Cell structure depends on both matrix strain and stiffness, but their interactive effects are poorly understood. We investigated the interactive roles of matrix properties and stretching patterns on cell structure by uniaxially stretching U2OS cells expressing GFP-actin on silicone rubber sheets supporting either a surface-adsorbed coating or thick hydrogel of type-I collagen. Cells and their actin stress fibers oriented perpendicular to the direction of cyclic stretch on collagen-coated sheets, but oriented parallel to the stretch direction on collagen gels. There was significant alignment parallel to the direction of a steady increase in stretch for cells on collagen gels, while cells on collagen-coated sheets did not align in any direction. The extent of alignment was dependent on both strain rate and duration. Stretch-induced alignment on collagen gels was blocked by the myosin light-chain kinase inhibitor ML7, but not by the Rho-kinase inhibitor Y27632. We propose that active orientation of the actin cytoskeleton perpendicular and parallel to direction of stretch on stiff and soft substrates, respectively, are responses that tend to maintain intracellular tension at an optimal level. Further, our results indicate that cells can align along directions of matrix stress without collagen fibril alignment, indicating that matrix stress can directly regulate cell morphology.
References
[1]
Hsu HJ, Lee CF, Locke A, Vanderzyl SQ, Kaunas R (2010) Stretch-induced stress fiber remodeling and the activations of JNK and ERK depend on mechanical strain rate, but not FAK. PLoS One 5: e12470. doi: 10.1371/journal.pone.0012470
[2]
Dartsch PC, Betz E (1989) Response of cultured endothelial cells to mechanical stimulation. Basic Res Cardiol 84: 268–281. doi: 10.1007/bf01907974
[3]
Kanda K, Matsuda T, Oka T (1992) Two-dimensional orientational response of smooth muscle cells to cyclic stretching. ASAIO J 38: M382–385. doi: 10.1097/00002480-199207000-00060
[4]
Tondon A, Hsu HJ, Kaunas R (2012) Dependence of cyclic stretch-induced stress fiber reorientation on stretch waveform. J Biomech 45: 728–735. doi: 10.1016/j.jbiomech.2011.11.012
[5]
Lee CF, Haase C, Deguchi S, Kaunas R (2010) Cyclic stretch-induced stress fiber dynamics - dependence on strain rate, Rho-kinase and MLCK. Biochem Biophys Res Commun 401: 344–349. doi: 10.1016/j.bbrc.2010.09.046
[6]
Jungbauer S, Gao H, Spatz JP, Kemkemer R (2008) Two characteristic regimes in frequency-dependent dynamic reorientation of fibroblasts on cyclically stretched substrates. Biophys J 95: 3470–3478. doi: 10.1529/biophysj.107.128611
[7]
Chicurel ME, Chen CS, Ingber DE (1998) Cellular control lies in the balance of forces. Curr Opin Cell Biol 10: 232–239. doi: 10.1016/s0955-0674(98)80145-2
[8]
Kaunas R, Deguchi S (2011) Sarcomeric model of stretch-induced stress fiber reorganization Cell Health and Cytoskeleton 3 13–22.
