European Biophysics Journal

, Volume 43, Issue 1, pp 11–23 | Cite as

Active contractions in single suspended epithelial cells

  • Markus Gyger
  • Roland Stange
  • Tobias R. Kießling
  • Anatol Fritsch
  • Katja B. Kostelnik
  • Annette G. Beck-Sickinger
  • Mareike Zink
  • Josef A. Käs
Original Paper
First Online:
Received:
Revised:
Accepted:

Abstract

Investigations of active contractions in tissue cells to date have been focused on cells that exert forces via adhesion sites to substrates or to other cells. In this study we show that also suspended epithelial cells exhibit contractility, revealing that contractions can occur independently of focal adhesions. We employ the Optical Stretcher to measure adhesion-independent mechanical properties of an epithelial cell line transfected with a heat-sensitive cation channel. During stretching the heat transferred to the ion channel causes a pronounced Ca2+ influx through the plasma membrane that can be blocked by adequate drugs. This way the contractile forces in suspended cells are shown to be partially triggered by Ca2+ signaling. A phenomenological mathematical model is presented, incorporating a term accounting for the active stress exerted by the cell, which is both necessary and sufficient to describe the observed increase in strain when the Ca2+ influx is blocked. The median and the shape of the strain distributions depend on the activity of the cells. Hence, it is unlikely that they can be described by a simple Gaussian or log normal distribution, but depend on specific cellular properties such as active contractions. Our results underline the importance of considering activity when measuring cellular mechanical properties even in the absence of measurable contractions. Thus, the presented method to quantify active contractions of suspended cells offers new perspectives for a better understanding of cellular force generation with possible implications for medical diagnosis and therapy.

Keywords

Active soft matter Epithelial contractions Calcium Optical Stretcher Cell rheology 
This is a preview of subscription content, log in to check access.

Notes

Acknowledgments

We thank B. Fabry (Friedrich Alexander University of Erlangen-Nürnberg) for helpful discussions and advice. Furthermore, we thank D. Julius (UCSF) for providing the TRPV1 transfected HEK293 cells. The project was funded by SAB-project 13403 (EFRE) and Agescreen-Biophotonics 5 Program [funded by the German Federal Ministry of Education and Research (BMBF)] and the graduate school Leipzig School of Natural Sciences—Building with Molecules and Nano-objects “BuildMoNa” of the Universität Leipzig.

Supplementary material

249_2013_935_MOESM1_ESM.pdf (1.9 mb)
PDF (1933 KB)
249_2013_935_MOESM2_ESM.pdf (1.9 mb)
PDF (1933 KB)

AVI (3129 KB)

AVI (3159 KB)

