Browsing by Subject "atomic force microscopy"
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Item Effects of mechanical forces on cytoskeletal remodeling and stiffness of cultured smooth muscle cells(2009-06-02) Na, SungsooThe cytoskeleton is a diverse, multi-protein framework that plays a fundamental role in many cellular activities including mitosis, cell division, intracellular transport, cell motility, muscle contraction, and the regulation of cell polarity and organization. Furthermore, cytoskeletal filaments have been implicated in the pathogenesis of a wide variety of diseases including cancer, blood disease, cardiovascular disease, inflammatory disease, neurodegenerative disease, and problems with skin, nail, cornea, hair, liver and colon. Increasing evidence suggests that the distribution and organization of the cytoskeleton in living cells are affected by mechanical stresses and the cytoskeleton determines cell stiffness. We developed a fully nonlinear, constrained mixture model for adherent cells that allows one to account separately for the contributions of the primary structural constituents of the cytoskeleton and extended a prior solution from the finite elasticity literature for use in a sub-class of atomic force microscopy (AFM) studies of cell mechanics. The model showed that the degree of substrate stretch and the geometry of the AFM tip dramatically affect the measured cell stiffness. Consistent with previous studies, the model showed that disruption of the actin filaments can reduce the stiffness substantially, whereas there can be little contribution to the overall cell stiffness by the microtubules or intermediate filaments. To investigate the effect of mechanical stretching on cytoskeletal remodeling and cell stiffness, we developed a simple cell-stretching device that can be combined with an AFM and confocal microscopy. Results demonstrate that cyclic stretching significantly and rapidly alters both cell stiffness and focal adhesion associated vinculin and paxillin, suggesting that focal adhesion remodeling plays a critical role in cell stiffness by recruiting and anchoring F-actin. Finally, we estimated cytoskeletal remodeling by synthesizing data on stretch-induced dynamic changes in cell stiffness and focal adhesion area using constrained mixture approach. Results suggest that the acute increase in stiffness in response to an increased cyclic stretch was probably due to an increased stretch of the original filaments whereas the subsequent decrease back towards normalcy was consistent with a replacement of the highly stretched original filaments with less stretched new filaments.Item Geometric Nanoconfinement Effects on the Electronic and Mechanical Properties of Self-Assembled Molecular Systems(2014-08-20) Ewers, Bradley WilliamWith the ongoing research and development of nanoscale technologies and materials, it becomes increasingly important to understand how local environment influences molecular and material properties. An important factor in this regard is geometric nanoconfinement, for example, the restriction of molecules to nanostructure surfaces. The bulk or average characteristics of materials and molecules do not appropriately define their behavior in these circumstances, and highly localized measurement techniques developed to specifically identify the influence of confinement on their properties is essential to understanding their characteristics and behavior. In this dissertation, two forms of geometric confinement are considered in the context of different molecular properties. First, the role of radial confinement on the tribological properties of self-assembled monolayers (SAMs) is considered. SAMs are an excellent model lubricant for experimental studies of boundary lubrication, and they have been employed as boundary lubricant additives and surface coatings. The lubricated contacts of technologically relevant surfaces, however, consist of asperity interactions, and the summit curvature of these asperities can impact the critical cohesive forces from which the properties of the SAM are derived. Molecular dynamics simulation was employed to understand the influence of nanoscopic surface curvature, as well as surface coverage density, two factors which together contribute to the cohesive forces of SAMs, on their tribological properties. In particular their dissipative potential and effective surface protection were examined, as well as the influence of these factors on the contact mechanics of functionalized nanoasperity contacts. Another mode of geometric confinement studiedin this work is two-dimensional nanoconfinement of molecules and its influence on the mechanism of charge transport in molecular systems. Effective control of charge transport in molecules is essential for molecular modification of CMOS technologies, and is critical in controlling charge carrier dynamics in dye-sensitized photovoltaics. In this work, the size dependence of the electronic properties of thiol-tethered zinc porphyrin aggregateson the Au(111) surface was investigated. AFM nanolithography was used to confine these molecules within an alkanethiol matrix on the Au(111) surface, forming molecular islands of specific dimensions to investigate the relationship between island size and charge transport, demonstrating a shift from tunneling based charge transport to the more tunable and efficient charge hopping based transport.Item The nanomechanics of polycystin-1: A kidney mechanosensor(2010-06-25) Liang Ma; Simon A. Lewis; Roger B. Sutton; Paul J. Boor; Guillermo A. Altenberg; Andres F. OberhauserMutations in polycystin-1 (PC1) can cause Autosomal Dominant Polycystic Kidney Disease (ADPKD), which is a leading cause of renal failure. The available evidence suggests that PC1 acts as a mechanosensor, receiving signals from the primary cilia, neighboring cells, and extracellular matrix. PC1 is a large membrane protein that has a long N-terminal extracellular region (about 3000 aa) with a multimodular structure including sixteen Ig-like PKD domains, which are targeted by many naturally occurring missense mutations. Nothing is known about the effects of these mutations on the biophysical properties of PKD domains. In addition, PC1 is expressed along the renal tubule, where it is exposed to a wide range of concentration of urea. Urea is known to destabilize proteins. Other osmolytes found in the kidney such as sorbitol, betaine and TMAO are known to counteract urea¡¯s negative effects on proteins. Nothing is known about how the mechanical properties of PC1 are affected by these osmolytes. Here I use nano-mechanical techniques to study the effects of missense mutations and effects of denaturants and various osmolytes on the mechanical properties of PKD domains. Several missense mutations were found to alter the mechanical stability of PKD domains resulting in distinct mechanical phenotypes. Based on these findings, I hypothesize that missense mutations may cause ADPKD by altering the stability of the PC1 ectodomain, thereby perturbing its ability to sense mechanical signals. I also found that urea has a significant impact on both the mechanical stability and refolding rate of PKD domains. It not only lowers their mechanical stability, but also slows down their refolding rate. Moreover, several osmolytes were found to effectively counteract the effects of urea. Our data provide the evidence that naturally occurring osmolytes can help to maintain Polycystin-1 mechanical stability and folding kinetics. This study has the potential to provide new therapeutic approaches (e.g. through the use of osmolytes or chemical chaperones) for rescuing destabilized and misfolded PKD domains.