[9]
Hsu HJ, Lee CF, Kaunas R (2009) A dynamic stochastic model of frequency-dependent stress fiber alignment induced by cyclic stretch. PLoS One 4: e4853. doi: 10.1371/journal.pone.0004853
[10]
Wang N, Tolic-Norrelykke IM, Chen J, Mijailovich SM, Butler JP, et al. (2002) Cell prestress. I. Stiffness and prestress are closely associated in adherent contractile cells. Am J Physiol Cell Physiol 282: C606–616. doi: 10.1152/ajpcell.00269.2001
[11]
Reinhart-King CA (2008) Endothelial cell adhesion and migration. Methods Enzymol 443: 45–64. doi: 10.1016/s0076-6879(08)02003-x
[12]
Reinhart-King CA, Dembo M, Hammer DA (2008) Cell-cell mechanical communication through compliant substrates. Biophys J 95: 6044–6051. doi: 10.1529/biophysj.107.127662
Yeung T, Georges PC, Flanagan LA, Marg B, Ortiz M, et al. (2005) Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesion. Cell Motil Cytoskeleton 60: 24–34. doi: 10.1002/cm.20041
[15]
Buxboim A, Rajagopal K, Brown AE, Discher DE (2010) How deeply cells feel: methods for thin gels. J Phys Condens Matter 22: 194116. doi: 10.1088/0953-8984/22/19/194116
[16]
Sen S, Engler AJ, Discher DE (2009) Matrix strains induced by cells: Computing how far cells can feel. Cell Mol Bioeng 2: 39–48. doi: 10.1007/s12195-009-0052-z
[17]
Gavara N, Roca-Cusachs P, Sunyer R, Farre R, Navajas D (2008) Mapping cell-matrix stresses during stretch reveals inelastic reorganization of the cytoskeleton. Biophys J 95: 464–471. doi: 10.1529/biophysj.107.124180
[18]
Throm Quinlan AM, Sierad LN, Capulli AK, Firstenberg LE, Billiar KL (2011) Combining dynamic stretch and tunable stiffness to probe cell mechanobiology in vitro. PLoS One 6: e23272. doi: 10.1371/journal.pone.0023272
[19]
Kang H, Kwak HI, Kaunas R, Bayless KJ (2011) Fluid shear stress and sphingosine 1-phosphate activate calpain to promote membrane type 1 matrix metalloproteinase (MT1-MMP) membrane translocation and endothelial invasion into three-dimensional collagen matrices. J Biol Chem 286: 42017–42026. doi: 10.1074/jbc.m111.290841
[20]
Fung YC (1994) A First Course in Continuum Mechanics for Physical and Biological Engineers and Scientists. Prentice-hall.
[21]
Kaunas R, Nguyen P, Usami S, Chien S (2005) Cooperative effects of Rho and mechanical stretch on stress fiber organization. Proc Natl Acad Sci U S A 102: 15895–15900. doi: 10.1073/pnas.0506041102
[22]
Xu F, Beyazoglu T, Hefner E, Gurkan UA, Demirci U (2011) Automated and adaptable quantification of cellular alignment from microscopic images for tissue engineering applications. Tissue Eng Part C Methods 17: 641–649. doi: 10.1089/ten.tec.2011.0038
[23]
Girton TS, Barocas VH, Tranquillo RT (2002) Confined compression of a tissue-equivalent: collagen fibril and cell alignment in response to anisotropic strain. J Biomech Eng 124: 568–575. doi: 10.1115/1.1504099
[24]
Vader D, Kabla A, Weitz D, Mahadevan L (2009) Strain-induced alignment in collagen gels. PLoS One 4: e5902. doi: 10.1371/journal.pone.0005902
[25]
Saez A, Ghibaudo M, Buguin A, Silberzan P, Ladoux B (2007) Rigidity-driven growth and migration of epithelial cells on microstructured anisotropic substrates. Proc Natl Acad Sci U S A 104: 8281–8286. doi: 10.1073/pnas.0702259104
[26]
Schwarz US, Erdmann T, Bischofs IB (2006) Focal adhesions as mechanosensors: the two-spring model. Biosystems 83: 225–232. doi: 10.1016/j.biosystems.2005.05.019
[27]
Zemel A, Rehfeldt F, Brown AE, Discher DE, Safran SA (2010) Optimal matrix rigidity for stress fiber polarization in stem cells. Nat Phys 6: 468–473. doi: 10.1038/nphys1613
[28]
Roby T, Olsen S, Nagatomi J (2008) Effect of sustained tension on bladder smooth muscle cells in three-dimensional culture. Ann Biomed Eng 36: 1744–1751. doi: 10.1007/s10439-008-9545-5
[29]
Guilak F, Cohen DM, Estes BT, Gimble JM, Liedtke W, et al. (2009) Control of stem cell fate by physical interactions with the extracellular matrix. Cell Stem Cell 5: 17–26. doi: 10.1016/j.stem.2009.06.016
[30]
Zhang W, Choi DS, Nguyen YH, Chang J, Qin L (2013) Studying cancer stem cell dynamics on PDMS surfaces for microfluidics device design. Sci Rep 3: 2332. doi: 10.1038/srep02332
[31]
Pang Y, Wang X, Lee D, Greisler HP (2011) Dynamic quantitative visualization of single cell alignment and migration and matrix remodeling in 3-D collagen hydrogels under mechanical force. Biomaterials 32: 3776–3783. doi: 10.1016/j.biomaterials.2011.02.003
[32]
Bellows CG, Melcher AH, Aubin JE (1982) Association between tension and orientation of periodontal ligament fibroblasts and exogenous collagen fibres in collagen gels in vitro. J Cell Sci 58: 125–138.