References

  1. Ananthakrishnan R, Guck J, Wottawah F, Schinkinger S, Lincoln B, Romeyke M, Käs J (2005) Modelling the structural response of an eukaryotic cell in the Optical Stretcher. Curr Sci India 88(9):1434–1440Google Scholar
  2. Bement WM, Forscher P, Mooseker MS (1993) A novel cytoskeletal structure involved in purse string wound closure and cell polarity maintenance. J Cell Biol 121(3):565–578. doi: 10.1083/jcb.121.3.565 PubMedCrossRefGoogle Scholar
  3. Bendix PM, Koenderink GH, Cuvelier D, Dogic Z, Koeleman BN, Brieher WM, Field CM, Mahadevan L, Weitz DA (2008) A quantitative analysis of contractility in active cytoskeletal protein networks. Biophys J 94(8):3126–3136. doi: 10.1529/biophysj.107.117960 PubMedCentralPubMedCrossRefGoogle Scholar
  4. Beningo KA, Hamao K, Dembo M, Wang YL, Hosoya H (2006) Traction forces of fibroblasts are regulated by the Rho-dependent kinase but not by the myosin light chain kinase. Arch Biochem Biophys 456(2):224–231. doi: 10.1016/j.abb.2006.09.025 PubMedCentralPubMedCrossRefGoogle Scholar
  5. Broedersz CP, MacKintosh FC (2011) Molecular motors stiffen non-affine semiflexible polymer networks. Soft Matter 7(7):3186–3191. doi: 10.1039/C0SM01004A CrossRefGoogle Scholar
  6. Caterina M, Schumacher M, Tominaga M, Rosen T, Levine J, Julius D (1997) The capsaicin receptor: a heat-activated ion channel in the pain pathway. Nature 389(6653):816–824PubMedCrossRefGoogle Scholar
  7. Caterina MJ, Rosen TA, Tominaga M, Brake AJ, Julius D (1999) A capsaicin-receptor homologue with a high threshold for noxious heat. Nature 398(6726):436–441PubMedCrossRefGoogle Scholar
  8. Cates ME, MacKintosh FC (2011) Active soft matter. Soft Matter 7(7):3050–3051. doi: 10.1039/C1SM90014E CrossRefGoogle Scholar
  9. Desprat N, Richert A, Simeon J, Asnacios A (2005) Creep function of a single living cell. Biophys J 88(3):2224–2233. doi: 10.1529/biophysj.104.050278 PubMedCentralPubMedCrossRefGoogle Scholar
  10. Efron B, Tibshirani RJ (1994) An introduction to the bootstrap. Chapman & Hall/CRC Monographs on statistics and applied probability. Taylor & Francis, London. http://books.google.de/books?id=gLlpIUxRntoC
  11. Fabry B, Maksym GN, Butler JP, Glogauer M, Navajas D, Fredberg JJ (2001) Scaling the microrheology of living cells. Phys Rev Lett 87(14):148102. doi: 10.1103/PhysRevLett.87.148102 PubMedCrossRefGoogle Scholar
  12. Fernández P, Ott A (2008) Single cell mechanics: stress stiffening and kinematic hardening. Phys Rev Lett 100(23):238102. doi: 10.1103/PhysRevLett.100.238102 PubMedCrossRefGoogle Scholar
  13. Fernández P, Pullarkat PA, Ott A (2006) A master relation defines the nonlinear viscoelasticity of single fibroblasts. Biophys J 90(10):3796–3805. doi: 10.1529/biophysj.105.072215 PubMedCentralPubMedCrossRefGoogle Scholar
  14. Friedl P, Gilmour D (2009) Collective cell migration in morphogenesis, regeneration and cancer. Nat Rev Mol Cell Bio 10(7):445–457CrossRefGoogle Scholar
  15. Fritsch A, Höckel M, Kiessling T, Nnetu KD, Wetzel F, Zink M, Käs JA (2010) Are biomechanical changes necessary for tumour progression? Nat Phys 6(10):730–732CrossRefGoogle Scholar
