Skip to main content
SpringerLink
Log in

Edge-Detection for Contractility Measurements with Cardiac Spheroids

  • Protocol
  • First Online:
Stem Cell-Derived Models in Toxicology

Part of the book series: Methods in Pharmacology and Toxicology ((MIPT))

Abstract

There is a growing appreciation of the requirement for a more physiological culture environment for in vitro research and toxicology testing. In the field of cardiovascular research, 3D cultures are often developed for regenerative medicine. However, recent innovations in 3D culture production and higher-throughput analysis methods, as well as the availability of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC) with disease-specific genotypes, have opened new possibilities for toxicological studies in vitro. Spheroid contractility and other parameters of cardiac physiology can be measured using a variety of methods. We have adapted a label-free, video-based method originally used for primary rodent cardiomyocytes and used it with a model of 3D co-culture of hiPSC-derived cardiomyocytes and cardiac fibroblasts for the evaluation and development of new therapies and detection of cardiotoxicity.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Roden DM, Lazzara R, Rosen M et al (1996) Multiple mechanisms in the long-QT syndrome. Current knowledge, gaps, and future directions. The SADS Foundation Task Force on LQTS. Circulation 94:1996–2012

    Article  CAS  PubMed  Google Scholar 

  2. Kondo RP, Apstein CS, Eberli FR et al (1998) Increased calcium loading and inotropy without greater cell death in hypoxic rat cardiomyocytes. Am J Physiol 275:H2272–H2282

    CAS  PubMed  Google Scholar 

  3. Timolati F, Anliker T, Groppalli V et al (2009) The role of cell death and myofibrillar damage in contractile dysfunction of long-term cultured adult cardiomyocytes exposed to doxorubicin. Cytotechnology 61:25–36

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Sawyer DB, Zuppinger C, Miller TA et al (2002) Modulation of anthracycline-induced myofibrillar disarray in rat ventricular myocytes by neuregulin-1 beta and anti-erbB2: potential mechanism for trastuzumab-induced cardiotoxicity. Circulation 105:1551–1554

    Article  CAS  PubMed  Google Scholar 

  5. Harder BA, Hefti MA, Eppenberger HM et al (1998) Differential protein localization in sarcomeric and nonsarcomeric contractile structures of cultured cardiomyocytes. J Struct Biol 122:162–175

    Article  CAS  PubMed  Google Scholar 

  6. Eschenhagen T, Mummery C, Knollmann BC (2015) Modelling sarcomeric cardiomyopathies in the dish: from human heart samples to iPSC cardiomyocytes. Cardiovasc Res 105:424–438

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Rajamohan D, Matsa E, Kalra S et al (2013) Current status of drug screening and disease modelling in human pluripotent stem cells. Bioessays 35:281–298

    Article  CAS  PubMed  Google Scholar 

  8. Suter-Dick L, Alves PM, Blaauboer BJ et al (2015) Stem cell-derived systems in toxicology assessment. Stem Cells Dev 24:1284–1296

    Article  PubMed  Google Scholar 

  9. Beauchamp P, Moritz W, Kelm JM et al (2015) Development and characterization of a scaffold-free 3D spheroid model of iPSC-derived human cardiomyocytes. Tissue Eng Part C Methods 21:852–861

    Article  CAS  PubMed  Google Scholar 

  10. Hinz B, Celetta G, Tomasek JJ et al (2001) Alpha-smooth muscle actin expression upregulates fibroblast contractile activity. Mol Biol Cell 12:2730–2741

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Eder A, Hansen A, Uebeler J. et al (2014) Basic Res Cardiol 109:436. doi:10.1007/s00395-014-0436-7

  12. Jahnke H-G, Steel D, Fleischer S et al (2013) A novel 3D label-free monitoring system of hES-derived cardiomyocyte clusters: a step forward to in vitro cardiotoxicity testing. PLoS One 8, e68971

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Rismani Yazdi S, Shadmani A, Bürgel SC et al (2015) Adding the “heart” to hanging drop networks for microphysiological multi-tissue experiments. Lab Chip 15:4138–4147

    Article  CAS  PubMed  Google Scholar 

  14. Mosadegh B, Xiong G, Dunham S et al (2015) Current progress in 3D printing for cardiovascular tissue engineering. Biomed Mater 10:034002

    Article  PubMed  Google Scholar 

  15. Kim J-Y, Fluri DA, Marchan R et al (2015) 3D spherical microtissues and microfluidic technology for multi-tissue experiments and analysis. J Biotechnol 205:24–35

    Article  CAS  PubMed  Google Scholar 

  16. Hirt MN, Hansen A, Eschenhagen T (2014) Cardiac tissue engineering: state of the art. Circ Res 114:354–367

    Article  CAS  PubMed  Google Scholar 

  17. Hansen A, Eder A, Bönstrup M et al (2010) Development of a drug screening platform based on engineered heart tissue. Circ Res 107:35–44

