Tracking molecules in stressed cells: multimodal microscopy and single molecule spectroscopy for mechanotransduction

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  • $450 Journal of Biomechanics 2006, Vol. 39 (Suppl 1)

    5738 Th, 12:00-12:15 (P42) Ultrasound elastography in consideration of finite deformations S. Reichling 1 , W. Khaled 2, O. Timme Bruhns 1 , H. Ermert 2. 1Institute of Mechanics, Ruhr-University Bochum, Bochum, Germany, 2Institute of High Frequency Engineering, Ruhr-University Bochum, Bochum, Germany

    Nowadays, palpation is the most frequently used screening method for the detection of breast or prostate tumours. The basis of this method is the difference between mechanical properties of healthy and diseased tissue. In most cases this ratio is significant. Commonly used imaging techniques, such as computer tomography or ultrasound, are not able to measure these properties. However, elastography based on ultrasound imaging provides a possibility to determine elastic material properties of soft tissue indirectly. In this imaging method, an investigated object is compressed quasi-statically. By comparing the ultrasound signals for two different load cases, the interior axial dis- placement field (the displacement component in compression direction) can be determined. Using the assumptions that the considered material is linear elastic, isotropic and incompressible, the shear modulus distribution can be calculated in the framework of inverse problems. Clearly, if the deformation of the object under investigation is too large, the theory of linear elasticity cannot be applied any more. Therefore, in this paper, a new approach is presented to solve the inverse problem of elastography taking into account fully elastic deformations. For this reason, the shear modulus distribution in the region of interest is calculated using an iterative method minimising the difference between the measured and the estimated displacement field. The required calculation of the displacement follows from the solution of a non-linear boundary value problem. In this connection, the material response is approximated by an Ogden material. The applicability as well as the performance of the approach is demonstrated by some 2D numerical examples.

    4533 Th, 12:15-12:30 (P42) Application of acoustoelasticty to biological tissues: a novel non-invasive technique for tissue property evaluation H. Kobayashi, R. Vanderby. Dept. of Orthopedics and Rehabilitation, University of Wisconsin-Madison, Wisconsin, USA

    Acoustoelasticity (AE) describes changes in acoustic characteristics in a deformed medium due to its strain dependent stiffness [1]. Hence, the wave propagation in a deformed medium can be rigorously analyzed only with AE. In the field of ultrasound-based medical imaging, evaluation and visualization of in-vivo tissue stiffness has become an important research topic. Recently, a new technique termed "elastography" is gaining popularity as an alternative to traditional sonography. Elastography evaluates strain distributions in tissues that are compressed by comparing the wave signals acquired pre- and post compression [2]. However, due to the simplified equations (not considering the AE effect), elastography can be problematic in tissues with strong strain- dependent stiffness [3]. As a theoretical basis, we developed a new strain energy function for nearly incompressible materials [4,5]. We then developed a novel non-invasive, ultrasound-based technique to evaluate the stiffness- strain relation and applied strain in compressed tissues. We use ultrasound echo signals transmitted and received by one transducer in the same direction as the applied tissue compression. We compute reflection coefficients (ratio of incident wave to reflected wave) and wave travel times through the tissue thickness from this system. From these data we simultaneously evaluate applied tissue strain and key parameters (stiffness-strain relation, damping factor) via AE theory. Since tissues are known to have characteristic stiffness- strain relations [6], stiffness-strain results can be used to identify tissue types orto quantify pathologic or healing properties. In this study, the newly proposed technique is applied to several different tumors and healthy tissues and tissue specific stiffness-strain relations and damping factors are successfully evaluated.

    References [1] Pao YH, et al. Physical acoustics Vol. XVII. 1984; Academic Press, ch. 2. [2] Ophir J, et al. Ultrasound in Med. Biol. 2000; 26(Suppl 1); $23-29. [3] Varghese T., et al. Ultrasound Med. Biol. 2000; 26: 839451. [4] Kobayashi H. and Vanderby R.J. Applied mechanics. 2005; 72: 843-851. [5] Kobayashi H. Ph.D. dissertation, University of Wisconsin-Madison 2005, ch. 6. [6] Krouskop TA, et al. Ultrasonic imaging. 1998; 20: 260-274.