  16. Fukata Y, Kaibuchi K, Amano M, Kaibuchi K (2001) Rho–Rho-kinase pathway in smooth muscle contraction and cytoskeletal reorganization of non-muscle cells. Trends Pharmacol Sci 22(1):32–39. doi: 10.1016/S0165-6147(00)01596-0 PubMedCrossRefGoogle Scholar
  17. Gardel ML, Shin JH, MacKintosh FC, Mahadevan L, Matsudaira P, Weitz DA (2004) Elastic behavior of cross-linked and bundled actin networks. Science 304(5675):1301–1305. doi: 10.1126/science.1095087 PubMedCrossRefGoogle Scholar
  18. Grishchuk EL, Molodtsov MI, Ataullakhanov FI, McIntosh JR (2005) Force production by disassembling microtubules. Nature 438(7066):384–388PubMedCrossRefGoogle Scholar
  19. Guck J, Schinkinger S, Lincoln B, Wottawah F, Ebert S, Romeyke M, Lenz D, Erickson HM, Ananthakrishnan R, Mitchell D, Käs J, Ulvick S, Bilby C (2005) Optical deformability as an inherent cell marker for testing malignant transformation and metastatic competence. Biophys J 88(5):3689–3698. doi: 10.1529/biophysj.104.045476 PubMedCentralPubMedCrossRefGoogle Scholar
  20. Gyger M, Rose D, Stange R, Kießling T, Zink M, Fabry B, Käs JA (2011) Calcium imaging in the Optical Stretcher. Opt Express 19(20):19212–19222. doi: 10.1364/OE.19.019212 PubMedCrossRefGoogle Scholar
  21. Hoffman BD, Massiera G, Van Citters KM, Crocker JC (2006) The consensus mechanics of cultured mammalian cells. Proc Natl Acad Sci USA 103(27):10259–10264. doi: 10.1073/pnas.0510348103 PubMedCrossRefGoogle Scholar
  22. Icard-Arcizet D, Cardoso O, Richert A, Hénon S (2008) Cell stiffening in response to external stress is correlated to actin recruitment. Biophys J 94(7):2906–2913. doi: 10.1529/biophysj.107.118265 PubMedCentralPubMedCrossRefGoogle Scholar
  23. Ingber DE (1997) Tensegrity: the architectural basis of cellular mechanotransduction. Annu Rev Physiol 59(1):575–599. doi: 10.1146/annurev.physiol.59.1.575 PubMedCrossRefGoogle Scholar
  24. Jacinto A, Wood W, Balayo T, Turmaine M, Martinez-Arias A, Martin P (2000) Dynamic actin-based epithelial adhesion and cell matching during Drosophila dorsal closure. Curr Biol 10(22):1420–1426. doi: 10.1016/S0960-9822(00)00796-X PubMedCrossRefGoogle Scholar
  25. Jonas O, Mierke CT, Käs JA (2011) Invasive cancer cell lines exhibit biomechanical properties that are distinct from their noninvasive counterparts. Soft Matter 7(24):11488–11495. doi: 10.1039/C1SM05532A CrossRefGoogle Scholar
  26. Kollmannsberger P, Mierke CT, Fabry B (2011) Nonlinear viscoelasticity of adherent cells is controlled by cytoskeletal tension. Soft Matter 7(7):3127–3132. doi: 10.1039/C0SM00833H CrossRefGoogle Scholar
  27. Kumar N, Tomar A, Parrill AL, Khurana S (2004) Functional dissection and molecular characterization of calcium-sensitive actin-capping and actin-depolymerizing sites in villin. J Biol Chem 279(43):45036–45046. doi: 10.1074/jbc.M405424200 PubMedCrossRefGoogle Scholar
  28. Larson L, Arnaudeau S, Gibson B, Li W, Krause R, Hao B, Bamburg JR, Lew DP, Demaurex N, Southwick F (2005) Gelsolin mediates calcium-dependent disassembly of Listeria actin tails. Proc Natl Acad Sci USA 102(6):1921–1926. doi: 10.1073/pnas.0409062102 PubMedCrossRefGoogle Scholar
  29. Lau AWC, Hoffman BD, Davies A, Crocker JC, Lubensky TC (2003) Microrheology, stress fluctuations, and active behavior of living cells. Phys Rev Lett 91(19):198101. doi: 10.1103/PhysRevLett.91.198101 PubMedCrossRefGoogle Scholar
