913

Beneficial and harmful effects of oscillatory

mechanical strain on airway smooth muscle1
Geoffrey N. Maksym, Linhong Deng, Nigel J. Fairbank, C.A. Lall, and
Sarah C. Connolly

Abstract: Airway smooth muscle (ASM) cells are constantly under mechanical strain as the lung cyclically expands
and deflates, and this stretch is now known to modulate the contractile function of ASM. However, depending on the
experimental conditions, stretch is either beneficial or harmful limiting or enhancing contractile force generation, re-
spectively. Stretch caused by a deep inspiration is known to be beneficial in limiting or reversing airway constriction in
healthy individuals, and oscillatory stretch lowers contractile force and stiffness or lengthens muscle in excised airway
tissue strips. Stretch in ASM culture has generally been reported to cause increased contractile function through in-
creases in proliferation, contractile protein content, and organization of the cell cytoskeleton. Recent evidence indicates
the type of stretch is critically important. Growing cells on flexible membranes where stretch is non-uniform and
anisotropic leads to pro-contractile changes, whereas uniform biaxial stretch causes the opposite effects. Furthermore,
the role of contractile tone might be important in modulating the response to mechanical stretch in cultured cells. This
report will review the contrasting evidence for modulation of contractile function of ASM, both in vivo and in vitro,
and summarize the recent evidence that mechanical stress applied either acutely within 2 h or chronically over 11 d is
a potent stimulus for cytoskeletal remodelling and stiffening. We will also point to new data suggesting that perhaps
some of the difference in response to stretch might lie with one of the fundamental differences in the ASM environ-
ment in asthma and in culture — the presence of elevated contractile tone.
Key words: length–tension, contraction, perturbed equilibrium of myosin binding, mechanical stress, cell phenotype,
myosin light chain kinase, tone and relaxation, asthma.

Résumé : Les cellules musculaires lisses des voies aériennes 922 (MLVA) sont soumises à une déformation mécanique
constante à chaque cycle de gonflement et de dégonflement du poumon, et l’on sait maintenant que cet étirement mo-
dule la fonction contractile des MLVA. Toutefois, selon les conditions expérimentales, l’étirement est soit bénéfique,
soit néfaste, en limitant ou en stimulant la production de la force contractile, respectivement. L’étirement causé par une
inspiration profonde a un effet bénéfique en limitant ou en renversant la constriction des voies aériennes chez les sujets
en santé, et l’étirement oscillatoire diminue la rigidité et la force contractile, ou allonge le muscle dans les bandes de
tissus excisés des voies aériennes. On indique généralement que, dans les MLVA en culture, l’étirement entraîne une
augmentation de la fonction contractile par le biais d’augmentations de la prolifération, de la teneur en protéines contrac-
tiles et de l’organisation du cytosquelette des cellules. De récentes observations indiquent que le type d’étirement a une
grande importance. La culture des cellules sur des membranes flexibles, où l’étirement est non uniforme et anisotrope,
entraîne des modifications procontractiles, alors que l’étirement biaxial uniforme a des effets contraires. Le tonus con-
tractile pourrait aussi jouer un rôle important en modulant la réponse à l’étirement mécanique dans les cellules culti-
vées. Ce compte rendu fera une synthèse des différentes données probantes sur la modulation de la fonction contractile
des MLVA, in vivo et in vitro, et résumera les récentes observations indiquant qu’un stress mécanique, appliqué de ma-
nière aiguë dans un délai de 2 h ou chroniquement pendant 11 jours, est un stimulus puissant pour le remodelage et la
rigidité du cytosquelette. Nous présentons aussi de nouveaux résultats qui semblent indiquer qu’une partie de la diffé-
rence dans la réponse à l’étirement pourrait résider dans l’une des différences fondamentales dans l’environnement du
MLVA de l’asthmatique et en culture, soit la présence d’un tonus contractile élevé.

Mots clés : longueur–tension, contraction, équilibre perturbé de la fixation de la myosine, stress mécanique, phénotype
cellulaire, kinase de la chaîne légère de la myosine, tonus et relaxation, asthme.

