc-cmsmentioning

confidence: 99%

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Efficient Cardiac Differentiation of Human Amniotic Fluid-Derived Stem Cells into Induced Pluripotent Stem Cells and Their Potential Immune Privilege

Fang

Wang

Gao

et al. 2020

IJMS

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Mature mammalian hearts possess very limited regenerative potential. The irreversible cardiomyocyte loss after heart injury can lead to heart failure and death. Pluripotent stem cells (PSCs) can differentiate into cardiomyocytes for cardiac repair, but there are obstacles to their clinical application. Among these obstacles is their potential for post-transplant rejection. Although human amniotic fluid-derived stem cells (hAFSCs) are immune privileged, they cannot induce cardiac differentiation. Thus, we generated hAFSC-derived induced PSCs (hAFSC-iPSCs) and used a Wnt-modulating differentiation protocol for the cardiac differentiation of hAFSC-iPSCs. In vitro studies using flow cytometry, immunofluorescence staining, and patch-clamp electrophysiological study, were performed to identify the characteristics of hAFSC-iPSC-derived cardiomyocytes (hAFSC-iPSC-CMs). We injected hAFSC-iPSC-CMs intramuscularly into rat infarcted hearts to evaluate the therapeutic potential of hAFSC-iPSC-CM transplantation. At day 21 of differentiation, the hAFSC-iPSC-CMs expressed cardiac-specific marker (cardiac troponin T), presented cardiomyocyte-specific electrophysiological properties, and contracted spontaneously. Importantly, these hAFSC-iPSC-CMs demonstrated low major histocompatibility complex (MHC) class I antigen expression and the absence of MHC class II antigens, indicating their low immunogenicity. The intramyocardial transplantation of hAFSC-iPSC-CMs restored cardiac function, partially remuscularized the injured region, and reduced fibrosis in the rat infarcted hearts. Therefore, hAFSC-iPSCs are potential candidates for the repair of infarcted myocardium.

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“…The established technologies to evaluate these functional parameters (e.g., patch clamp electrophysiology, multi-electrode arrays, optical imaging, and traction force microscopy) typically rely on low-to medium-throughput assays using single cells, 3D organoid, and/or engineered heart tissues [5,26]. However, recent developments have enabled measurement of contractility across all these configurations, based on quantification of pixel displacement in high-speed movies using a publicly available software [27]. Remarkably, it is now even possible to measure action potentials, cytosolic calcium flux, and contractility simultaneously [28].…”

Section: Introductionmentioning

confidence: 99%

High-Throughput Phenotyping Toolkit for Characterizing Cellular Models of Hypertrophic Cardiomyopathy In Vitro

Mosqueira

Lis-Slimak

Denning

2019

MPs

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Hypertrophic cardiomyopathy (HCM) is a prevalent and complex cardiovascular disease characterised by multifarious hallmarks, a heterogeneous set of clinical manifestations, and several molecular mechanisms. Various disease models have been developed to study this condition, but they often show contradictory results, due to technical constraints and/or model limitations. Therefore, new tools are needed to better investigate pathological features in an unbiased and technically refined approach, towards improving understanding of disease progression. Herein, we describe three simple protocols to phenotype cellular models of HCM in vitro, in a high-throughput manner where technical artefacts are minimized. These are aimed at investigating: (1) Hypertrophy, by measuring cell volume by flow cytometry; (2) HCM molecular features, through the analysis of a hypertrophic marker, multinucleation, and sarcomeric disarray by high-content imaging; and (3) mitochondrial respiration and content via the Seahorse™ platform. Collectively, these protocols comprise straightforward tools to evaluate molecular and functional parameters of HCM phenotypes in cardiomyocytes in vitro. These facilitate greater understanding of HCM and high-throughput drug screening approaches and are accessible to all researchers of cardiac disease modelling. Whilst HCM is used as an exemplar, the approaches described are applicable to other cellular models where the investigation of identical biological changes is paramount.

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Musclemotion

Abstract: Supplemental Digital Content is available in the text.

Cited by 267 publications

References 35 publications

Quantitative Evaluation of the Sarcomere Network of Human hiPSC-Derived Cardiomyocytes Using Single-Molecule Localization Microscopy

Quantitative Evaluation of the Sarcomere Network of Human hiPSC-Derived Cardiomyocytes Using Single-Molecule Localization Microscopy

Efficient Cardiac Differentiation of Human Amniotic Fluid-Derived Stem Cells into Induced Pluripotent Stem Cells and Their Potential Immune Privilege

High-Throughput Phenotyping Toolkit for Characterizing Cellular Models of Hypertrophic Cardiomyopathy In Vitro

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