Dielectrophoresis (DEP) is the phenomenon in which a particle, such as a living cell, is polarized and moved by electrical gravity in a non-uniform electric field. In the present study, the DEP force is utilized to act on the cells to induce spatial movement for investigating the correlation between the cell adhesion force and activation level of focal adhesion kinase (FAK). The DEP force produced by the non-uniform electric field was used to measure the cell adhesion force of ECV304 cells, on type 1 collagen (COL1)- and fibronectin (FN)-coated polydimethylsiloxane (PDMS) membranes. For COL1-coating, ECV304 cells revealed weak and variable adhesion force (0.343–0.760 nN) in the first eight hours of incubation. Interestingly, the cell adhesion force of ECV304 at two and five hours of cultivation was significantly high and matched their FAK activation level. In comparison, ECV304 on FN-coated membrane had higher and more stable cell adhesion force (0.577–2.053 nN). FN coating intensified the cell adhesion force of ECV304 with culture time and similar outcome was present on the activation level of FAK. Therefore, this study demonstrated a relationship between cell adhesion force and FAK activation level that was dependant on the choice of the extracellular matrix (ECM) component. Subsequently, two tyrosine kinase inhibitors (AG18 and genistein) and one PI3K inhibitor (LY294002) were applied to study the influence of protein phosphorylation on the cell adhesion force. FAK plays an important role on cell attachment and DEP force measurement is a useful technique for studying cell adhesion.
References
[1]
Rosso, F.; Giordano, A.; Barbarisi, M.; Barbarisi, A. From cell-ECM interactions to tissue engineering. J. Cell. Physiol. 2004, 199, 174–180, doi:10.1002/jcp.10471. 15039999
[2]
Badylak, S.F. The extracellular matrix as a biologic scaffold material. Biomaterials 2007, 28, 3587–3593, doi:10.1016/j.biomaterials.2007.04.043. 17524477
[3]
Sage, E.H. Regulation of interactions between cells and extracellular matrix: A command performance on several stages. J. Clin. Invest. 2001, 107, 781–783, doi:10.1172/JCI12683. 11285292
[4]
Zamir, E.; Geiger, B. Molecular complexity and dynamics of cell-matrix adhesions. J. Cell Sci. 2001, 114, 3583–3590. 11707510
[5]
Plopper, G.E.; McNamee, H.P.; Dike, L.E.; Bojanowski, K.; Ingber, D.E. Convergence of integrin and growth factor receptor signaling pathways within the focal adhesion complex. Mol. Biol. Cell 1995, 6, 1349–1365. 8573791
[6]
Richardson, A.; Parsons, J.T. A mechanism for regulation of the adhesion-associated protein tyrosine kinase pp125FAK. Nature 1996, 380, 538–540, doi:10.1038/380538a0. 8606775
[7]
Gilmore, A.P.; Romer, L.H. Inhibition of focal adhesion kinase (FAK) signaling in focal adhesions decreases cell motility and proliferation. Mol. Biol. Cell 1996, 7, 1209–1224. 8856665
Thoumine, O.; Ott, A.; Louvard, D. Critical centrifugal forces induce adhesion rupture or structural reorganization in cultured cells. Cell Motil. Cytoskelet. 1996, 33, 276–287, doi:10.1002/(SICI)1097-0169(1996)33:4<276::AID-CM4>3.0.CO;2-7.
[10]
Thoumine, O.; Ott, A. Comparison of the mechanical properties of normal and transformed fibroblasts. Biorheology 1997, 34, 309–326, doi:10.1016/S0006-355X(98)00007-9. 9578806
[11]
Truskey, G.A.; Proulx, T.L. Relationship between 3T3 cell spreading and the strength of adhesion on glass and silane surfaces. Biomaterials 1993, 14, 243–254, doi:10.1016/0142-9612(93)90114-H. 7682850
[12]
Leonenko, Z.; Finot, E.; Amrein, M. Adhesive interaction measured between AFM probe and lung epithelial type II cells. Ultramicroscopy 2007, 107, 948–953, doi:10.1016/j.ultramic.2007.02.036. 17561346
[13]
Zhang, H.; Liu, K.-K. Optical tweezers for single cells. J. R. Soc. Interface 2008, 5, 671–690, doi:10.1098/rsif.2008.0052. 18381254
[14]
Rico, F.; Roca-Cusachs, P.; Sunyer, R.; Farré, R.; Navajas, D. Cell dynamic adhesion and elastic properties probed with cylindrical atomic force microscopy cantilever tips. J. Mol. Recogn. 2007, 20, 459–466, doi:10.1002/jmr.829.
[15]
Shen, Y.; Nakajima, M.; Ahmad, M.R.; Kojima, S.; Homma, M.; Fukuda, T. Effect of ambient humidity on the strength of the adhesion force of single yeast cell inside environmental-SEM. Ultramicroscopy 2011, 111, 1176–1183, doi:10.1016/j.ultramic.2011.02.008. 21763235
[16]
Shen, Y.; Nakajima, M.; Kojima, S.; Michio Homma, M.; Fukuda, T. Study of the time effect on the strength of cell-cell adhesion force by a novel nano-picker. BBRC 2011, 409, 160–165. 21510921
[17]
Pethig, R. Dielectrophoresis: Using inhomogeneous AC electric fields separate and manipulate cells. Crit. Rev. Biotechnol. 1996, 16, 331–348, doi:10.3109/07388559609147425.
