This report describes an approach to extend indefinitely the duration of traction force measurements with cells cultured on soft polyacrylamide gels. Typical observation times in traction force measurements on similar substrates have been limited to 24–48 h, but cell differentiation or responses to external stimuli often occur over much longer periods of several days or weeks. This study describes a method for covalently linking fluorescent marker beads to a polyacrylamide matrix that renders the hydrogels useful for traction force measurements over several days. This approach was validated by comparing the contractility of C2C12 murine skeletal muscle cells prior to myotube formation, after one day in culture, with that of myotubes after 7 days in culture. Measured tractions increased concurrent with the differentiation of C2C12 cells to the contractile, myotube phenotype. Covalent bead linkage thus extends the useful period during which traction force data can be obtained with cells cultured on optically transparent polyacrylamide hydrogels with controlled elastic moduli.
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Butler, J. P., I. M. Tolic-Norrelykke, B. Fabry, and J. J. Fredberg. Traction fields, moments, and strain energy that cells exert on their surroundings. Am. J. Physiol. Cell Physiol. 282:C595–C605, 2002.
Chen, C. S., J. Tan, and J. Tien. Mechanotransduction at cell–matrix and cell–cell contacts. Ann. Rev. Biomed. Eng. 6:275–302, 2004.
Choquet, D., D. P. Felsenfeld, and M. P. Sheetz. Extracellular matrix rigidity causes strengthening of integrin-cytoskeleton linkages. Cell 88:39–48, 1997.
Domke, J., and M. Radmacher. Measuring the elastic properties of thin polymer films with the atomic force microscope. Langmuir 14:3320–3325, 1998.
Dong, R., T. W. Jensen, K. Engberg, R. G. Nuzzo, and D. E. Leckband. Variably elastic hydrogel patterned via capillary action in microchannels. Langmuir 23:1483–1488, 2007.
Engler, A. J., S. Sen, H. L. Sweeney, and D. E. Discher. Matrix elasticity directs stem cell lineage specification. Cell 126:677–689, 2006.
Ganz, A., M. Lambert, A. Saez, P. Silberzan, A. Buguin, R. M. Mage, and B. Ladoux. Traction forces exerted through N-cadherin contacts. Biol. Cell 98:721–730, 2006.
Gunawan, R. C., E. R. Choban, J. E. Conour, J. Silvestre, L. B. Schook, H. R. Gaskins, D. E. Leckband, and P. J. A. Kenis. Regiospecific control of protein expression in cells cultured on two-component counter gradients of extracellular matrix proteins. Langmuir 21:3061–3068, 2005.
Harris, A. K., P. Wild, and D. Stopak. Silicone rubber substrata: a new wrinkle in the study of cell locomotion. Science 208:177–179, 1980.
Hinterberger, T. J., and K. F. Barald. Fusion between myoblasts and adult muscle fibers promotes remodeling of fibers into myotubes in vitro. Development 109:139–147, 1990.
Kim, D.-H., P. K. Wong, J. Park, A. Levchenko, and Y. Sun. Microengineered platforms for cell mechanobiology. Ann. Rev. Biomed. Eng. 11:203–233, 2009.
Kumar, S., and V. Weaver. Mechanics, malignancy, and metastasis: the force journey of a tumor cell. Cancer Metastasis Rev. 28:113–127, 2009.
Li, L., N. Sharma, U. Chipada, X. Jiang, R. Schloss, M. Yarmush, and N. Langrana. Functional modulation of ES-derived hepatocyte lineage cells via substrate compliance alteration. Ann. Biomed. Eng. 36:865–876, 2008.
Lombardi, M. L., D. A. Knect, M. Dembo, and J. Lee. Traction force microscopy in Dictyostelium reveals distinct roles for myosin II motor and actin-crosslinking activity in polarized cell movement. J. Cell Sci. 120:1624–1634, 2007.
McMahon, D. K., P. A. Anderson, R. Nassar, J. B. Bunting, Z. Saba, A. E. Oakeley, and N. N. Malouf. C2C12 cells: biophysical, biochemical, and immunocytochemical properties. Am. J. Physiol. Cell Physiol. 266:C1795–C1802, 1994.
Pless, D. D., Y. C. Lee, S. Roseman, and R. L. Schnaar. Specific cell adhesion to immobilized glycoproteins demonstrated using new reagents for protein and glycoprotein immobilization. J. Biol. Chem. 258:2340–2349, 1983.
Reinhart-King, C. A., M. Dembo, and D. A. Hammer. The dynamics and mechanics of endothelial cell spreading. Biophys. J. 89:676–689, 2005.
Rowlands, A. S., P. A. George, and J. J. Cooper-White. Directing osteogenic and myogenic differentiation of MSCs: interplay of stiffness and adhesive ligand presentation. Am. J. Physiol. Cell Physiol. 295:C1037–C1044, 2008.
Saha, K., A. J. Keung, E. Irwin, L. Yang, L. Little, D. V. Schaffer, and K. E. Healy. Substrate modulus directs neural stem cell behavior. Biophys. J. 95:4426–4438, 2008.
Teixeira, A. I., S. Ilkhanizadeh, J. A. Wigenius, J. K. Duckworth, O. Inganas, and O. Hermanson. The promotion of neuronal maturation on soft substrates. Biomaterials 30:4567–4572, 2009.
Tempelman, L. A. Quantifying Receptor-Mediated Cell Adhesion Under Fluid Flow Using a Model Granulocyte Cell Line. Dissertation, Cornell Univeristy, New York, p. 328, 1993.
Tolic-Norrelykke, I. M., J. P. Butler, J. Chen, and N. Wang. Spatial and temporal traction response in human airway smooth muscle cells. Am. J. Physiol. Cell Physiol. 283:C1254–C1266, 2002.
Tolic-Norrelykke, I. M., and N. Wang. Traction in smooth muscle cells varies with cell spreading. J. Biomech. 38:1405–1412, 2005.
Vogel, V. Mechanotransduction involving multimodular proteins: converting force into biochemical signals. Ann. Rev. Biophys. Biomol. Struct. 35:459–488, 2006.
Wang, Y.-L., R. J. Pelham, and B. V. Richard. Preparation of a flexible, porous polyacrylamide substrate for mechanical studies of cultured cells. Methods Enzymol. 298:489–496, 1998.
Yang, Z., J. S. Lin, J. Chen, and J. H. C. Wang. Determining substrate displacement and cell traction fields—a new approach. J. Theor. Biol. 242:607–616, 2006.
This work was supported by the Reid T. Milner Professorship to DEL and CM was supported by 5 P01 HL060678. The authors have no financial interests in this work.
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Mann, C., Leckband, D. Measuring Traction Forces in Long-Term Cell Cultures. Cel. Mol. Bioeng. 3, 40–49 (2010). https://doi.org/10.1007/s12195-010-0108-0
- Traction force microscopy
- Polyacrylamide hydrogel
- Cell contractility