[Traduit par la Rédaction] Maksym et al.

Received 12 January 2005. Published on the NRC Research Press Web site at http://cjpp.nrc.ca on 8 November 2005.
G.N. Maksym,2 N.J. Fairbank, C.A. Lall, and S.C. Connolly. School of Biomedical Engineering, Dalhousie University,
5981 University Ave., Halifax, NS B3H 1W2, Canada.
L. Deng. Physiology Program, Harvard School of Public Health, Boston, MA 02115, USA.
1
This paper is one of a selection of papers published in this Special Issue, entitled Models of smooth muscle contraction, and has
undergone the Journal’s usual peer review process.
2
Corresponding author (e-mail: gmaksym@dal.ca).

Can. J. Physiol. Pharmacol. 83: 913–922 (2005) doi: 10.1139/Y05-091 © 2005 NRC Canada
914
Introduction
Asthma is associated with airway hyperresponsiveness be-
cause of inhaled agonists, inflammation of the airways, and
airway remodelling including increases in airway smooth
muscle (ASM) cells and possibly changes in the mechanical
stresses in the airways. Of these contributing factors, it is the
abnormal enhanced constriction of the ASM, or the inability
to reverse this constriction, that makes asthma a dangerous
disease. How the contractile behaviour of the ASM lining the
airways becomes altered in asthma is an unanswered ques-
tion. In this report, we will describe contrasting behaviour re-
ported for mechanical stretch on ASM function. Stretch can
be beneficial to ASM function, leading to muscle relaxation
and larger airway diameters, and stretch can be harmful or
maladaptive,3 leading to stronger, stiffer muscle with an in-
creased ability to shorten. Understanding the role of stretch
to ASM function is important, as stretch is a normal part of
the environment of the airway, stretching approximately 3%
in length with tidal breathing and much more with deep in-
flation in healthy lungs.
Stretch is beneficial to ASM function in fresh tissue
and in vivo
In healthy individuals, a deep breath prior to inhalation of a
spasmogen attenuates subsequent constriction (Kapsali et al.
2000). Also, a deep inspiration following provoked broncho-
constriction results in residual dilation lasting for many sec-
onds before renarrowing in healthy individuals (Pellegrino
et al. 1996; Salome et al. 2003). These are known, respec-
tively, as the bronchoprotective and bronchodilatory effect of
deep inspirations and both of these effects are significantly
attenuated in asthma (Fredberg 2004; Skloot and Togias
2003; Salome et al. 2003).
The mechanisms for these effects and their attenuation in
asthma are unknown but have been associated with hyper-
responsiveness in asthma (Mitzner and Brown 2000). Al-
though airway hyperresponsiveness is not explained by the
lack of bronchoprotective or bronchodilatory effects alone
(Brusasco and Pellegrino 2003), the following behaviour of
ASM might explain the dilatory effect of deep inspiration. In
vitro experiments on activated ASM strips show that tidal
oscillations of sufficient magnitude lead to either a softening
or lengthening of ASM or both, depending on the loading
conditions (Fredberg et al. 1996; Latourelle et al. 2002;
Raboudi et al. 1998). Thus, the length of activated ASM is
dynamically determined and is regulated by the load, the
state of muscular activation, and by the degree of length per-
turbation continuously afforded to the muscle by the action
of breathing and periodic deep inspirations. However, if this
mechanism explains the bronchodilatory effect of deep in-
spirations, it is unknown why this is impaired in asthma.
Using computed tomography (CT) imaging in mild asthma,
Brown et al. (2001) observed that while the larger airways
do dilate in response to a deep inspiration, they subsequently
rapidly renarrow indicating perhaps a change in ASM con-
tractile properties. In more severe asthma, the amount of di-
3
In this report, maladaptive is used to describe changes in ASM that increase its contractile function. Indeed, like the appendix, it is unclear
what benefit if any ASM surrounding the airways gives, thus enhanced contractility has no clear function other than to be harmful in airway
pathologies such as asthma (Mitzner 2004).
Can. J. Physiol. Pharmacol. Vol. 83, 2005
lation with a deep inspiration varies considerably amongst
airways such that it is possible that constricted airways do
not receive sufficient stretch, indicating airway stiffness may
be increased in those airways relative to parenchymal tether
stiffness (Spector et al. 1996). Although other effects such
as a change in parenchymal properties could contribute to a
reduced bronchodilator effect from a deep inspiration, these
authors concluded that in subjects with mild asthma, the re-
sponsible mechanisms are likely to be intrinsic to changes in
the smooth muscle (Spector et al. 1996; Mitzner and Brown
2000).
The beneficial aspects of stretch in normal tissue or
healthy individuals contrast with the effects of stretch on
cultured ASM cells (described in the next section).
Stretch is maladaptive to ASM cell function in culture
ASM cells grown in static culture are different then cells
in vivo. Cultured cells retain much of their physiologic re-
sponses to contractile or relaxant agonists to some degree,
but they decrease their production of specific contractile pro-
teins that continues with cell generation number. In a series
of studies conducted by Smith et al., it was shown that cy-
clic stretching of ASM cells on flexible silastic membranes
caused several cellular functional and structural changes in-
cluding increased contractile specific proteins. With cyclic
stretch applied for up to 14 d, this type of strain increases
protein markers of cell contractility such as smooth muscle
myosin, smooth muscle myosin light chain kinase (MLCK),
and desmin, while non-contractile proteins such as non-
muscle myosin, actin, tubulin, and vimentin are unaltered
(Smith et al. 1997b). The activity of contractile enzymes
such as MLCK are also increased (Smith et al. 1994). Me-
chanical strain also causes an alteration in both smooth mus-
cle shape and cytoskeletal structure. Cytoskeletal filaments
become more aligned, and the cells become elongated and
more tubular (Smith et al. 1997a). These changes are associ-
ated with increased force generation, calcium sensitivity
(Smith et al. 2000), increased velocity of shortening, and an
increased shortening capacity (Smith et al. 1999). These
cytoskeletal and functional changes have been shown to in-
volve RhoA activation, a key regulator of actin formation as
well as contractile function (Smith et al. 2003b).
We have shown that the cytoskeletal stiffness of cultured
smooth muscle cells is increased with mechanical strain.
