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Living cell study at the single-molecule and single-cell levels by atomic force microscopy

    Xiaoli Shi

    Beijing National Laboratory for Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, 2 Zhongguancun North First Street, 100190 Beijing, PR China

    ,
    Xuejie Zhang

    Beijing National Laboratory for Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, 2 Zhongguancun North First Street, 100190 Beijing, PR China

    ,
    Tie Xia

    Beijing National Laboratory for Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, 2 Zhongguancun North First Street, 100190 Beijing, PR China

    &
    Xiaohong Fang

    * Author for correspondence

    Beijing National Laboratory for Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, 2 Zhongguancun North First Street, 100190 Beijing, PR China.

    Published Online:https://doi.org/10.2217/nnm.12.130

    Atomic force microscopy (AFM) has been emerging as a multifunctional molecular tool in nanobiology and nanomedicine. This review summarizes the recent advances in AFM study of living mammalian cells at the single-molecule and single-cell levels. Besides nanoscale imaging of cell membrane structure, AFM-based force measurements on living cells are mainly discussed. These include the development and application of single-molecule force spectroscopy to investigate ligand–receptor binding strength and dissociation dynamics, and the characterization of cell mechanical properties in a physiological environment. Molecular manipulation of cells by AFM to change the cellular process is also described. Living-cell AFM study offers a new approach to understand the molecular mechanisms of cell function, disease development and drug effect, as well as to develop new strategies to achieve single-cell-based diagnosis.

    Papers of special note have been highlighted as: ▪ of interest ▪▪ of considerable interest

