The 5 th Korean-US NanoForum: Nanobiotechnology. Jeju Island, April 17 th -19 th , 2008 Focal Adhesion Mechanotransduction: Focal Adhesion Mechanotransduction: Cellular Response to Nanoscale Cellular Response to Nanoscale Mechanical Factors Mechanical Factors Nathan J. Sniadecki, Ph.D. Mechanical Engineering Department University of Washington Seattle, Washington
Mechanotransduction at Focal Adhesions Mechanotransduction at Focal Adhesions Focal adhesions are force sensitive integrin-receptor complexes (100 nm – 1 μ m). Structural proteins link integrins to actin to Structural proteins develop traction force for cellular migration or Actin contraction α -Act Signaling proteins activated with integrin- Signaling proteins Src p130 FAK FAK pax matrix binding and force vin vin tal tal tal Integrins Integrins Traction forces or external forces are transduced into biological signals that Extracellular matrix (ECM) can regulate cell function Traction Force Myosin Actin Cell External Force ) F A ( o n s i h e d A c a l o F Extracellular Matrix (ECM)
Traction Force Sensor Array Traction Force Sensor Array Arrays of flexible microposts to measure traction forces Made with soft lithography of polydimethlysiloxane (PDMS) 20 nN 10 μ m DAPI Actin PDMS Vinculin 3- μ m diameter, 10- μ m height microposts
Magnetic Microposts Magnetic Microposts Sensor and Actuator System Sensor and Actuator System • Measure traction forces with microposts π d 3 E = = F K δ δ tr 3 64L • Apply forces with magnetic nanowires τ + 3 ( L L ) τ = µ ⋅ = B w F ⊥ Mag 2 + + 2 2( L L L L ) w w Cobalt Nanowires Cobalt Nanowires • 300 nm diameter, 5-7 μ m long High magnetic moment ( µ ) along long axis • Nanowire Nanowires N.J. Sniadecki, et al., PNAS , 2007, 104:114553-8 N.J. Sniadecki, et al., Rev Sci Instr , 2008, 79, 044302
Magnetic Nanowires Magnetic Nanowires Cobalt Electrodeposition Vibrating Sample Magnetometer Template Co 2+ Co 2+ Co 2+ Co 2+ Co 2e - Co 2+ (aq) + 2e - → Co(s) 2 µm N.J. Sniadecki, et al., PNAS , 2007, 104:114553-8 N.J. Sniadecki, et al., Rev Sci Instr , 2008, 79, 044302
Fabrication Fabrication Nanowires magnetically embedded into PDMS mold 1. Lithography 4. Deposit Nanowires SEM Image 2. Develop 5. Encase in PDMS EDS Measurement 3. Cast in PDMS 6. Release N.J. Sniadecki, et al., PNAS , 2007, 104:114553-8 N.J. Sniadecki, et al., Rev Sci Instr , 2008, 79, 044302
Magnetic Stimulation Magnetic Stimulation Magnetic actuation for external force Stimulation relaxes traction forces Focal adhesion protein recruitment N.J. Sniadecki, et al., PNAS , 2007, 104:114553-8
Spatial Response Spatial Response Relaxation occurs at remote sites along cell periphery but not at interior posts Magnetic Post Jump Gradual Jump Gradual Binomial Analysis: Edge posts have significant loss versus interior posts (p<0.05) Post local to magnetic posts have no significant difference to distal posts (p>0.25) Control Control N.J. Sniadecki, et al., PNAS , 2007, 104:114553-8
Nanoposts Nanoposts High Spatial Resolution of Traction Forces 830 nm dia, 3.3 μ m tal 830 nm dia, 3.0 μ m tall 16 nN/ μ m 28 nN/ μ m Sloped-wall cantilever 670 nm dia, 3.3 μ m tal 8 nN/ μ m M.T. Yang, N.J. Sniadecki, C.S. Chen, Adv Mat , 2007, 104:114553-8
Acknowledgements Acknowledgements Nathan J. Sniadecki Sangyoon Han Aimi Ahmad Shukri Collaborators: SUPPORT: National Institutes of Health Christopher S. Chen (UPenn) National Science Foundation Daniel H. Reich (JHU) The Hartwell Foundation UW Royalty Research Fund
Micropost Force Sensors Micropost Force Sensors Post Array: No adhesion blocking: Microcontact printing: Adhesion only on tips: PEO PPO Fibronectin Measurements: Posts deflects as simple cantilever springs: π 4 3 Ed = δ = δ F K 3 64L F … Cellular Force K … Spring Stiffness (32 nN/ μ m) δ … Displacement E … Elasticity Modulus of PDMS d … Post diameter (3 μ m dia) L … Post Length (10 μ m) J. L. Tan, et al., Proc. Nat. Acad. Sci. , 2003, 100:1484-1489 C. A. Lemmon, et al., Mech. & Chem. Biosystems. , 2005, 2(1) 1-16
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