Furthermore, we showed that mechanical strain increases the
response to contractile agonists by more than 50%, as mea-
sured by changes in cytoskeletal stiffness (Smith et al.
2003a). Recently, we have investigated whether the chroni-
cally strained ASM cells have an altered response to acute
oscillatory stretch. After 7 d of approximately 10% oscilla-
tory mechanical strain or 7 d of static culture, we lifted sin-
gle cells off their substrate using glass micropipettes
attached to a force transducer and length actuator (Smith et
al. 2000). We then exposed the individual cells to oscillatory
stretches of increasing amplitude (bottom of Fig. 1) similar
in fashion to studies previously performed on tracheal
© 2005 NRC Canada
Maksym et al.
Fig. 1. Acute stiffness following oscillatory cycling of activated
single airway smooth muscle (ASM) cells grown either in static
culture (䊉) or nearly 10% cyclic chronic strain (䊐) on collagen-
coated flexible membranes. After 7 d in culture, 6 representative
single cells from each condition were lifted from the membrane
using micromanipulators fitted with force and length transducers
and were subjected to 100 cycles of 1 Hz oscillatory strain at
2%, 4%, 8%, 10%, and returned to 2% strain sequentially. Stiff-
ness at each amplitude was calculated as the real part of the ra-
tio of the Fourier transforms of all 100 cycles of force and
displacement except for the final 2% strain where the stiffness
was computed from the last 25 cycles. Both control and chroni-
cally strained cells decreased in stiffness for strains greater than
4% (*p < 0.001). However, when stiffness was reduced to 2%
from 10%, control cells remained relaxed at 44% of their origi-
nal stiffness, while chronically strained cells recovered to more
than 85% of their original stiffness (#p < 0.01).
0.14
0.12
Stiffness (µN/µm)
0.10
0.08
#
0.06
0.04
0.02 * *
0.00
2 4 8 10 2'
Oscillation amplitude (%Lo)
strain
smooth muscle strips (Fredberg et al. 1997). We found that
cells both grown in static culture or grown with chronic cy-
clic strain relaxed significantly (p < 0.001, ANOVA, Fig. 1)
when oscillated with amplitudes of length oscillation >4%.
The relaxation is in agreement with the findings previously
reported in tissue strips (Fredberg et al. 1997). However,
when the oscillation was reduced again to 2%, the cell stiff-
ness of control cells remained depressed, but the stiffness of
the previously chronically strained cells significantly recov-
ered to more than 85% of their initial stiffness at 2% (Fig. 1).
This difference in stiffness recovery after the large oscilla-
tory stretch of 10% may be important as this behaviour
mimics the lack of maintained bronchodilatory response to a
deep inspiration in asthma. Although the strained cells are
stretched to the same extent as the control cells, control cells
maintained their stretch-induced relaxation while strained
cells rapidly regained their stiffness. In asthma, although the
airways are reportedly stretched by a deep inspiration, there
is no lasting bronchodilatory effect following the deep inspi-
ration, and the airways rapidly renarrow within a breath
(Jensen et al. 2001).
915
Cultured ASM cells exposed to stretch have features
similar to those observed in asthma or in cells obtained
from subjects with asthma
Not only does the response to length perturbation just de-
scribed have a behavioural correlate in asthma, but several
other features of the chronically mechanically strained ASM
are mimicked in asthma or airway hyperresponsiveness (Ta-
ble 1). It is largely agreed that increased ASM cell number
is a cardinal characteristic of asthma, although some recent
doubts have been raised (McParland et al. 2004). Also, ASM
cells from subjects with asthma are more proliferative (John-
son et al. 2001), correlating with increased proliferation in
response to strain. Furthermore, in asthma, airways narrow
too easily (asthmatics are more sensitive to inhaled agonists)
and too much (enhanced narrowing) (Woolcock et al. 1984),
corresponding to the increased sensitivity of mechanically
strained cells to calcium (Smith et al. 2000) and the in-
creased capacity of shortening, respectively (Smith et al.
1999). Recent data show that constitutive changes in ASM
occur in asthma, demonstrated by the increase in MLCK
protein from 2 to 4 times (Benayoun et al. 2003), which cor-
responds to strain-induced increases in culture (Table 1;
Smith et al. 1997b). Interestingly, although protein levels
were found to be increased in intermittent to moderate and
severe asthma (Benayoun et al. 2003), differing results mea-
suring mRNA for MLCK have been reported, finding either
a significant increase of MLCK with a molecular mass of
108 kDa in mild to moderate asthma (Ma et al. 2002) or no
difference in MLCK relative to healthy subjects (Ma et al.
2002; Woodruff et al. 2004).
MLCK is now known to occur with different molecular
masses, with a lower molecular mass form ranging between
108 and 155 kDa and a higher molecular mass form between
206 and 220 kDa (Blue et al. 2002). Both are widely ex-
pressed in various tissues with shorter MLCK more predom-
inant in ASM. Interestingly MLCK of the shorter form
decreases more rapidly with cell culture (Blue et al. 2002).
Functionally, shorter MLCK is more associated with smooth
muscle contraction (Bao et al. 2002), whereas longer MLCK
may have a role in stress fibre bundling (Blue et al. 2002).
In response to mechanical strain in cultured ASM cells,
we have preliminary data suggesting that the increase in
MLCK protein, irrespective of type, was considerably under-
estimated, and when factors such as strain profile were taken
into account, the effect was increased from a factor of 2 to a
factor of approximately 5 (discussed below). Increases in
MLCK are important functionally as MLCK (at least of the
shorter type) is rate limiting to actin-myosin binding, which
might directly explain increased velocity of shortening
(Smith et al. 1999) and possibly increased force-generation
capacity (Smith et al. 2000). Additionally, in agreement with
the increased MLCK, velocity of shortening of ASM taken
from patients with asthma is elevated (Ma et al. 2002) as it
is in mechanically strained cells (Smith et al. 1999). Airway
stiffness is functionally increased in asthma (Brackel et al.
2000), which may have many causes including airway re-
modelling, but this does correspond to the increased stiffness
we reported in mechanically strained cells (Smith et al.
2003a). Finally, the response to a deep inspiration in asthma
is impaired as we described above, which is similar to the
© 2005 NRC Canada
916 Can. J. Physiol. Pharmacol. Vol. 83, 2005