    Article  CAS  PubMed  Google Scholar 

  18. Zuppinger C (2016) 3D culture for cardiac cells. Biochim Biophys Acta 1863(7):1873–1881

    Google Scholar 

  19. Schaaf S, Eder A, Vollert I et al (2014) Generation of strip-format fibrin-based engineered heart tissue (EHT). Methods Mol Biol 1181:121–129

    Article  PubMed  Google Scholar 

  20. Hirschhaeuser F, Menne H, Dittfeld C et al (2010) Multicellular tumor spheroids: an underestimated tool is catching up again. J Biotechnol 148:3–15

    Article  CAS  PubMed  Google Scholar 

  21. Kelm JM, Ehler E, Nielsen LK et al (2004) Design of artificial myocardial microtissues. Tissue Eng 10:201–214

    Article  CAS  PubMed  Google Scholar 

  22. Rimann M, Graf-Hausner U (2012) Synthetic 3D multicellular systems for drug development. Curr Opin Biotechnol 23:803–809

    Article  CAS  PubMed  Google Scholar 

  23. Desroches BR, Zhang P, Choi B-R et al (2012) Functional scaffold-free 3-D cardiac microtissues: a novel model for the investigation of heart cells. Am J Physiol Heart Circ Physiol 302:H2031–H2042

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Frey O, Misun PM, Fluri DA et al (2014) Reconfigurable microfluidic hanging drop network for multi-tissue interaction and analysis. Nat Commun 5:4250

    Article  CAS  PubMed  Google Scholar 

  25. Drewitz M, Helbling M, Fried N et al (2011) Towards automated production and drug sensitivity testing using scaffold-free spherical tumor microtissues. Biotechnol J 6:1488–1496

    Article  CAS  PubMed  Google Scholar 

  26. Messner S, Agarkova I, Moritz W et al (2013) Multi-cell type human liver microtissues for hepatotoxicity testing. Arch Toxicol 87:209–213

    Article  CAS  PubMed  Google Scholar 

  27. Edling Y, Sivertsson LK, Butura A et al (2009) Increased sensitivity for troglitazone-induced cytotoxicity using a human in vitro co-culture model. Toxicol In Vitro 23:1387–1395

    Article  CAS  PubMed  Google Scholar 

  28. Prange JA, Bieri M, Segerer S et al (2016) Human proximal tubule cells form functional microtissues. Pflugers Arch 468(4):739–50

    Google Scholar 

  29. Tirziu D, Giordano FJ, Simons M (2010) Cell communications in the heart. Circulation 122:928–937

    Article  PubMed  PubMed Central  Google Scholar 

  30. Souders CA, Bowers SLK, Baudino TA (2009) Cardiac fibroblast: the renaissance cell. Circ Res 105:1164–1176

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Desmoulière A, Gabbiani G (1995) Myofibroblast differentiation during fibrosis. Exp Nephrol 3(2):134–139

    Google Scholar 

  32. Rohr S, Kucera JP, Fast VG et al (1997) Paradoxical improvement of impulse conduction in cardiac tissue by partial cellular uncoupling. Science 275:841–844

    Article  CAS  PubMed  Google Scholar 

  33. Kelm JM, Diaz Sanchez-Bustamante C, Ehler E et al (2005) VEGF profiling and angiogenesis in human microtissues. J Biotechnol 118:213–229

    Article  CAS  PubMed  Google Scholar 

  34. Bichsel CA, Hall SRR, Schmid RA et al (2015) Primary human lung pericytes support and stabilize in vitro perfusable microvessels. Tissue Eng Part A 21:2166–2176

    Article  CAS  PubMed  Google Scholar 

  35. Kelm JM, Djonov V, Hoerstrup SP et al (2006) Tissue-transplant fusion and vascularization of myocardial microtissues and macrotissues implanted into chicken embryos and rats. Tissue Eng 12:2541–2553

    Article  CAS  PubMed  Google Scholar 

  36. Veerman CC, Kosmidis G, Mummery CL et al (2015) Immaturity of human stem-cell-derived cardiomyocytes in culture: fatal flaw or soluble problem? Stem Cells Dev 24(9):1035–1052

    Article  CAS  PubMed  Google Scholar 

  37. van den Berg CW, Elliott DA, Braam SR et al (2016) Differentiation of human pluripotent stem cells to cardiomyocytes under defined conditions. Methods Mol Biol 1353:163–180