[18]
Lapizco-Encinas, B.H.; Simmons, B.A.; Cummings, E.B.; Fintschenko, Y. Dielectrophoretic concentration and separation of live and dead bacteria in an array of Insulators. Anal. Chem. 2004, 76, 1571–1579, doi:10.1021/ac034804j. 15018553
[19]
Dalir, H.; Yanagida, Y.; Hatsuzawa, T. Probing DNA mechanical characteristics by dielectrophoresis. Sens. Actuat. B Chem. 2009, 136, 472–478, doi:10.1016/j.snb.2008.11.004.
[20]
Jones, T.B. Electromechanics of Particles, 2nd ed. ed.; Cambridge University Press: Cambridge, UK, 2005; pp. 34–62.
[21]
Morgan, H.; Green, N.G. AC Electrokinetics: Colloids and Nanoparticles; Research Studies Press Ltd.: Hertfordshire, UK, 2003; pp. 39–43.
[22]
Hung, M.S.; Chen, Y.W.; Lin, C.J.; Ay, C.; Chiou, C.P. Measurement of human endothelial cell adhesion using dielectrphoresis. J. Chin. Soc. Mech. Eng. 2009, 30, 393–400.
[23]
Chen, C.N.; Li, Y.S.; Yeh, Y.T.; Lee, P.L.; Usami, S.; Chien, S.; Chiu, J.J. Synergistic roles of platelet-derived growth factor-BB and interleukin-1beta in phenotypic modulation of human aortic smooth muscle cells. Proc. Natl. Acad. Sci. USA 2006, 103, 2665–2670, doi:10.1073/pnas.0510973103. 16477012
[24]
Yeh, C.C.; Chang, H.I.; Chiang, J.K.; Tsai, W.T.; Chen, L.M.; Wu, C.P.; Chien, S.; Chen, C.N. Regulation of plasminogen activator inhibitor 1 expression in human osteoarthritic chondrocytes by fluid shear Stress: Role of PKCα. Arthritis Rheum. 2009, 60, 2350–2361, doi:10.1002/art.24680. 19644850
[25]
Seger, R.; Biener, Y.; Feinstein, R.; Hanoch, T.; Gazit, A.; Zick, Y. Differential activation of mitogen-activated protein kinase and S6 kinase signaling pathways by 12-O-tetradecanoylphorbol-13-acetate (TPA) and insulin. Evidence for involvement of a TPA-stimulated protein-tyrosine kinase. J. Biol. Chem. 1995, 270, 28325–28330, doi:10.1074/jbc.270.47.28325. 7499332
[26]
Sawai, H.; Okada, Y.; Funahashi, H.; Matsuo, Y.; Takahashi, H.; Takeyama, H.; Manabe, T. Activation of focal adhesion kinase enhances the adhesion and invasion of pancreatic cancer cells via extracellular signal-regulated kinase-1/2 signaling pathway activation. Mol. Cancer 2005, 4, doi:10.1186/1476-4598-4-37.
[27]
Heinz, R.R.; Kao, S.C.; Cary, L.A.; Guan, J.L.; Lai, J.F. Requirement of phophatidylinositol 3-kinase in focal adhesion kinase-promoted cell migration. J. Biol. Chem. 1999, 274, 12361–12366, doi:10.1074/jbc.274.18.12361. 10212207
[28]
Garcia, A.J.; Huber, F.; Boettiger, D. Force required to break α5β1 integrin-fibronectin bonds in intact adherent cells is sensitive to integrin activation state. J. Biol. Chem. 1998, 273, 10988–10993, doi:10.1074/jbc.273.18.10988. 9556578
Elineni, K.K.; Gallant, N.D. Regulation of cell adhesion strength by peripheral focal adhesion distribution. Biophys. J. 2011, 101, 2903–2911, doi:10.1016/j.bpj.2011.11.013. 22208188
[31]
Weder, G.; V？r？s, J.; Giazzon, M.; Matthey, N.; Heinzelmann, H.; Liley, M. Measuring cell adhesion forces during the cell cycle by force spectroscopy. Biointerphases 2009, 4, 27–34, doi:10.1116/1.3139962. 20408720
[32]
Thie, M.; R？spel, R.; Dettmann, W.; Benoit, M.; Ludwig, M.; Gaub, H.E.; Denker, H.W. Interactions between trophoblast and uterine epithelium: Monitoring of adhesive forces. Hum. Reprod. 1998, 13, 3211–3219, doi:10.1093/humrep/13.11.3211. 9853883
[33]
Wang, H.B.; Dembo, M.; Hanks, S.K.; Wang, Y. Focal adhesion kinase is involved in mechanosensing during fibroblast migration. Proc. Natl. Acad. Sci. USA 2001, 98, 11295–11300, doi:10.1073/pnas.201201198. 11572981
[34]
von Sengbusch, A.; Gassmann, P.; Fisch, K.M.; Enns, A.; Nicolson, G.L.; Haier, J. Focal adhesion kinase regulates metastatic adhesion of carcinoma cells within liver sinusoids. Am. J. Pathol. 2005, 166, 585–596, doi:10.1016/S0002-9440(10)62280-8. 15681841
[35]
van Nimwegen, M.J.; Verkoeijen, S.; van Buren, L.; Burg, D.; van de water, B. requirement for focal adhesion kinase in the early phase of mammary adenocarcinoma lung metastasis formation. Cancer Res. 2005, 65, 4698–4706, doi:10.1158/0008-5472.CAN-04-4126. 15930288
