title: A microfluidic system with optical laser tweezers to study mechanotransduction and focal adhesion recruitment.
abstract: We present a new method to locally apply mechanical tensile and compressive force on single cells based on integration of a microfluidic device with an optical laser tweezers. This system can locate a single cell within customized wells exposing a square-like membrane segment to a functionalized bead. Beads are coated with extracellular matrix (ECM) proteins of interest (e.g. fibronectin) to activate specific membrane receptors (e.g. integrins). The functionalized beads are trapped and manipulated by optical tweezers to apply mechanical load on the ECM-integrin-cytoskeleton linkage. Activation of the receptor is visualized by accumulation of expressed fluorescent proteins. This platform facilitates isolation of single cells and excitation by tensile/compressive forces applied directly to the focal adhesion via specific membrane receptors. Protein assembly or recruitment in a focal adhesion can then be monitored and identified using fluorescent imaging. This platform is used to study the recruitment of vinculin upon the application of external tensile force to single endothelial cells. Vinculin appears to be recruited above the forced bead as an elliptical cloud, centered 2.1 ± 0.5 μm from the 2 μm bead center. The mechanical stiffness of the membrane patch inferred from this measurement is 42.9 ± 6.4 pN μm(-1) for a 5 μm × 5 μm membrane segment. This method provides a foundation for further studies of mechanotransduction and tensile stiffness of single cells.
authors.affiliation: Massachusetts Institute of Technology (MIT), Department of Biological Engineering, Cambridge, MA 02139, USA.
authors.forename: Peyman
authors.initials: P
authors.lastname: Honarmandi
authors.forename: Hyungsuk
authors.initials: H
authors.lastname: Lee
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Lang
authors.forename: Roger D
authors.initials: RD
authors.lastname: Kamm
mesh_terms: D002469:Cell Separation; D002478:Cells, Cultured; D003198:Computer Simulation; D004548:Elasticity; D004867:Equipment Design; D022001:Focal Adhesions / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D040542:Mechanotransduction, Cellular / Q000502:physiology*; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D008856:Microscopy, Fluorescence; D008858:Microscopy, Phase-Contrast; D052898:Optical Tweezers; D016596:Vinculin / Q000378:metabolism
chemical_list: D016596:Vinculin