    References

    • Binnig G, Quate CF, Gerber C. Atomic force microscopy. Phys. Rev. Lett.56,930–933 (1986).Crossref, Medline, CASGoogle Scholar
    • Paige MF, Rainey JK, Goh MC. A study of fibrous long spacing collagen ultrastructure and assembly by atomic force microscopy. Micron32(3),341–353 (2001).Crossref, Medline, CASGoogle Scholar
    • Müller DJ, Dufrêne YF. Atomic force microscopy as a multifunctional molecular toolbox in nanobiotechnology. Nat Nanotechnol.3(5),261–269 (2008).▪▪ Comprehensively outlines the fascinating opportunities offered by the rapid advances in atomic force microscopy (AFM).Crossref, Medline, CASGoogle Scholar
    • Hinterdorfer P, Dufrêne YF. Detection and localization of single molecular recognition events using atomic force microscopy. Nat Methods3(5),347–355 (2006).Crossref, Medline, CASGoogle Scholar
    • Engel A, Gaub HE. Structure and mechanics of membrane proteins. Annu. Rev. Biochem.77,127–148 (2008).Crossref, Medline, CASGoogle Scholar
    • Müller DJ, Anderson K. Biomolecular imaging using atomic force microscopy. Trends Biotech.20(8),S45–S49 (2002).Crossref, MedlineGoogle Scholar
    • Alsteens D, Dupres V, Andre G, Dufrêne YF. Frontiers in microbial nanoscopy. Nanomedicine6(2),395–403 (2011).LinkGoogle Scholar
    • Vadillo-Rodriguez V, Dutcher JR. Viscoelasticity of the bacterial cell envelope. Soft Matter7,4101–4110 (2011).Crossref, CASGoogle Scholar
    • Bai C, Tian F, Luo K. Scanning Force Microscope. Science Press, Beijing, China (2000).▪▪ Systematically describes AFM techniques from its principle, experimental methods to application.Google Scholar
    • 10  Oberleithner H. Is the vascular endothelium under the control of aldosterone? Facts and hypothesis. Pflug. Arch. Eur. J. Phy.454(2),187–193 (2007).Crossref, Medline, CASGoogle Scholar
    • 11  Heu C, Berquand A, Elie-Caille C, Nicod L. Glyphosate-induced stiffening of HaCaT keratinocytes, a Peak Force Tapping study on living cells. J. Struc. Biol.178(1),1–7 (2012).Crossref, Medline, CASGoogle Scholar
    • 12  Oberleithner H, Riethmüller C, Ludwig T et al. Differential action of steroid hormones on human endothelium. J. Cell Sci.119(9),1926–1932 (2006).Crossref, Medline, CASGoogle Scholar
    • 13  Hillebrand U, Lang D, Telgmann RG et al. Nebivolol decreases endothelial cell stiffness via the estrogen receptor beta: a nano-imaging study. J. Hypertens.27(3),517–526 (2009).Crossref, Medline, CASGoogle Scholar
    • 14  Oberleithner H, Callies C, Kusche-Vihrog K et al. Potassium softens vascular endothelium and increases nitric oxide release. Proc. Natl Acad. Sci. USA106(8),2829–2834 (2009).Crossref, Medline, CASGoogle Scholar
    • 15  Oberleithner H, Riethmüller C, Ludwig T, Hausberg M, Schillers H. Aldosterone remodels human endothelium. Acta Physiol.187(1–2),305–312 (2006).Crossref, CASGoogle Scholar
    • 16  Städler B, Blättler TM, Franco-Obregón A. Time-lapse imaging of in vitro myogenesis using atomic force microscopy. J. Microsc.237(1),63–69 (2010).Crossref, Medline, CASGoogle Scholar
    • 17  Wang DC, Chen KY, Tsai CH, Chen GY, Chen CH. AFM membrane roughness as a probe to identify oxidative stress-induced cellular apoptosis. J. Biomech.44(16),2790–2794 (2011).Crossref, MedlineGoogle Scholar
    • 18  Prasanth R, Nair G, Girish CM. Enhanced endocytosis of nano-curcumin in nasopharyngeal cancer cells: an atomic force microscopy study. Appl. Phys. Lett.99,163706 (2011).CrossrefGoogle Scholar
    • 19  Friedrichs J, Taubenberger A, Franz CM, Muller DJ. Cellular remodelling of individual collagen fibrils visualized by time-lapse AFM. J. Mol. Biol.372(3),594–607 (2007).Crossref, Medline, CASGoogle Scholar
    • 20  Kim KS, Cho CH, Park EK et al. AFM-detected apoptotic changes in morphology and biophysical property caused by paclitaxel in Ishikawa and HeLa cells. PLoS One7(1),e30066 (2012).Crossref, Medline, CASGoogle Scholar
    • 21  Müller DJ, Helenius J, Alsteens D, Dufrêne YF. Force probing surfaces of living cells to molecular resolution. Nat. Chem. Biol.5(6),383–390 (2009).Crossref, Medline, CASGoogle Scholar