  30. Lin YC, Koenderink GH, MacKintosh FC, Weitz DA (2011) Control of non-linear elasticity in F-actin networks with microtubules. Soft Matter 7(3):902–906CrossRefGoogle Scholar
  31. Lincoln B, Schinkinger S, Travis K, Wottawah F, Ebert S, Sauer F, Guck J (2007) Reconfigurable microfluidic integration of a dual-beam laser trap with biomedical applications. Biomed Microdev 9(5):703–710CrossRefGoogle Scholar
  32. Liverpool TB, Marchetti MC, Joanny JF, Prost J (2009) Mechanical response of active gels. Europhys Lett 85(1):18007CrossRefGoogle Scholar
  33. Lo CM, Buxton DB, Chua GC, Dembo M, Adelstein RS, Wang YL (2004) Nonmuscle myosin IIB is involved in the guidance of fibroblast migration. Mol Biol Cell 15(3):982–989. doi: 10.1091/mbc.E03-06-0359 PubMedCentralPubMedCrossRefGoogle Scholar
  34. Lo CM, Wang HB, Dembo M, Wang YL (2000) Cell movement is guided by the rigidity of the substrate. Biophys J 79(1):144–152. doi: 10.1016/S0006-3495(00)76279-5 PubMedCentralPubMedCrossRefGoogle Scholar
  35. Lodish H (2000) Molecular cell biology. W.H. Freeman and Company, New YorkGoogle Scholar
  36. MacKintosh FC, Levine AJ (2008) Nonequilibrium mechanics and dynamics of motor-activated gels. Phys Rev Lett 100(1):018104. doi: 10.1103/PhysRevLett.100.018104 PubMedCrossRefGoogle Scholar
  37. Mader CC, Hinchcliffe EH, Wang YL (2007) Probing cell shape regulation with patterned substratum: requirement of myosin II-mediated contractility. Soft Matter 3(3):357–363. doi: 10.1039/B606590B CrossRefGoogle Scholar
  38. Maloney JM, Nikova D, Lautenschläger F, Clarke E, Langer R, Guck J, Vliet KJV (2010) Mesenchymal stem cell mechanics from the attached to the suspended state. Biophys J 99(8):2479–2487. doi: 10.1016/j.bpj.2010.08.052 PubMedCentralPubMedCrossRefGoogle Scholar
  39. Mann HB, Whitney DR (1947) On a test of whether one of two random variables is stochastically larger than the other. Ann Math Statist 18(1):50–60CrossRefGoogle Scholar
  40. Martin AC, Kaschube M, Wieschaus EF (2009) Pulsed contractions of an actin-myosin network drive apical constriction. Nature 457(7228):495–499PubMedCentralPubMedCrossRefGoogle Scholar
  41. Mierke CT, Rösel D, Fabry B, Brábek J (2008) Contractile forces in tumor cell migration. Eur J Cell Biol 87(8–9):669–676. doi: 10.1016/j.ejcb.2008.01.002 PubMedCentralPubMedCrossRefGoogle Scholar
  42. Mitrossilis D, Fouchard J, Guiroy A, Desprat N, Rodriguez N, Fabry B, Asnacios A (2009) Single-cell response to stiffness exhibits muscle-like behavior. Proc Natl Acad Sci USA 106(43):18243–18248. doi: 10.1073/pnas.0903994106 PubMedCrossRefGoogle Scholar
  43. Mizuno D, Tardin C, Schmidt CF, MacKintosh FC (2007) Nonequilibrium mechanics of active cytoskeletal networks. Science 315(5810):370–373. doi: 10.1126/science.1134404 PubMedCrossRefGoogle Scholar
  44. Mogilner A, Keren K (2009) The shape of motile cells. Curr Biol 19(17):R762–R771. doi: 10.1016/j.cub.2009.06.053 PubMedCentralPubMedCrossRefGoogle Scholar
  45. Mogilner A, Oster G (2003) Force generation by actin polymerization II: the elastic ratchet and tethered filaments. Biophys J 84(3):1591–1605PubMedCentralPubMedCrossRefGoogle Scholar
