Probing the subcellular nanostructure of engineered human cardiomyocytes in 3D tissue

Javor, Josh; Ewoldt, Jourdan; Cloonan, Paige E.; Chopra, Anant; Luu, Rebeccah J.; Freychet, Guillaume; Zhernenkov, Mikhail; Ludwig, Karl; Seidman, Jonathan G.; Chen, Christopher; Bishop, David J.

doi:10.1038/s41378-020-00234-x

Cited by 9 publications

(29 citation statements)

References 37 publications

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“…One such example is Beamline I22 at the DIAMOND light source in the UK that was recently used for a skeletal muscle diffraction study [ 44 ]. A recent study of engineered heart tissue [ 45 ] was done on the LIXS beamline at NSLS II at Brookhaven National Laboratory (Upton, NY, USA).…”

Section: Introductionmentioning

confidence: 99%

Small Angle X-ray Diffraction as a Tool for Structural Characterization of Muscle Disease

Irving

2022

IJMS

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Small angle X-ray fiber diffraction is the method of choice for obtaining molecular level structural information from striated muscle fibers under hydrated physiological conditions. For many decades this technique had been used primarily for investigating basic biophysical questions regarding muscle contraction and regulation and its use confined to a relatively small group of expert practitioners. Over the last 20 years, however, X-ray diffraction has emerged as an important tool for investigating the structural consequences of cardiac and skeletal myopathies. In this review we show how simple and straightforward measurements, accessible to non-experts, can be used to extract biophysical parameters that can help explain and characterize the physiology and pathology of a given experimental system. We provide a comprehensive guide to the range of the kinds of measurements that can be made and illustrate how they have been used to provide insights into the structural basis of pathology in a comprehensive review of the literature. We also show how these kinds of measurements can inform current controversies and indicate some future directions.

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Section: Introductionmentioning

confidence: 99%

Small Angle X-ray Diffraction as a Tool for Structural Characterization of Muscle Disease

Irving

2022

IJMS

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“…Engineered Cardiac Platform: Polydimethylsiloxane (PDMS) milli-tug were cast from PDMS (Sylgard 184, Dow-Corning) with a curing agent to base ratio of 1:25 via replica molding from polypropylene master. The method and milli-tug dimension were modified based on Javor et al's [23] protocols. Detailedly, milli-tugs were baked overnight at 70 °C before immersed in ethanol to be peeled off from the master.…”

Section: Methodsmentioning

confidence: 99%

Effect of Electrical Stimulation on Spontaneously Beating Dynamics of Cardiac Tissues: An Analysis Using Digital Image Correlation

Lou

Rubfiaro

et al. 2021

Adv Materials Technologies

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promoting cardiomyocyte maturity as indicated by sarcomere size, gene expression, metabolism, and Ca 2+ waves. Similarly, Khodabukus et al. [2] found electrical stimulation (70 mA amplitude, 2 ms, and 1 or 10 Hz) improved protein expression, force generation, calcium handling, and fatty acid oxidation in human skeletal muscles, which demonstrated the enhancement in maturation, structure, and metabolism. However, there are still concerns about the effectiveness of external stimuli. For example, in a review summarized by Love et al., [3] electrical stimulations can trigger an increase, decrease, or neutral effect on cell proliferation and apoptosis. This is perhaps due to non-optimized electrical and material parameters: voltage, type, number of pauses, frequency, duration, treatment period, and cell type. Thus, the underlying mechanism has not thoroughly been studied and understood.Simple detection or verification methods are necessary to fully decipher the mechanism and optimize the function of electric stimuli. Currently used techniques such as gene expression and Ca 2+ detection methods are mature but complex. Mechanical measurement methods (e.g., tensile, hardness, bending, burst testing) have been explored, which were nonideal for soft non-uniform cardiac tissues due to their [4] small size, viscoelasticity, hierarchical and porous structures, hydration levels, complex compositions, and high surface roughness. Novel methods, including patch-clamp, indentation, [5] optical tweezer, [6] multifunctional membrane, [7] and voltagesensitive dye, have also been applied for mechanic and beating properties' measurement. However, these methods are invasive; and the results exhibit high variations and deviations due to the viscoelasticity and non-uniformity characteristics of soft biomaterials. [6] Compared to invasive mechanical measuring methods, digital image correlation (DIC) [8] is a non-invasive and comprehensive method to detect cardiac beating mechanics, measuring nano-and micro-deformation of the tested object through the change of gray value patterned subsets. For example, Shradhanjali et al. [9] designed an adaptive reference-DIC method to study the mechanics of cardiomyocyte contraction.The mechanism of DIC analysis involves tracking and imaging micro-deformation of the tested object through detecting the corresponding subsets' differences between the reference and deformed images. [8] More specifically, it detects the correlations among all the subsets in the reference and Electrical pacing/stimulations (EP) have been widely adopted to promote the maturation of hiPSC-derived cardiomyocytes. However, there is a debate about their functions and effectiveness due to non-optimized pacing conditions. Here, the effectiveness of EP (13 V cm −1 , 2 ms in width, and 5 Hz frequency) on cardiac tissue beating mechanics are analyzed using digital image correlation (DIC). The cardiac tissues with and without EP at tissue culture time from day 2 to 11 (D2-D11) are characterized and compared. The results indicate EP dec...

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“…Compared to EM, small angle X-ray scattering (SAXS) has fewer disadvantages in studying biological tissue if radiation damage is managed 21 . This can be done with a combination of exposure time, microfocusing, beam intensity, and beam energy.…”

Section: Introductionmentioning

confidence: 99%

“…Information about the nanostructure of cardiomyocytes has accumulated from SAXS studies performed on cardiac tissue from humans, rats, and zebrafish [21][22][23] . The capabilities of SAXS for the characterization of the dynamics of the nanostructure of live cardiomyocytes were showcased by Brunello et al 23 .…”

Section: Introductionmentioning

confidence: 99%

Structural maturation of myofilaments in engineered 3D cardiac microtissues characterized using small angle X-ray scattering

Javor

Ewoldt

Zhernenkov

et al. 2023

Preprint

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Understanding the structural and functional development of human-induced pluripotent stem-cell-derived cardiomyocytes is essential to engineering cardiac tissue that enables pharmaceutical testing, modeling diseases, and designing therapies. Here, we used a method not commonly applied to biological materials, small angle X-ray scattering to characterize the structural development of human induced pluripotent stem-cell-derived cardiomyocytes within 3D engineered tissues during their preliminary stages of maturation. An innovative X-ray scattering experimental setup enabled the visualization of a systematic variation in the cardiomyocyte myofilament spacing with maturation time. The myofilament lattice spacing monotonically decreased as the tissue matured from its initial post-seeding state over the span of ten days. Visualization of the spacing at a grid of positions in the tissue provides a new approach to characterizing the maturation and organization of cardiomyocyte myofilaments and has the potential to help elucidate mechanisms of pathophysiology, disease progression, thereby stimulating new biological hypotheses in stem cell engineering.

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Probing the subcellular nanostructure of engineered human cardiomyocytes in 3D tissue

Cited by 9 publications

References 37 publications

Small Angle X-ray Diffraction as a Tool for Structural Characterization of Muscle Disease

Small Angle X-ray Diffraction as a Tool for Structural Characterization of Muscle Disease

Effect of Electrical Stimulation on Spontaneously Beating Dynamics of Cardiac Tissues: An Analysis Using Digital Image Correlation

Structural maturation of myofilaments in engineered 3D cardiac microtissues characterized using small angle X-ray scattering

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