    • 22  Kuznetsova TG, Starodubtseva MN, Yegorenkov NI et al. Atomic force microscopy probing of cell elasticity. Micron38(8),824–833 (2007).Crossref, Medline, CASGoogle Scholar
    • 23  Butt HJ, Cappella B, Kappl M. Force measurements with the atomic force microscope: technique, interpretation and applications. Surf. Sci. Rep.59(1–6),1–152 (2005).Crossref, CASGoogle Scholar
    • 24  Kada G, Kienberger F, Hinterdorfer P. Atomic force microscopy in bionanotechnology. Nanotoday3(1–2),12–19 (2008).Crossref, Medline, CASGoogle Scholar
    • 25  Jiang Y, Fang X, Bai C et al. Specific aptamer-protein interaction studied by atomic force microscopy. Anal. Chem.75(9),2112–2116 (2003).Crossref, Medline, CASGoogle Scholar
    • 26  Lee CK, Wang YM, Huang LS, Lin S. Atomic force microscopy: determination of unbinding force, off rate and energy barrier for protein–ligand interaction. Micron38(5),446–461 (2007).Crossref, Medline, CASGoogle Scholar
    • 27  Helenius J, Heisenberg CP, Gaub HE, Muller DJ. Single-cell force spectroscopy. J. Cell Sci.121(11),1785–1791 (2008).▪▪ Describes current implementations of single-cell force spectroscopy in providing insights into the forces, energetics and kinetics of cell adhesion processes and discusses the potential pitfalls.Crossref, Medline, CASGoogle Scholar
    • 28  Moy VT, Florin EL, Gaub HE. Intermolecular forces and energies between ligands and receptors. Science266(5183),257–259 (1994).Crossref, Medline, CASGoogle Scholar
    • 29  Lee GU, Kidwell DA, Colton RJ. Sensing discrete streptavidin-biotin interactions with atomic force microscopy. Langmuir10(2),354–357 (1994).Crossref, CASGoogle Scholar
    • 30  Borgia A, Steward A, Clarke J. An effective strategy for the design of proteins with enhanced mechanical stability. Angew. Chem. Int. Ed. Engl.47(36),6900–6903 (2008).Crossref, Medline, CASGoogle Scholar
    • 31  Alegre-Cebollada J, Perez-Jimenez R, Kosuri P, Fernandez JM. Single-molecule force spectroscopy approach to enzyme catalysis. J. Biol. Chem.285(25),18961–18966 (2010).Crossref, Medline, CASGoogle Scholar
    • 32  Puchner EM, Gaub HE. Force and function: probing proteins with AFM-based force spectroscopy. Curr. Opin. Struct. Biol.19(5),605–614 (2009).Crossref, Medline, CASGoogle Scholar
    • 33  Rief M, Oesterhelt F, Heymann B, Gaub HE. Single molecule force spectroscopy on polysaccharides by AFM. Science275(5304),1295–1297 (1997).Crossref, Medline, CASGoogle Scholar
    • 34  Dufrêne YF, Evans E, Engel A, Helenius J, Gaub HE, Müller DJ. Five challenges to bringing single-molecule force spectroscopy into living cells. Nat. Methods8(2),123–127 (2011).▪▪ Discusses the problems and challenges that need to be addressed to bring single-molecule force spectroscopy (SMFS) into living cells and to learn how cellular machinery is controlled in vivo.Crossref, Medline, CASGoogle Scholar
    • 35  Morgan MR, Humphries MJ, Bass MD. Synergistic control of cell adhesion by integrins and syndecans. Nat. Rev. Mol. Cell Biol.8(12),957–969 (2007).Crossref, Medline, CASGoogle Scholar
    • 36  Benoit M, Gabriel D, Gerisch G, Gaub HE. Discrete interactions in cell adhesion measured by single-molecule force spectroscopy. Nat. Cell Biol.2(6),313–317 (2000).▪▪ The first AFM work that measured cell–cell interaction force with living cells.Crossref, Medline, CASGoogle Scholar
    • 37  Hanley W, McCarty O, Jadhav S et al. Single molecule characterization of P-selectin/ligand binding. J. Biol. Chem.278(12),10556–10561 (2003).Crossref, Medline, CASGoogle Scholar
    • 38  du Roure O, Buquin A, Feracci H, Silberzan P. Homophilic interactions between cadherin fragments at the single molecule level: an AFM study. Langmuir22(10),4680–4684 (2006).Crossref, Medline, CASGoogle Scholar
    • 39  Panorchan P, George JP, Wirtz D. Probing intercellular interactions between vascular endothelial cadherin pairs at single-molecule resolution and in living cells. J. Mol. Biol.358(3),665–674 (2006).Crossref, Medline, CASGoogle Scholar
