Biomedical Engineering Reference
In-Depth Information
a
b
1.0
n =
6
MEAN ± S.D.
0.8
0.6
0.4
8.0
7.0
6.0
5.0
4.0
3.0
2.0
1.0
0.0
Distance from the leading edge
x
[µm]
Fig. 4.4
Distribution profile of actin filament density of a keratocyte (Adachi et al.
2009
).
(
a
) Image of keratocyte analyzed for actin density distribution. (
b
) Mean actin network distribu-
tion analyzed over the
boxed region
in (
a
). Actin network density decreases from a peak at the
leading edge (
right-hand side
) toward the back of the lamellipodia (
left-hand side
) (Adapted with
permission from Elsevier Ltd.: [Journal of Biomechanics], copyright (2008))
back of the lamellipodia where the myosin II motors are most abundant is in agreement
with the dynamic network contraction model (Svitkina et al.
1997
).
It should be noted that although the normal incremental strain, ∆ε
n
()
(Fig.
4.4c
) is nearly zero even at the back of the lamellipodia where SFs are local-
ized, this does not contradict the generation of tension along the SFs because ten-
sion along the SFs is generated by isometric contraction, which does not necessarily
translate into a noticeable strain along the fibers.
4.6
Role of Mechanical Strain in Actin Network
Reorganization
4.6.1
Distribution of Actin Network Density
To understand the role of the mechanical strain in the actin network, we correlated
it with actin network density. First, we elaborate on how to analyze the distribution
of actin filament network in the lamellipodia. For this purpose, actin filament net-
work in the lamellipodia of fish keratocytes can be labeled using an appropriate dye
such as rhodamine phalloidin.
For the live-cell staining of the actin cytoskeleton, keratocytes are fixed in 5 %
paraformaldehyde, permeabilized with 0.2 % Triton-X for 10 min and rinsed with
PBS before staining with rhodamine phalloidin (25 nM) diluted in PBS for 150 min
at 37 ﾰC. After rinsing 3-4 times with PBS, the cells are imaged while immersed in
PBS to avoid drying. A sample image of fish keratocyte stained with rhodamine