Table 1. A comparison of changes observed in cultured ASM exposed to cyclic mechanical strain compared with
selected characteristics of asthma or smooth muscle from subjects with asthma.
Changes due to cyclic mechanical strain of ASM Observations of airway function in asthma or from ASM
cells in vitro cells obtained from subjects with asthma
8 Proliferation (Smith et al. 1994) Hyperplasia in vivo (King et al. 1999) 8 cell proliferation
in culture (Johnson et al. 2001)
8 Ca2+ sensitivity (Smith et al. 2000) Airways narrow too easily (Woolcock et al. 1984)
8 Shortening capacity (Smith et al. 1999) Airways narrow too much (Woolcock et al. 1984) and
8 shortening capacity (Ma et al. 2002)
8 Shortening velocity (Smith et al. 1999) 8 Shortening velocity (Ma et al. 2002)
8 MLCK (-2×) (Smith et al. 1997b) 8 MLCK 2 to 4× (Benayoun et al. 2003; Ma et al. 2002)
8 Stiffness chronic (Smith et al. 2003a) and acute 8 Airway stiffness (Brackel et al. 2000)
(Deng et al. 2004b)
Softening response to stretch impaired (Maksym Dilatory response to DI impaired (Scichilone et al. 2000;
et al. 2001) and Fig. 1 Jensen et al. 2001)
Note: ASM, airway smooth muscle; MLCK, smooth muscle myosin light chain; 8, means the quantity is increased; and DI, deep
inspiration.