    • 40  Yang H, Fang X, He C et al. Interaction between single molecules of Mac-1 and ICAM-1 in living cells: an atomic force microscopy study. Exp. Cell Res.313(16),3497–3504 (2007).▪▪ Demonstrated the use of SMFS to study the conformation changes of membrane protein.Crossref, Medline, CASGoogle Scholar
    • 41  Li Y, Shi X, Liu H, Yi S, Zhang X, Fang X. Study of the effect of atorvastatin on the interaction between ICAM-1 and CD11b by live-cell single-molecule force spectroscopy. Sci. China Chem.53(4),752–758 (2010).Crossref, CASGoogle Scholar
    • 42  Yu J, Wang Q, Fang X et al. Single-molecule force spectroscopy study of interaction between transforming growth factor β1 and its receptor in living cells. J. Phys. Chem.111(48),13619–13625 (2007).Crossref, CASGoogle Scholar
    • 43  Yang Y, Xu Y, Fang X et al. A single-molecule study of the inhibition effect of Naringenin on transforming growth factor-β ligand–receptor binding. Chem Comm.47,5440–5442 (2011).▪▪ Combined SMFS with theoretical simulation and single-molecule fluorescence imaging to investigate the inhibition mechanism of the small-molecule inhibitor of signal transduction.Crossref, Medline, CASGoogle Scholar
    • 44  Wildling L, Rankl C, Haselgrubler T et al. Probing binding pocket of serotonin transporter by single molecular force spectroscopy on living cells. J. Biol. Chem.287(1),105–113 (2012).Crossref, Medline, CASGoogle Scholar
    • 45  Shi X, Xu L, Fang X et al. Study of inhibition effect of herceptin on interaction between heregulin and ErbB receptors HER3/HER2 by single-molecule force spectroscopy. Exp. Cell Res.315(16),2487–2855 (2009).▪▪ First use of AFM SMFS to study the inhibition mechanism of a clinical anticancer drug.Crossref, MedlineGoogle Scholar
    • 46  Lee S, Mandic J, Van Vliet KJ. Chemomechanical mapping of ligand-receptor binding kinetics on cells. Proc. Natl Acad. Sci. USA104(23),9609–9614 (2007).Crossref, Medline, CASGoogle Scholar
    • 47  Roduit C, van der Goot FG, De Los Rios P et al. Elastic membrane heterogeneity of living cells revealed by stiff nanoscale membrane domains. Biophys. J.94(4),1521–1532 (2008).Crossref, Medline, CASGoogle Scholar
    • 48  Chtcheglova LA, Hinterdorfer P. Simultaneous topography and recognition imaging on endothelial cells. J. Mol. Recognit.24(5),788–794 (2011).▪▪ Describes the principles of simultaneous topography and recognition imaging and its recent applications in single-cell studies.Crossref, Medline, CASGoogle Scholar
    • 49  Chtcheglova LA, Waschke J, Wildling L, Drenckhahn D, Hinterdorfer P. Nano-scale dynamic recognition imaging on vascular endothelial cells. Biophys. J.93(2),L11–L13 (2007).Crossref, Medline, CASGoogle Scholar
    • 50  Pletikapic G, Berquand A, Radic TM, Svetlicic V. Microrheology of human lung epithelial cells measured by atomic force microscopy. J. Phycol.48,174–185 (2012).Crossref, MedlineGoogle Scholar
    • 51  Alcaraz J, Buscemi L, Grabulosa M et al. Microrheology of human lung epithelial cells measured by atomic force microscopy. Biophys. J.84,2071–2079 (2003).Crossref, Medline, CASGoogle Scholar
    • 52  Moreno-Flores S, Benitez R, Vivanco M, Toca-Herrera JL. Stress relaxation and creep on living cells with the atomic force microscope: a means to calculate elastic moduli and viscosities of cell components. Nanotechnology21(44),445101 (2010).Crossref, MedlineGoogle Scholar
    • 53  Liu F, Tschumperlin DJ. Micro-mechanical characterization of lung tissue using atomic force microscopy. J. Vis. Exp.54,1–7 (2011).Google Scholar
    • 54  Qiu HY, Zhu Y, Sun Z et al. Vascular smooth muscle cell stiffness as a mechanism for increased aortic stiffness with aging. Circ. Res.107,615–619 (2010).Crossref, Medline, CASGoogle Scholar
    • 55  Lulevich V, Zimmer CC, Hong HS, Jin LW, Liu GY. Single-cell mechanics provides a sensitive and quantitative means for probing amyloid-β peptide and neuronal cell interactions. Proc. Natl Acad. Sci. USA107(31),13872–13877 (2010).Crossref, Medline, CASGoogle Scholar
