Dataset Information

Microscopy-based cellular contractility assay for adult, neonatal, and hiPSC cardiomyocytes.

ABSTRACT: This protocol provides instructions to acquire high-quality cellular contractility data from adult, neonatal, and human induced pluripotent stem cell-derived cardiomyocytes. Contractility parameters are key to unravel mechanisms underlying cardiac pathologies, yet difficulties in acquiring data can compromise measurement accuracy and reproducibility. We provide optimized steps for microscope and camera setup, as well as cellular selection criteria for different cardiomyocyte cell types, aiming to obtain robust and reliable data. Moreover, we use CONTRACTIONWAVE software to analyze and show the optimized results. For complete details on the use and execution of this profile, please refer to Scalzo et al. (2021).

SUBMITTER: Scalzo S

PROVIDER: S-EPMC8850799 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Microscopy-based cellular contractility assay for adult, neonatal, and hiPSC cardiomyocytes.

Scalzo Sérgio S Mendonça Carolina A T F CATF Kushmerick Christopher C Agero Ubirajara U Guatimosim Silvia S

STAR protocols 20220211 1

This protocol provides instructions to acquire high-quality cellular contractility data from adult, neonatal, and human induced pluripotent stem cell-derived cardiomyocytes. Contractility parameters are key to unravel mechanisms underlying cardiac pathologies, yet difficulties in acquiring data can compromise measurement accuracy and reproducibility. We provide optimized steps for microscope and camera setup, as well as cellular selection criteria for different cardiomyocyte cell types, aiming t ...[more]

PMID: 35199028

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Project description:The mechanical output of contracting cardiomyocytes, the muscle cells of the heart, relates to healthy and disease states of the heart. Culturing cardiomyocytes on arrays of elastomeric microposts can enable inexpensive and high-throughput studies of heart disease at the single-cell level. However, cardiomyocytes weakly adhere to these microposts, which limits the possibility of using biomechanical assays of single cardiomyocytes to study heart disease. We hypothesized that a stable covalent attachment of laminin to the surface of microposts improves cardiomyocyte contractility. We cultured cells on polydimethylsiloxane microposts with laminin covalently bonded with the organosilanes 3-glycidoxypropyltrimethoxysilane and 3-aminopropyltriethoxysilane with glutaraldehyde. We measured displacement of microposts induced by the contractility of mouse neonatal cardiomyocytes, which attach better than mature cardiomyocytes to substrates. We observed time-dependent changes in contractile parameters such as micropost deformation, contractility rates, contraction and relaxation speeds, and the times of contractions. These parameters were affected by the density of laminin on microposts and by the stability of laminin binding to micropost surfaces. Organosilane-mediated binding resulted in higher laminin surface density and laminin binding stability. 3-glycidoxypropyltrimethoxysilane provided the highest laminin density but did not provide stable protein binding with time. Higher surface protein binding stability and strength were observed with 3-aminopropyltriethoxysilane with glutaraldehyde. In cultured cardiomyocytes, contractility rate, contraction speeds, and contraction time increased with higher laminin stability. Given these variations in contractile function, we conclude that binding of laminin to microposts via 3-aminopropyltriethoxysilane with glutaraldehyde improves contractility observed by an increase in beating rate and contraction speed as it occurs during the postnatal maturation of cardiomyocytes. This approach is promising for future studies to mimic in vivo tissue environments.

Dataset Information

Microscopy-based cellular contractility assay for adult, neonatal, and hiPSC cardiomyocytes.

Publications

Microscopy-based cellular contractility assay for adult, neonatal, and hiPSC cardiomyocytes.

Similar Datasets

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