Fig. 2. Cells were mechanically stimulated by oscillating ferromagnetic beads adherent to the cytoskeleton of canine ASM cells in an
incubator for up to 2 h and samples removed at different time points, stiffness measured by optical magnetic twisting optometry
(OMTC) and stained for actin by phalloidin. (A) The stiffness (G′) increases with the time of mechanical stimulation compared with
unstimulated control cells, reaching a plateau after approximately 1 h. (B) The number of beads staining positively for actin above the
background increases with time and is much greater in mechanically stimulated cells than in unstimulated control cells. Actin also in-
creases with a similar time scale as the stiffness (from Deng et al. 2004b, reproduced with permission of Am. J. Physiol. Cell Physiol.,
Vol. 287, pp. C440–C448, © 2004 The American Physiological Society).
3.0
% beads with positive actin staining

MS (A) 100 (B)

MS
2.5
Controls
*
Controls *
* 80

* *
G' (Pa /nm)

2.0
60
1.5 *
40
1.0

0.5 20

0.0 0
0 5 15 30 60 120 0 5 15 30 60 120
Time (min) Time (min)

lack of depressed stiffness with a large amplitude stretch in
chronically strained cultured ASM cells. This last observa-
tion may be quite important. It suggests that somehow
chronic mechanical stretch may be involved in altering ASM
function in some way that makes an acute stretch no longer
beneficial, which is the behaviour observed in asthma. The
relationships summarized in Table 1 are not presented as
causal relationships, as the role of mechanical strain in vivo
on ASM function is not understood. However, the evidence
suggests that mechanical stress and strain may have a role in
altering ASM structure and function in asthma.
Measurements of mechanically induced cytoskeletal
remodelling in cultured ASM cells
To investigate more closely the ability for mechanical
stretch to influence ASM structure and function, we recently
measured the response of the ASM cytoskeleton to localized
oscillatory forces applied directly to the cytoskeleton via fo-
cal adhesions. We applied Arg-Gly-Asp (RGD) coated ferri-
magnetic beads to cells, which bind avidly within 15 min to
the cytoskeleton via integrin receptors, then applied oscilla-
tory torques to the beads (mechanical torque amplitude per
bead volume = 56 Pa at 0.3 Hz), resulting in mechanical
stimulation of the cytoskeleton continuously for up to 2 h.
We then measured the mechanical stiffness of the cells at
the sites of the applied stress using a method known as opti-
cal magnetic twisting cytometry. By using subpixel arithme-
tic, beads oscillated by magnetic fields can be tracked to sub-
2 nm resolution for the purposes of measuring cytoskeletal
mechanical properties (Fabry et al. 2001). This method has
been used in several studies with different cell types, and its
results are in close agreement with other methods such as