    Oral Presentations

    T4.7 Quantitative Functional Imaging 7459 We, 11:00-11:15 (P33) Tracking molecules in stressed cells: multimodal microscopy and single molecule spectroscopy for mechanotransduction P.J. Butler, R. Gullapalli, T. Tabouillot. Ceil and Tissue Mechanotransduction Lab; The Pennsylvania State University; Department of Bioengineering; University Park, PA, USA

    Forces induce coordinated biochemical signaling cascades in cells over a wide range of spatial and temporal scales. We have developed new integrated tools based on multimodal microscopy and single photon counting to elucidate the molecular basis of mechanotransduction over large temporal ranges in cells subjected to well-defined forces. Total internal reflection, epi-fluorescence, differential interference contrast, and 3-D deconvolution provide the means to correlate the single molecule spectroscopic measurements from fluorescence correlation spectroscopy (FCS) and time-resolved fluorescence with precise spatial locations in the cell. With this new hybrid microscopy system, precise effects of force on single molecule dynamics can be mapped spatially in response well-defined forces imposed by micropipette aspiration, optically trapped beads, and fluid flow. We describe the construction of our multi-modal microscope in detail and the calibrations necessary to define molecular dynam- ics in cells and model membranes. Finally, we discuss the potential applications of the system and its implications for the field of mechanotransduction.

    4817 We, 11:15-11:30 (P33) A novel cell tracking method for in vivo biomechanical assessment of healing murine tendon: a pilot study J.G. Snedeker 1 , G. Pelled 2, ~ Zilberman 2, R. MLiller 1 , D. Gazit 2. 1Institute for Biomedical Engineering, University and ETH Zurich, Switzerland, 2Skeletal Biotechnology Laboratory, Hebrew University, Hadassah Medical Center, Jerusalem, Israel

    This study explores and describes a novel method to quantify in vivo tendon biomechanics from endoscopic confocal fluorescence microscope images of externally loaded tendon. The proposed method was developed to enable pe- riodic, on-line assessment of healing in genetically engineered mesenchymal stem cell tendon injury therapies. In an initial step, a single female C3H/HeN mouse, aged 10 weeks, was anes- thetized and the cells of the right Achilles tendon were stained in the elastic region using a fluorescent nucleic acid stain. The femur and tibia of the mouse were physically constrained in a loading jig that provided a quantified rotational moment at the ankle joint. During loading, the relative motions tenocytes were recorded using a CelI-Vizio (Mauna Kea Technologies, USA) endoscopic micro-imaging system. The cellular displacements were then analyzed using a custom developed, cross-correlation based, tracking algorithm to extract and map the tissue strain field. The veracity and repeatability of both the tendon loading protocol and automatic tracking algorithm were established by manually measuring and analyzing tenocyte displacement within a prominent superficial tendon structure. Over repeated trials, the primary engineering strain in the reference tendon structure was 12.41.3%. Thus, reproducibility of the tendon loading device was established. The full field cellular displacements and corresponding local tissue strains revealed primary engineering strains in the tissue to range from -3 to 18%, indicating a highly inhomogenous strain state. Close inspection of the full field cellular displacements showed that the tendon structure is complex, with stratified tissue structures that alternately load and unload according to the degree of ankle rotation. In conclusion, a novel method for in vivo assessment of tendon stiffness has been proposed and partially validated. While some aspects require refinement, the method shows exciting promise for application in longitudinal study of healing in regenerative connective tissue therapies.

    7296 We, 11:30-11:45 (P33) Trabecular bone density and strength assessment using non-invasive scanning confocal ultrasound imaging ~-X. Qin, ~ Xia, W. Lin, E. Mittra, B. Gruber, C. Rubin. Department of Biomedical Engineering, State University of New York at Stony Brook, Stony Brook, NY, USA

    Conclusive evidence indicates that osteopenia occurs due to aging and micro- gravity. Early diagnosis of such skeletal disorders leads to prompt treatment and dramatically reduces the risk of complications. Bone integrity is dependant not only on the mineral density, but on the quality, i.e., the strength and structural parameters. Advents in quantitative ultrasound (QUS) provide a unique method for evaluating both bone strength and density. Using our newly developed scanning confocal acoustic diagnostic (SCAD) system, the objectives of this study were to evaluate the ability of QUS to predict trabecular

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