    • 56  Stolz M, Gottardi R, Raiteri R et al. Early detection of aging cartilage and osteoarthritis in mice and patient samples using atomic force microscopy. Nat. Nanotechnol.4(3),186–192 (2009).Crossref, Medline, CASGoogle Scholar
    • 57  Faria EC, Ma N, Gazi E et al. Measurement of elastic properties of prostate cancer cells using AFM. Analyst133(11),1498–1500 (2008).Crossref, Medline, CASGoogle Scholar
    • 58  Iyer S, Gaikwad RM, Subba-Rao V, Woodworth CD, Sokolov I. Atomic force microscopy detects differences in the surface brush of normal and cancerous cells. Nat. Nanotechnol.4(6),389–393 (2009).Crossref, Medline, CASGoogle Scholar
    • 59  Cross SE, Jin YS, Rao J, Gimzewski JK. Nanomechanical analysis of cells from cancer patients. Nat. Nanotechnol.2(12),780–783 (2007).▪▪ Demonstrated that AFM nanomechanical analysis correlated well with immunohistochemical testing currently used for detecting cancers.Crossref, Medline, CASGoogle Scholar
    • 60  Lekka M, Gil D, Pogoda K et al. Cancer cell detection in tissue sections using AFM. Arch. Biochem. Biophys.518(2),151–156 (2012).Crossref, Medline, CASGoogle Scholar
    • 61  Shi X, Qin L, Fang X et al. Elasticity of cardiac cells on the polymer substrates with different stiffness: an atomic force microscopy study. Phys. Chem. Chem. Phys.13(6),7540–7545 (2011).Crossref, Medline, CASGoogle Scholar
    • 62  Li QS, Lee GYH, Ong CN, Lim CT. AFM indentation study of breast cancer cells. Biochem. Biophys. Res. Commun.374(4),609–613 (2008).Crossref, Medline, CASGoogle Scholar
    • 63  Cross SE, Jin YS, Tondre J et al. AFM-based analysis of human metastatic cancer cells. Nanotechnology19(38),384003 (2008).Crossref, MedlineGoogle Scholar
    • 64  Obataya I, Nakamura C, Han SW, Nakamura N, Miyake J. Nanoscale operation of a living cell using an atomic force microscope with a nanoneedle. Nano Lett.5(1),27–30 (2005).Crossref, Medline, CASGoogle Scholar
    • 65  Cuerrier CM, Lebel R, Grandbois M. Single cell transfection using plasmid decorated AFM probes. Biochem. Biophys. Res. Commun.355(3),632–636 (2007).Crossref, Medline, CASGoogle Scholar
    • 66  Chen X, Kis A, Zettl A, Carolyn RB. A cell nanoinjector based on carbon nanotubes. Proc. Natl Acad. Sci. USA104(20),8218–8222 (2007).Crossref, Medline, CASGoogle Scholar
    • 67  Vakarelski IU, Brown SC, Higashitani K, Moudgil BM. Penetration of living cell membranes with fortified carbon nanotube tips. Langmuir23(22),10893–10896 (2007).Crossref, Medline, CASGoogle Scholar
    • 68  Guo XE, Takai E, Jiang XY et al. Intracellular calcium waves in bone cell networks under single cell nanoindentation. Mol. Cell Biomech.3(3),95–107 (2006).MedlineGoogle Scholar
    • 69  Huo B, Lu XL, Costa KD, Xu Q, Guo XE. An ATP-dependent mechanism mediates intercellular calcium signaling in bone cell network under single cell nanoindentation. Cell Calcium47(3),234–241 (2010).Crossref, Medline, CASGoogle Scholar
    • 70  Shi L, Shi S, Han D et al. AFM and fluorescence imaging of nanomechanical response in periodontal ligament cells. Front Biosci. (Elite Ed).2,1028–1041 (2010).Crossref, MedlineGoogle Scholar
    • 71  He K, Shi X, Fang X et al. Long-distance intercellular connectivity between cardiomyocytes and cardiofibroblasts mediated by membrane nanotubes. Cardiovasc. Res.92(1),39–47 (2011).Crossref, Medline, CASGoogle Scholar
    • 72  Dixon IM, Davies JJ. Fibroblasts are coupled to myocytes in heart muscle by nanotubes: a bigger and better syncytium? Cardiovasc. Res.92(1),5–6 (2011).Crossref, Medline, CASGoogle Scholar
    • 73  Katan AL, Dekker C. High-speed AFM reveals the dynamics of single biomolecules at the nanometer scale. Cell147(5),979–982 (2011).Crossref, Medline, CASGoogle Scholar
    • 74  Harke B, Chacko JV, Haschke H, Canale C, Diaspro A. A novel nanoscopic tool by combining AFM with STED microscopy. Optical Nanoscopy doi:10.1186/2192-2853-2851-2853(2012) (Epub ahead of print).Google Scholar