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luorescence imaging techniques with force-application methods, to observe
structural intracellular changes in response to extracellular perturbations.
Among these studies are the observations of changes in the actin and MT
cytoskeleton of live ibroblast cells in response to deformations produced
by glass needles, which were visualized using GFP-tagged cytoskeletal
proteins.
Deformations of the IF cytoskeleton were analysed by visualizing
GFP-vimentin in live endothelial cells before and after the application
of shear stress in a low chamber.
66
In another study that combined the
magnetic bead twisting technique with GFP-luorescent imaging, application
of forces to focal adhesions by the use of speciically coated beads resulted
in displacements of actin ilament bundles at distances of 20-30 μm from
the beads.
67
A similar technique was used to visualize displacements
of intracellular organelles such as mitochondria 69 and to analyse the
propagation of forces to the nucleus by quantifying displacements of
nucleolar structures in response to load. 70 Visualization of responses to
extracellular perturbations is not limited to the tracking of natural organelles
or cytoskeletal components: in a recent study, AFM was used to apply
perturbations onto live, adherent cells, while quantifying stress propagation
through the cell by tracking of integrin-bound luorescent microspheres. 71
Here, we will review some of our previous work
68
on the application
of simultaneous AFM and luorescence microscopy or laser scanning
confocal microscopy (LSCM) in the context of living mammalian cells. Three
examples will be presented which demonstrate the utility of simultaneous
AFM and optical approaches to understand the origin and control of force
transmission inside and through living mammalian cells to the underlying
substrate.
18,19,72-74
18.3 CELLULAR NANOMECHANICS AND FORCE TRANSDUCTION
THROUGH THE CELLULAR ARCHITECTURE
18.3.1 Mitochondrial Displacements in Response to Force
Mitochondria are semi-autonomous and highly dynamic organelles, which
have the ability to change their shape and their location inside the living
cell. 75 Localization and rearrangement of mitochondria in higher eukaryotes
is known to be dependent on the MT. More recent research suggests
that actin ilaments have an important role as well, such as facilitating
mitochondrial organization in yeast and vertebrate neurons, 76,77 and
controlling mitochondrial movement and morphology. 78 Given the strong
association of mitochondria with the cytoskeleton, it is predicted that
forces locally applied by the AFM tip will affect their arrangement through
mechanical transduction. 79-81
 
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