    Article  PubMed  Google Scholar 

  38. Hayakawa T, Kunihiro T, Ando T et al (2014) Image-based evaluation of contraction-relaxation kinetics of human-induced pluripotent stem cell-derived cardiomyocytes: Correlation and complementarity with extracellular electrophysiology. J Mol Cell Cardiol 77:178–191

    Article  CAS  PubMed  Google Scholar 

  39. Lipp P, Hüser J, Pott L et al (1996) Spatially non-uniform Ca2+ signals induced by the reduction of transverse tubules in citrate-loaded guinea-pig ventricular myocytes in culture. J Physiol 497(Pt 3):589–597

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Steadman BW, Moore KB, Spitzer KW et al (1988) A video system for measuring motion in contracting heart cells. IEEE Trans Biomed Eng 35:264–272

    Article  CAS  PubMed  Google Scholar 

  41. Delbridge LM, Roos KP (1997) Optical methods to evaluate the contractile function of unloaded isolated cardiac myocytes. J Mol Cell Cardiol 29:11–25

    Article  CAS  PubMed  Google Scholar 

  42. Pieperhoff S, Wilson KS, Baily J et al (2014) Heart on a plate: histological and functional assessment of isolated adult zebrafish hearts maintained in culture. PLoS One 9, e96771

    Article  PubMed  PubMed Central  Google Scholar 

  43. Brito-Martins M, Harding SE, Ali NN (2008) beta(1)- and beta(2)-adrenoceptor responses in cardiomyocytes derived from human embryonic stem cells: comparison with failing and non-failing adult human heart. Br J Pharmacol 153:751–759

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Kloss D, Fischer M, Rothermel A et al (2008) Drug testing on 3D in vitro tissues trapped on a microcavity chip. Lab Chip 8:879–884

    Article  CAS  PubMed  Google Scholar 

  45. Seferidis VE, Ghanbari M (1993) General approach to block-matching motion estimation. Opt Eng 32:1464–1474

    Article  Google Scholar 

  46. Ahola A, Kiviaho AL, Larsson K et al (2014) Video image-based analysis of single human induced pluripotent stem cell derived cardiomyocyte beating dynamics using digital image correlation. Biomed Eng Online 13:39

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

Funding was provided by the Swiss Heart Foundation. This study was performed with the support of the Microscopy Imaging Center (MIC), University of Bern. I am thanking Prof. Hugues Abriel and the entire channelopathies group (Department of Clinical Research, Univ. Bern) for valuable advice, stimulating discussions, and experimental support.

Author information

Authors and Affiliations

  1. Cardiology, Department of Clinical Research MEM G803b, Bern University Hospital, Murtenstrasse 35, 3008, Bern, Switzerland

    Christian Zuppinger

Authors
  1. Christian Zuppinger
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Christian Zuppinger .

Editor information

Editors and Affiliations

  1. Axion BioSystems, Inc., Atlanta, Georgia, USA

    Mike Clements

  2. Cell Applications, GE Healthcare, Cardiff, United Kingdom

    Liz Roquemore

1 Electronic Supplementary Material

Below is the link to the electronic supplementary material.

337872_1_En_11_MOESM1_ESM.zip

This movie shows a variety of cardiac models that are currently in use to assess cardiac contractility. Top left: an isolated rat heart shows contractions while being perfused by calcium-containing buffer solution. Top middle: a freshly isolated adult rat cardiomyocyte shows shortening induced by electrical field pacing. Top right: adult rat cardiomyocytes have been cultured for 9 days in a serum-containing medium and have formed a 2D layer of spontaneously contracting cells. Middle: 6 different cardiac spheroids made of hiPSC-derived cardiomyocytes (Cellular Dynamics) and cardiac fibroblasts show spontaneous contractions. Right: screenshot from the IonWizard (IonOptix) interface showing the edge of a spheroid with a red cursor following the motion and drawing the red line graph in real-time. (AVI 2,072 kb)

Glossary

3D

Three-dimensional

DMSO

Dimethyl sulfoxide

FCS

Fetal Calf Serum

HBSS

Hank’s balanced salt solution

HCF

Human cardiac fibroblasts

hESC

Human embryonic stem cells

HEPES

4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid

hiPSC

Human induced pluripotent stem cells

PBS

Phosphate buffered saline

PE

Phenylephrine

ROI

Region of interest

Rights and permissions

Reprints and permissions

Copyright information

© 2017 Springer Science+Business Media New York

About this protocol

Cite this protocol

Zuppinger, C. (2017). Edge-Detection for Contractility Measurements with Cardiac Spheroids. In: Clements, M., Roquemore, L. (eds) Stem Cell-Derived Models in Toxicology. Methods in Pharmacology and Toxicology. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6661-5_11

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-6661-5_11

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6659-2

  • Online ISBN: 978-1-4939-6661-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation