Annals of Biomedical Engineering

, Volume 40, Issue 5, pp 1061–1072 | Cite as

Fibroblast Morphology on Dynamic Softening of Hydrogels

  • Michelle L. Previtera
  • Kevin L. Trout
  • Devendra Verma
  • Uday Chippada
  • Rene S. Schloss
  • Noshir A. LangranaEmail author


Despite cellular environments having dynamic characteristics, many laboratories utilized static polyacrylamide hydrogels to study the ECM–cell relationship. To attain a more in vivo like environment, we have developed a dynamic, DNA-crosslinked hydrogel (DNA gel). Through the controlled delivery of DNA, we can temporally decrease or increase gel stiffness while expanding or contracting the gel, respectively. These dual mechanical changes make DNA gels a cell–ECM model for studying dynamic mechano-regulated processes, such as wound healing. Here, we characterized DNA gels on a mechanical and cellular level. In contrast to our previous publication, in which we examined the increasing stiffness effects on fibroblast morphology, we examined the effects of decreased matrix stiffness on fibroblast morphology. In addition, we quantified the bulk and/or local stress and strain properties of dynamic gels. Gels generated about 0.5 Pa stress and about 6–11% strain upon softening to generate larger and more circular fibroblasts. These results complemented our previous study, where dynamic gels contracted upon stiffening to generate smaller and longer fibroblasts. In conclusion, we developed a biomaterial that increases and decreases in stiffness while contracting and expanding, respectively. We found that the dynamic deformation directionality of the matrix determined the fibroblast morphology and possibly influences function.


Stress Strain ECM Compliance Stiffness DNA gel Wound healing 



This work was supported by NJSCR #08-3080-SCR-E-0. K.T. supported by Rutgers RiSE summer program, Rutgers REU-CB (NSF EEC-0851831), and The College of St. Scholastica Ronald E. McNair Postbaccalaureate Achievement Program. We would like to thank Christopher Liu for analysis of cell density and editing of this manuscript. We would like to thank Dr. David I. Shreiber for the GFP fibroblasts. Lastly, we would like to thank Namrata Kulkarni and Vikas Shah for editing this manuscript.

Conflict of interest

Authors have no conflict of interest.

Supplementary material

10439_2011_483_MOESM1_ESM.tif (79 kb)
Supplemental Figure 1: Aspect ratio and perimeter. Mean + SEM values of aspect ratio (left) and perimeter (right) for fibroblasts grown on various DNA gels (TIFF 79 kb)


  1. 1.
    Aarabi, S., K. A. Bhatt, Y. Shi, J. Paterno, E. I. Chang, S. A. Loh, J. W. Holmes, M. T. Longaker, H. Yee, and G. C. Gurtner. Mechanical load initiates hypertrophic scar formation through decreased cellular apoptosis. FASEB J. 21:3250, 2007.PubMedCrossRefGoogle Scholar
  2. 2.
    Balaban, N. Q., U. S. Schwarz, D. Riveline, P. Goichberg, G. Tzur, I. Sabanay, D. Mahalu, S. Safran, A. Bershadsky, L. Addadi, and B. Geiger. Force and focal adhesion assembly: a close relationship studied using elastic micropatterned substrates. Nat. Cell Biol. 3:466–472, 2001.PubMedCrossRefGoogle Scholar
  3. 3.
    Banerjee, I., K. Yekkala, T. K. Borg, and T. A. Baudino. Dynamic interactions between myocytes, fibroblasts, and extracellular matrix. Ann. N. Y. Acad. Sci. 1080:76–84, 2006.PubMedCrossRefGoogle Scholar
  4. 4.
    Brown, X. Q., K. Ookawa, and J. Y. Wong. Evaluation of polydimethylsiloxane scaffolds with physiologically-relevant elastic moduli: interplay of substrate mechanics and surface chemistry effects on vascular smooth muscle cell response. Biomaterials 26:3123–3129, 2005.PubMedCrossRefGoogle Scholar
  5. 5.
    Chippada, U. Non-Intrusive Characterization of Properties of Hydrogels. Rutgers University Electronic Theses and Dissertations, 2010.Google Scholar
  6. 6.
    Chippada, U., B. Yurke, and N. A. Langrana. Simultaneous determination of young’s modulus, shear modulus, and Poisson’s ratio of soft hydrogels. J. Mater. Res. 25:545–555, 2010.CrossRefGoogle Scholar
  7. 7.
    Cukierman, E., R. Pankov, D. R. Stevens, and K. M. Yamada. Taking cell–matrix adhesions to the third dimension. Science. 294:1708–1712, 2001.PubMedCrossRefGoogle Scholar
  8. 8.
    Dembo, M., and Y. L. Wang. Stresses at the cell-to-substrate interface during locomotion of fibroblasts. Biophys. J. 76:2307–2316, 1999.PubMedCrossRefGoogle Scholar
  9. 9.
    Eastwood, M., D. A. McGrouther, and R. A. Brown. Fibroblast responses to mechanical forces. Proc. Inst. Mech. Eng. H: J. Eng. Med. 212:85–92, 1998.CrossRefGoogle Scholar
  10. 10.
    Engler, A., L. Bacakova, C. Newman, A. Hategan, M. Griffin, and D. Discher. Substrate compliance versus ligand density in cell on gel responses. Biophys. J. 86:617–628, 2004.PubMedCrossRefGoogle Scholar
  11. 11.
    Flanagan, L. A., Y. E. Ju, B. Marg, M. Osterfield, and P. A. Janmey. Neurite branching on deformable substrates. Neuroreport. 13:2411, 2002.PubMedCrossRefGoogle Scholar
  12. 12.
    Frank, C., D. McDonald, J. Wilson, D. Eyre, and N. Shrive. Rabbit medial collateral ligament scar weakness is associated with decreased collagen pyridinoline crosslink density. J. Orthop. Res. 13:157–165, 1995.PubMedCrossRefGoogle Scholar
  13. 13.
    Friedland, J. C., M. H. Lee, and D. Boettiger. Mechanically activated integrin switch controls alpha(5)beta(1) function. Science. 323:642–644, 2009.PubMedCrossRefGoogle Scholar
  14. 14.
    Gabbiani, G. The myofibroblast in wound healing and fibrocontractive diseases. J. Pathol. 200:500–503, 2003.PubMedCrossRefGoogle Scholar
  15. 15.
    Galbraith, C. G., and M. P. Sheetz. A micromachined device provides a new bend on fibroblast traction forces. Proc. Natl Acad. Sci. USA 94:9114–9118, 1997.PubMedCrossRefGoogle Scholar
  16. 16.
    Galbraith, C. G., and M. P. Sheetz. Forces on adhesive contacts affect cell function. Curr. Opin. Cell Biol. 10:566–571, 1998.PubMedCrossRefGoogle Scholar
  17. 17.
    Galbraith, C. G., R. Skalak, and S. Chien. Shear stress induces spatial reorganization of the endothelial cell cytoskeleton. Cell Motil. Cytoskelet. 40:317–330, 1998.CrossRefGoogle Scholar
  18. 18.
    Geiger, B., and A. Bershadsky. Exploring the neighborhood: adhesion-coupled cell mechanosensors. Cell 110:139–142, 2002.PubMedCrossRefGoogle Scholar
  19. 19.
    Genes, N. G., J. A. Rowley, D. J. Mooney, and L. J. Bonassar. Effect of substrate mechanics on chondrocyte adhesion to modified alginate surfaces. Arch. Biochem. Biophys. 422:161–167, 2004.PubMedCrossRefGoogle Scholar
  20. 20.
    Haston, W. S., J. M. Shields, and P. C. Wilkinson. The orientation of fibroblasts and neutrophils on elastic substrata. Exp. Cell Res. 146:117–126, 1983.PubMedCrossRefGoogle Scholar
  21. 21.
    Huang, S., and D. E. Ingber. The structural and mechanical complexity of cell-growth control. Nat. Cell Biol. 1:E131–E138, 1999.PubMedCrossRefGoogle Scholar
  22. 22.
    Hynes, R. O. The dynamic dialogue between cells and matrices: implications of fibronectin’s elasticity. Proc. Natl Acad. Sci. USA 96:2588–2590, 1999.PubMedCrossRefGoogle Scholar
  23. 23.
    Ingber, D. E. Mechanosensation through integrins: cells act locally but think globally. Proc. Natl Acad. Sci. USA 100:1472–1474, 2003.PubMedCrossRefGoogle Scholar
  24. 24.
    Inoh, H., N. Ishiguro, S. Sawazaki, H. Amma, M. Miyazu, H. Iwata, M. Sokabe, and K. Naruse. Uni-axial cyclic stretch induces the activation of transcription factor nuclear factor kappab in human fibroblast cells. FASEB J. 16:405–407, 2002.PubMedGoogle Scholar
  25. 25.
    Jean, C., P. Gravelle, J. J. Fournie, and G. Laurent. Influence of stress on extracellular matrix and integrin biology. Oncogene. 30:2697–2706, 2011.PubMedCrossRefGoogle Scholar
  26. 26.
    Jiang, X., P. C. Georges, B. Li, Y. Du, M. K. Kutzing, M. L. Previtera, N. A. Langrana, and B. L. Firestein. Cell growth in response to mechanical stiffness is affected by neuron-astroglia interactions. Open Neurosci. J. 1:7–14, 2007.Google Scholar
  27. 27.
    Jiang, F. X., B. Yurke, B. L. Firestein, and N. A. Langrana. Neurite outgrowth on a DNA crosslinked hydrogel with tunable stiffnesses. Ann. Biomed. Eng. 36:1565–1579, 2008.PubMedCrossRefGoogle Scholar
  28. 28.
    Jiang, F. X., B. Yurke, R. S. Schloss, B. L. Firestein, and N. A. Langrana. Effect of dynamic stiffness of the substrates on neurite outgrowth by using a DNA-crosslinked hydrogel. Tissue Eng. A 16:1873–1889, 2010.CrossRefGoogle Scholar
  29. 29.
    Jiang, F. X., B. Yurke, R. S. Schloss, B. L. Firestein, and N. A. Langrana. The relationship between fibroblast growth and the dynamic stiffnesses of a DNA crosslinked hydrogel. Biomaterials 31:1199–1212, 2010.PubMedCrossRefGoogle Scholar
  30. 30.
    Kano, Y., K. Katoh, and K. Fujiwara. Lateral zone of cell–cell adhesion as the major fluid shear stress-related signal transduction site. Circ. Res. 86:425–433, 2000.PubMedGoogle Scholar
  31. 31.
    Kolodney, M. S., and R. B. Wysolmerski. Isometric contraction by fibroblasts and endothelial cells in tissue culture: a quantitative study. J. Cell Biol. 117:73–82, 1992.PubMedCrossRefGoogle Scholar
  32. 32.
    Kuo, S. C., and M. P. Sheetz. Forces of single kinesin molecules measured with optical tweezers. Science 260:232–234, 1993.PubMedCrossRefGoogle Scholar
  33. 33.
    Lauffenburger, D. A., and A. F. Horwitz. Cell migration: a physically integrated molecular process. Cell 84:359–369, 1996.PubMedCrossRefGoogle Scholar
  34. 34.
    Letourneau, P. C., M. L. Condic, and D. M. Snow. Interactions of developing neurons with the extracellular matrix. J. Neurosci. 14:915, 1994.PubMedGoogle Scholar
  35. 35.
    Li, L., A. E. Davidovich, J. M. Schloss, U. Chippada, R. R. Schloss, N. A. Langrana, and M. L. Yarmush. Neural lineage differentiation of embryonic stem cells within alginate microbeads. Biomaterials 32:4489–4497, 2011.PubMedCrossRefGoogle Scholar
  36. 36.
    Lin, D. C., B. Yurke, and N. A. Langrana. Mechanical properties of a reversible, DNA-crosslinked polyacrylamide hydrogel. J. Biomech. Eng. 126:104, 2004.PubMedCrossRefGoogle Scholar
  37. 37.
    Lin, D. C., B. Yurke, and N. A. Langrana. Inducing reversible stiffness changes in DNA-crosslinked gels. J. Mater. Res. 20:1456–1464, 2005.CrossRefGoogle Scholar
  38. 38.
    Lin, D. C., B. Yurke, and N. A. Langrana. Use of rigid spherical inclusions in young’s moduli determination: application to DNA-crosslinked gels. J. Biomech. Eng. 127:571, 2005.PubMedCrossRefGoogle Scholar
  39. 39.
    Lo, C. M., H. B. Wang, M. Dembo, and Y. L. Wang. Cell movement is guided by the rigidity of the substrate. Biophys. J. 79:144–152, 2000.PubMedCrossRefGoogle Scholar
  40. 40.
    MacKenna, D., S. R. Summerour, and F. J. Villarreal. Role of mechanical factors in modulating cardiac fibroblast function and extracellular matrix synthesis. Cardiovasc. Res. 46:257–263, 2000.PubMedCrossRefGoogle Scholar
  41. 41.
    Neidlinger-Wilke, C., E. S. Grood, J.-C. Wang, R. A. Brand, and L. Claes. Cell alignment is induced by cyclic changes in cell length: studies of cells grown in cyclically stretched substrates. J. Orthop. Res. 19:286–293, 2001.PubMedCrossRefGoogle Scholar
  42. 42.
    Pelham, R. J., Jr., and Y. Wang. Cell locomotion and focal adhesions are regulated by substrate flexibility. Proc. Natl Acad. Sci. USA 94:13661, 1997.PubMedCrossRefGoogle Scholar
  43. 43.
    Polio, S. R., K. E. Rothenberg, D. Stamenović, and M. L. Smith. A micropatterning and image processing approach to simplify measurement of cellular traction force. Acta Biomater. 8:82–88, 2012.PubMedCrossRefGoogle Scholar
  44. 44.
    Previtera, M. L., C. G. Langhammer, and B. L. Firestein. Effects of substrate stiffness and cell density on primary hippocampal cultures. J. Biosci. Bioeng. 110:459–470, 2010.PubMedCrossRefGoogle Scholar
  45. 45.
    Previtera, M. L., C. G. Langhammer, N. A. Langrana, and B. L. Firestein. Dendrite arborization by substrate stiffness is mediated by glutamate receptor. Ann. Biomed. Eng. 38:3733–3743, 2010.PubMedCrossRefGoogle Scholar
  46. 46.
    Raghow, R. The role of extracellular matrix in postinflammatory wound healing and fibrosis. FASEB J. 8:823, 1994.PubMedGoogle Scholar
  47. 47.
    Sheetz, M. P., D. P. Felsenfeld, and C. G. Galbraith. Cell migration: regulation of force on extracellular-matrix-integrin complexes. Trends Cell Biol. 8:51–54, 1998.PubMedCrossRefGoogle Scholar
  48. 48.
    Singer, A. J., and R. A. F. Clark. Cutaneous wound healing. N. Engl. J. Med. 341:738, 1999.PubMedCrossRefGoogle Scholar
  49. 49.
    Skutek, M., M. van Griensven, J. Zeichen, N. Brauer, and U. Bosch. Cyclic mechanical stretching modulates secretion pattern of growth factors in human tendon fibroblasts. Eur. J. Appl. Physiol. 86:48–52, 2001.PubMedCrossRefGoogle Scholar
  50. 50.
    Solon, J., I. Levental, K. Sengupta, P. C. Georges, and P. A. Janmey. Fibroblast adaptation and stiffness matching to soft elastic substrates. Biophys. J. 93:4453–4461, 2007.PubMedCrossRefGoogle Scholar
  51. 51.
    Sundararaghavan, H. G., G. A. Monteiro, B. L. Firestein, and D. I. Shreiber. Neurite growth in 3D collagen gels with gradients of mechanical properties. Biotechnol. Bioeng. 102:632–643, 2009.PubMedCrossRefGoogle Scholar
  52. 52.
    Thibeault, S. L., S. D. Gray, D. M. Bless, R. W. Chan, and C. N. Ford. Histologic and rheologic characterization of vocal fold scarring. J. Voice 16:96–104, 2002.PubMedCrossRefGoogle Scholar
  53. 53.
    Tomasek, J. J., G. Gabbiani, B. Hinz, C. Chaponnier, and R. A. Brown. Myofibroblasts and mechano-regulation of connective tissue remodelling. Nat. Rev. Mol. Cell Biol. 3:349–363, 2002.PubMedCrossRefGoogle Scholar
  54. 54.
    Wakatsuki, T., M. S. Kolodney, G. I. Zahalak, and E. L. Elson. Cell mechanics studied by a reconstituted model tissue. Biophys. J. 79:2353–2368, 2000.PubMedCrossRefGoogle Scholar
  55. 55.
    Wang, N., J. P. Butler, and D. E. Ingber. Mechanotransduction across the cell-surface and through the cytoskeleton. Science. 260:1124–1127, 1993.PubMedCrossRefGoogle Scholar
  56. 56.
    Wang, H. B., M. Dembo, S. K. Hanks, and Y. Wang. Focal adhesion kinase is involved in mechanosensing during fibroblast migration. Proc. Natl Acad. Sci. USA 98:11295, 2001.PubMedCrossRefGoogle Scholar
  57. 57.
    Yeung, T., P. C. Georges, L. A. Flanagan, B. Marg, M. Ortiz, M. Funaki, N. Zahir, W. Ming, V. Weaver, and P. A. Janmey. Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesion. Cell Motil. Cytoskelet. 60:24–34, 2005.CrossRefGoogle Scholar
  58. 58.
    Zaman, M. H., L. M. Trapani, A. L. Sieminski, D. MacKellar, H. Gong, R. D. Kamm, A. Wells, D. A. Lauffenburger, and P. Matsudaira. Migration of tumor cells in 3D matrices is governed by matrix stiffness along with cell–matrix adhesion and proteolysis. Proc. Natl Acad. Sci. USA 103:10889, 2006.PubMedCrossRefGoogle Scholar

Copyright information

© Biomedical Engineering Society 2011

Authors and Affiliations

  • Michelle L. Previtera
    • 1
  • Kevin L. Trout
    • 2
  • Devendra Verma
    • 1
  • Uday Chippada
    • 3
  • Rene S. Schloss
    • 1
  • Noshir A. Langrana
    • 1
    • 3
    Email author
  1. 1.Department of Biomedical EngineeringRutgers UniversityPiscatawayUSA
  2. 2.Department of Chemistry and BiochemistryThe College of St. ScholasticaDuluthUSA
  3. 3.Department of Mechanical Engineering and AerospaceRutgers UniversityPiscatawayUSA

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