  46. Molodtsov M, Grishchuk E, McIntosh J, Ataullakhanov F (2007) Measurement of the force developed by disassembling microtubule during calcium-induced depolymerization. Dokl Biochem Biophys 412(1):18–21PubMedCrossRefGoogle Scholar
  47. Munevar S, Wang YL, Dembo M (2001) Traction force microscopy of migrating normal and H-ras transformed 3T3 fibroblasts. Biophys J 80(4):1744–1757. doi: 10.1016/S0006-3495(01)76145-0 PubMedCentralPubMedCrossRefGoogle Scholar
  48. Park SW, Schapery RA (1999) Methods of interconversion between linear viscoelastic material functions. Part I–a numerical method based on Prony series. Int J Solids Struct 36(11):1653–1675. doi: 10.1016/S0020-7683(98)00055-9 CrossRefGoogle Scholar
  49. Pelham RJ, Wang YL (1999) High resolution detection of mechanical forces exerted by locomoting fibroblasts on the substrate. Mol Biol Cell 10(4):935–945PubMedCentralPubMedCrossRefGoogle Scholar
  50. Remmerbach TW, Wottawah F, Dietrich J, Lincoln B, Wittekind C, Guck J (2009) Oral cancer diagnosis by mechanical phenotyping. Cancer Res 69(5):1728–1732. doi: 10.1158/0008-5472.CAN-08-4073 PubMedCrossRefGoogle Scholar
  51. Ridley AJ, Schwartz MA, Burridge K, Firtel RA, Ginsberg MH, Borisy G, Parsons JT, Horwitz AR (2003) Cell migration: Integrating signals from front to back. Science 302(5651):1704–1709. doi: 10.1126/science.1092053 PubMedCrossRefGoogle Scholar
  52. Saitoh M, Ishikawa T, Matsushima S, Naka M, Hidaka H (1987) Selective inhibition of catalytic activity of smooth muscle myosin light chain kinase. J Biol Chem 262(16):7796–7801PubMedGoogle Scholar
  53. Salmon ED, Segall RR (1980) Calcium-labile mitotic spindles isolated from sea urchin eggs (Lytechinus variegatus). J Cell Bio 86(2):355–365. doi: 10.1083/jcb.86.2.355 CrossRefGoogle Scholar
  54. Somlyo AP, Somlyo AV (2003) Ca2+ sensitivity of smooth muscle and nonmuscle myosin II: modulated by g proteins, kinases, and myosin phosphatase. Physiol Rev 83(4):1325–1358. doi: 10.1152/physrev.00023.2003 PubMedGoogle Scholar
  55. Stamenovic D, Suki B, Fabry B, Wang N, Fredberg JJ, Buy JE (2004) Rheology of airway smooth muscle cells is associated with cytoskeletal contractile stress. J Appl Physiol 96(5):1600–1605. doi: 10.1152/japplphysiol.00595.2003 PubMedCrossRefGoogle Scholar
  56. Sun SX, Walcott S, Wolgemuth CW (2010) Cytoskeletal cross-linking and bundling in motor-independent contraction. Curr Biol 20(15):R649–R654. doi: 10.1016/j.cub.2010.07.004 PubMedCentralPubMedCrossRefGoogle Scholar
  57. Tamada M, Perez TD, Nelson WJ, Sheetz MP (2007) Two distinct modes of myosin assembly and dynamics during epithelial wound closure. J Cell Biol 176(1):27–33. doi: 10.1083/jcb.200609116 PubMedCrossRefGoogle Scholar
  58. Thoumine O, Ott A (1997) Time scale dependent viscoelastic and contractile regimes in fibroblasts probed by microplate manipulation. J Cell Sci 110(17):2109–2116PubMedGoogle Scholar
  59. Tominaga M, Caterina MJ, Malmberg AB, Rosen TA, Gilbert H, Skinner K, Raumann BE, Basbaum AI, Julius D (1998) The cloned capsaicin receptor integrates multiple pain-producing stimuli. Neuron 21(3):531–543. doi: 10.1016/S0896-6273(00)80564-4 PubMedCrossRefGoogle Scholar
