Abstract
Atomic force microscopy (AFM) is not only a high-resolution imaging technique but also a sensitive tool able to study biomechanical properties of bio-samples (biomolecules, cells) in native conditions—i.e., in buffered solutions (culturing media) and stable temperature (mostly 37 °C). Micromechanical transducers (cantilevers) are often used to map surface stiffness distribution, adhesion forces, and viscoelastic parameters of living cells; however, they can also be used to monitor time course of cardiomyocytes contraction dynamics (e.g. beating rate, relaxation time), together with other biomechanical properties. Here we describe the construction of an AFM-based biosensor setup designed to study the biomechanical properties of cardiomyocyte clusters, through the use of standard uncoated silicon nitride cantilevers. Force-time curves (mechanocardiograms, MCG) are recorded continuously in real time and in the presence of cardiomyocyte-contraction affecting drugs (e.g., isoproterenol, metoprolol) in the medium, under physiological conditions. The average value of contraction force and the beat rate, as basic biomechanical parameters, represent pharmacological indicators of different phenotype features. Robustness, low computational requirements, and optimal spatial sensitivity (detection limit 200 pN, respectively 20 nm displacement) are the main advantages of the presented method.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Aistrup GL, Shiferaw Y, Kapur S et al (2009) Mechanisms Underlying the Formation and Dynamics of Subcellular Calcium Alternans in the Intact Rat Heart. Circ Res 104:639–649. https://doi.org/10.1161/CIRCRESAHA.108.181909
Stienen GJM (2015) Pathomechanisms in heart failure: the contractile connection. J Muscle Res Cell Motil 36:47–60. https://doi.org/10.1007/s10974-014-9395-8
Holzmann M, Nicko A, Kühl U et al (2008) Complication rate of right ventricular endomyocardial biopsy via the femoral approach: a retrospective and prospective study analyzing 3048 diagnostic procedures over an 11-year period. Circulation 118:1722–1728. https://doi.org/10.1161/CIRCULATIONAHA.107.743427
Imamura T, Kinugawa K, Nitta D et al (2015) Is the internal jugular vein or femoral vein a better approach site for endomyocardial biopsy in heart transplant recipients? Int Heart J 56:67–72. https://doi.org/10.1536/ihj.14-156
Thomson JA, Itskovitz-eldor J, Shapiro SS et al (1998) Embryonic stem cell lines derived from human blastocysts. Science 282:1145–1148. https://doi.org/10.1126/science.282.5391.1145
Takahashi K, Tanabe K, Ohnuki M et al (2007) Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors. Cell 131:861–872. https://doi.org/10.1016/j.cell.2007.11.019
Mummery C, Ward D, Van den Brink CE et al (2002) Cardiomyocyte differentiation of mouse and human embryonic stem cells. J Anat 200:233–242
Acimovic I, Vilotic A, Pesl M et al (2014) Human pluripotent stem cell-derived cardiomyocytes as research and therapeutic tools. Biomed Res Int 2014:512831. https://doi.org/10.1155/2014/512831
Pesl M, Pribyl J, Acimovic I et al (2016) Atomic force microscopy combined with human pluripotent stem cell derived cardiomyocytes for biomechanical sensing. Biosens Bioelectron 85:751–757. https://doi.org/10.1016/j.bios.2016.05.073
Pesl M, Pribyl J, Caluori G et al (2016) Phenotypic assays for analyses of pluripotent stem cell-derived cardiomyocytes. J Mol Recognit. https://doi.org/10.1002/jmr.2602
Dahlmann J, Kensah G, Kempf H et al (2013) The use of agarose microwells for scalable embryoid body formation and cardiac differentiation of human and murine pluripotent stem cells. Biomaterials 34:2463–2471. https://doi.org/10.1016/j.biomaterials.2012.12.024
Pesl M, Ivana A, Pribyl J et al (2014) Molecular and functional characterization of uniform-sized beating embryoid bodies and cardiomyocytes from human embryonic and induced pluripotent stem cells. Biophys J 106:565A–565A
Acknowledgments
We wish to thank Anton Salykin for providing assistance on data evaluation, Stanislava Koskova, Tereza Jurakova, and Aleksandra Vilotic for kind assistance and reliable preparation of cardiomyocyte clusters.
This work was supported by the Ministry of Education, Youth and Sports of the Czech Republic under the projects CEITEC 2020 (LQ1601) and FNUSA-ICRC (LQ1605), FNUSA- ICRC no. CZ.1.05/1.1.00/02.0123 (OP VaVpI), as well as CIISB research infrastructure project LM2015043, project MSM0021622430 and Grant Agency of the Czech Republic (Grant No. P302/12/G157). This work was also supported by the European Regional Development Fund Project “CIISB4HEALTH” no.CZ.02.1.01/0.0/0.0/16_013/0001776. Martin Pešl and Guido Caluori were supported by Grant Agency of the Masaryk University (MUNI/A/1010/2016).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Pribyl, J. et al. (2019). Biomechanical Characterization of Human Pluripotent Stem Cell-Derived Cardiomyocytes by Use of Atomic Force Microscopy. In: Santos, N., Carvalho, F. (eds) Atomic Force Microscopy. Methods in Molecular Biology, vol 1886. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8894-5_20
Download citation
DOI: https://doi.org/10.1007/978-1-4939-8894-5_20
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-8893-8
Online ISBN: 978-1-4939-8894-5
eBook Packages: Springer Protocols