[33]
Tanner K, Boudreau A, Bissell MJ, Kumar S (2010) Dissecting regional variations in stress fiber mechanics in living cells with laser nanosurgery. Biophys J 99: 2775–2783. doi: 10.1016/j.bpj.2010.08.071
[34]
Katoh K, Kano Y, Amano M, Kaibuchi K, Fujiwara K (2001) Stress fiber organization regulated by MLCK and Rho-kinase in cultured human fibroblasts. Am J Physiol Cell Physiol 280: C1669–1679.
[35]
Ronan W, Deshpande VS, McMeeking RM, McGarry JP (2013) Cellular contractility and substrate elasticity: a numerical investigation of the actin cytoskeleton and cell adhesion. Biomech Model Mechanobiol.
[36]
Lee SL, Nekouzadeh A, Butler B, Pryse KM, McConnaughey WB, et al. (2012) Physically-induced cytoskeleton remodeling of cells in three-dimensional culture. PLoS One 7: e45512. doi: 10.1371/journal.pone.0045512
[37]
Krishnan R, Park CY, Lin YC, Mead J, Jaspers RT, et al. (2009) Reinforcement versus fluidization in cytoskeletal mechanoresponsiveness. PLoS One 4: e5486. doi: 10.1371/journal.pone.0005486
[38]
Deshpande VS, McMeeking RM, Evans AG (2006) A bio-chemo-mechanical model for cell contractility. Proc Natl Acad Sci U S A 103: 14015–14020. doi: 10.1073/pnas.0605837103
[39]
Trepat X, Deng L, An SS, Navajas D, Tschumperlin DJ, et al. (2007) Universal physical responses to stretch in the living cell. Nature 447: 592–595. doi: 10.1038/nature05824
[40]
Chen C, Krishnan R, Zhou E, Ramachandran A, Tambe D, et al. (2010) Fluidization and resolidification of the human bladder smooth muscle cell in response to transient stretch. PLoS One 5: e12035. doi: 10.1371/journal.pone.0012035
[41]
Faust U, Hampe N, Rubner W, Kirchgessner N, Safran S, et al. (2011) Cyclic stress at mHz frequencies aligns fibroblasts in direction of zero strain. PLoS One 6: e28963. doi: 10.1371/journal.pone.0028963
[42]
Plotnikov SV, Pasapera AM, Sabass B, Waterman CM (2012) Force Fluctuations within Focal Adhesions Mediate ECM-Rigidity Sensing to Guide Directed Cell Migration. Cell 151: 1513–1527. doi: 10.1016/j.cell.2012.11.034
[43]
Trichet L, Le Digabel J, Hawkins RJ, Vedula SR, Gupta M, et al. (2012) Evidence of a large-scale mechanosensing mechanism for cellular adaptation to substrate stiffness. Proc Natl Acad Sci U S A 109: 6933–6938. doi: 10.1073/pnas.1117810109