© 2005 NRC Canada

Maksym et al.
Fig. 3. Formation of different actin structures in response to 2 h
of cyclic mechanical stimulation by magnetically oscillating the
beads (dark circles) at 0.3 Hz with a mechanical torque per bead
volume = 56 Pa. Actin forms in a variety of structures such as
the (A) halo, (B) spindle, (C) filament, or (D) ring features
(from Deng et al. 2004b, reproduced with permission of Am. J.
Physiol. Cell Physiol., Vol. 287, pp. C440–C448, © 2004 The
American Physiological Society).
atomic force microscopy (Alcaraz et al. 2003; Deng et al.
2004b; Fabry et al. 2000; An et al. 2004). Furthermore, the
cytoskeletal (CSK) stiffness is linearly related to the traction
force of the cell on the substrate, indicating that the CSK
stiffness is a good measure of internal cell stress or prestress
(Wang et al. 2002).
We found that the stiffness (G′) increased by more than
2-fold within 1 h of stimulation compared with unstimulated
controls (Fig. 2A) (Deng et al. 2004b). By imaging similarly
stimulated cells that had been fixed and fluorescently la-
belled for actin, we found that actin accumulated at the
beads progressively with the duration of mechanical stimula-
tion (Fig. 2B). These data correspond to the increased CSK
stiffness, and agree with findings from other cell types using a
variety of mechanical perturbation techniques from bead pull-
ing to optical laser trapping. In particular, mechanical stresses
applied to fibroblasts and endothelial cells via surface-bound
beads have been found to induce bead-associated local re-
modelling in CSK proteins, including accumulation of actin
filaments and formation of focal adhesion complexes contain-
ing actin, vinculin, and talin (Glogauer and Ferrier 1998;
Glogauer et al. 1998; Chicurel et al. 1998; Galbraith et al.
2002). We also found that the actin remodelling near the
beads varied considerably (Fig. 3). Actin formed on beads
either only as a narrow hairline actin feature or as larger
more intense actin structures that appeared to link to other
structures within the cell. The larger more intense structures
were predominantly associated with mechanical stimulation,
while in unstimulated cells either no actin was discernible or
917
Fig. 4. Very few larger actin structures are found with KCl
(80 mmol/L) or acetylcholine (10–5 mol/L, not shown) stimula-
tion (Deng et al. 2004b) compared with mechanical stimulation
(white bars), whereas the number of hairline ring structures are
comparable (grey bars). For each group, n > 100 beads, and er-
ror bars indicate SE. Very few larger structures also occur in
time matched controls, numbering less than 11% of positively
stained beads (a portion of this data is from Deng et al. 2004b,
reproduced with permission of Am. J. Physiol. Cell Physiol.,
Vol. 287, pp. C440–C448, © 2004 The American Physiological
Society).
100
% beads accumulating actin
80
Hairline
60 rings
40
Total Large
structures
20
0
KCl Mech
80 mmol/L Stress
actin predominantly formed on the bead in a hairline struc-
ture. Furthermore, stimulation of the cells with contractile
agonists KCl (80 mmol/L) or acetylcholine (10–5 mol/L)
caused the number of beads accumulating actin to increase
to only 38.1% (SE 1.6%) and 60.7% (SE 2.0%), respec-
tively, which was less than that caused by mechanical stimu-
lation (68.1%, SE 1.6%) after 1 h (p < 0.01). However, there
was a large difference in the intensity and extent of the actin
accumulation at the beads. More than half of the beads me-
chanically stimulated formed large actin structures, while
contractile activation produced only a few large structures
(Fig. 4). These data indicate that contractile activation pro-
motes actin remodelling in agreement with other findings
(Mehta and Gunst 1999; Hirshman and Emala 1999), and
more importantly that mechanical stress is a potent stimulus
for actin remodelling and stiffening of the cytoskeleton in
ASM cells (Deng et al. 2004b).
Elevated tone in culture and in asthma
If mechanical stretch (strain) is beneficial in healthy indi-
viduals, why is strain provided by deep inspirations or large
breaths no longer beneficial in asthma, and why does me-
chanical strain in culture produce a myriad of deleterious ef-
fects in ASM? One perhaps underappreciated aspect is that
smooth muscle cells in culture have considerably elevated
contractile tone, a feature in common with asthma. Tone is
by definition a characteristic feature of asthma recognized
by reversible airway obstruction. That is, asthma is indicated
if an inhaled bronchodilator results in at least typically 20%
greater forced expired volume in 1 s, or FEV1.0, the stan-
dard measure of airway obstruction in asthma. The induced
© 2005 NRC Canada
918 Can. J. Physiol. Pharmacol. Vol. 83, 2005

