The Nanoscale World

Analysis of Cytoskeleton-Destabilizing Agents by Optimized Optical Navigation and AFM Force Measurements


Tue, Feb 23 2010

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Analysis of Cytoskeleton-Destabilizing Agents by Optimized Optical Navigation and AFM Force Measurements

Mechanical properties of cells are determined by the dynamic behavior of the cytoskeleton and physical interactions with the environment. The cytoskeleton, composed of actin filaments, intermediate filaments, and microtubules, is vital for numerous key cellular processes, such as cell division, vesicle trafficking, cell contraction, cell motility, and cell signaling. There is increasing evidence that deregulation of cytoskeletal components like disassembly of actin and tubulin filaments is an important parameter in cellular pathology. Thus, significant alterations of the mechanical phenotype of the cell and its surrounding microenvironment are reported to be involved in aberrant cellular processes and successively contribute to onset and progression of diseases such as cancer, malaria, and possibly neurodegeneration [1–5]. In vitro and ex vivo biomechanical studies have shown that cancer cells have significantly decreased elastic moduli than their normal counterparts, a characteristic that is attributed to the ability of cancer cells to metastasize or spread [6, 7].

 

Along with other techniques, atomic force microscopy (AFM) has become a powerful method to explore mechanical properties of living cells in nearly physiological conditions [8,9] and even of ex vivo cancer cells [6]. Living cells typically show a Young’s modulus between 0.5 and 200 kPa depending on sub-cellular region and cell type. These specific mechanobiological properties can be quantified by AFM force measurements and subsequently used to assess and correlate effects of drug treatment, aging, or pathology [10–14].

 

In this article, we used Multiple Image Registration Overlay (MIRO) software and the Bioscope Catalyst Atomic Force Microscope (Veeco Instruments, Santa Barbara, CA) to facilitate optical navigation targeting the best locations for force measurements in living cells. This approach was used to explore real-time changes in cell integrity of two cancer cell lines, HeLa and U2-O2 osteosarcoma, upon treatment with the cytoskeleton-destabilizing drugs, nocodazole and latrunculin B. The apparent Young’s moduli of these cells were quantified and compared. This allowed us to differentiate the visco-elastic changes caused by the disassembly of the actin cytoskeleton from those caused by the disassembly of the microtubule network. Furthermore, it enabled us to discriminate the two types of cancer cells based on their elastic properties.

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