  60. Tschoegl NW (1989) The phenomenological theory of linear viscoelastic behavior. Springer-Verlag, New YorkCrossRefGoogle Scholar
  61. Vicente-Manzanares M, Ma X, Adelstein RS, Horwitz AR (2009) Non-muscle myosin II takes centre stage in cell adhesion and migration. Nat Rev Mol Cell Bio 10(11):778–790CrossRefGoogle Scholar
  62. Walsh TP, Weber A, Davis K, Bonder E, Mooseker M (1984) Calcium dependence of villin-induced actin depolymerization. Biochemistry (US) 23(25):6099–6102. doi: 10.1021/bi00320a030 CrossRefGoogle Scholar
  63. Wang N, Butler J, Ingber D (1993) Mechanotransduction across the cell surface and through the cytoskeleton. Science 260(5111):1124–1127. doi: 10.1126/science.7684161 PubMedCrossRefGoogle Scholar
  64. Wang HB, Dembo M, Hanks SK, Wang Yl (2001a) Focal adhesion kinase is involved in mechanosensing during fibroblast migration. Proc Natl Acad Sci USA 98(20):11295–11300. doi: 10.1073/pnas.201201198 PubMedCrossRefGoogle Scholar
  65. Wang N, Naruse K, Stamenović D, Fredberg JJ, Mijailovich SM, Tolić-Norrelykke IM, Polte T, Mannix R, Ingber DE (2001b) Mechanical behavior in living cells consistent with the tensegrity model. Proc Natl Acad Sci USA 98(14):7765–7770. doi: 10.1073/pnas.141199598 PubMedCrossRefGoogle Scholar
  66. Wei C, Wang X, Chen M, Ouyang K, Song LS, Cheng H (2009) Calcium flickers steer cell migration. Nature 457(7231):901–905PubMedCentralPubMedCrossRefGoogle Scholar
  67. Wottawah F, Schinkinger S, Lincoln B, Ananthakrishnan R, Romeyke M, Guck J, Käs J (2005a) Optical rheology of biological cells. Phys Rev Lett 94(9):098103. doi: 10.1103/PhysRevLett.94.098103 PubMedCrossRefGoogle Scholar
  68. Wottawah F, Schinkinger S, Lincoln B, Ebert S, Müller K, Sauer F, Travis K, Guck J (2005b) Characterizing single suspended cells by optorheology. Acta Biomater 1(3):263–271. doi: 10.1016/j.actbio.2005.02.010 PubMedCrossRefGoogle Scholar
  69. Wysolmerski RB, Lagunoff D (1990) Involvement of myosin light-chain kinase in endothelial cell retraction. Proc Natl Acad Sci USA 87(1):16–20PubMedCrossRefGoogle Scholar
  70. Xu J, Tseng Y, Wirtz D (2000) Strain hardening of actin filament networks. J Biol Chem 275(46):35886–35892. doi: 10.1074/jbc.M002377200 PubMedCrossRefGoogle Scholar
  71. Yamada S, Wirtz D, Kuo SC (2000) Mechanics of living cells measured by laser tracking microrheology. Biophys J 78(4):1736–1747. doi: 10.1016/S0006-3495(00)76725-7 PubMedCentralPubMedCrossRefGoogle Scholar
  72. Yeung T, Georges PC, Flanagan LA, Marg B, Ortiz M, Funaki M, Zahir N, Ming W, Weaver V, Janmey PA (2005) Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesion. Cell Motil Cytoskel 60(1):24–34. doi: 10.1002/cm.20041 CrossRefGoogle Scholar

Copyright information

© European Biophysical Societies' Association 2013

Authors and Affiliations

  • Markus Gyger
    • 1
  • Roland Stange
    • 1
  • Tobias R. Kießling
    • 1
  • Anatol Fritsch
    • 1
  • Katja B. Kostelnik
    • 2
  • Annette G. Beck-Sickinger
    • 2
  • Mareike Zink
    • 1
  • Josef A. Käs
    • 1
  1. 1.Abteilung für Physik der weichen Materie, Institut für Experimentelle Physik I Universität LeipzigLeipzigGermany
  2. 2.Institut für BiochemieUniversität LeipzigLeipzigGermany

Personalised recommendations