Fig. 5. (A) Shows the cells roughly as they orient around the periphery of the flexible membrane. (B) Shows the strain profiles as a
function of position from the centre (0) to the edge of the flexible membrane (from Williams et al. 1992, adapted with permission of
J. Biomech. Eng., Vol. 114, pp. 377–384, © 1992 The American Society of Mechanical Engineers). Strain is nearly constant in the ra-
dial direction, but is decreasing from a maximum of 10% in the centre to 0 at the edge in the circumferential direction, increasing in
strain anisotropy to nearly uniaxial strain in the edge region indicated by the box. Note that the strain gradient or rate of change of
strain is also the greatest in this vicinity. (C) Images of cells after 7 d of 10% strain at 0.25 Hz under Hoffman contrast showing cells
from images taken at the centre, middle, and edge regions of the flexible membranes. Also shown are the directions and relative mag-
nitudes of the stretch in these regions.
A B Region of near
uniaxial strain
13

Radial strain
% Strain

3 Circumferential strain

-2 0 0.2 0.4 0.6 0.8 1

Normalized radial position

7 days stretch

airway dilation in asthma can be compared with the response
of cultured ASM cells to various relaxant agonists. Numer-
ous relaxant agonists are effective in reducing the stiffness
or contractile force exerted by ASM cells in culture by more
than 50% (Fabry et al. 2000; Maksym et al. 2000; Laporte
et al. 1999; Hubmayr et al. 1996).
In addition to reducing contractile force or stiffness, beta-
agonists also cause depolymerization of actin filaments of
ASM cells in culture (Hirshman et al. 2001). It is therefore
reasonable to suggest that the inhibition of contractile tone
by relaxant agonists might have an effect on the polymeriza-
tion of actin in response to mechanical stress. Indeed, when
we incubated cultured ASM cells with formoterol, a potent
long-acting beta-agonist, we found that mechanically in-
duced stiffening could be completely ablated (Deng et al.
2004a, 2005). That is, we saw no increase in mechanical stiff-
ness of the cytoskeleton with mechanical twisting stresses
applied for more than 2 h when relaxed with formoterol,
compared with the more than 2-fold increase we saw previ-
ously in the presence of contractile tone (Fig. 2A). While
not proven, this result is suggestive that other stress-
dependent effects could be modulated by the state of con-
tractile activation of the ASM cell. Is it possible that ele-
vated contractile tone in vivo turns mechanical stretch from
being beneficial in healthy individuals to maladaptive in
asthma? While it is too early to suggest this as a causal link
between the similar behaviours of strain and asthma listed in
Table 1, the evidence warrants further investigation. For
example, it would be important to establish whether the re-
duction of tone in cultured ASM cells can inhibit longer
term phenotypic changes such as those listed in the left col-
umn of Table 1.
Mechanically induced structure-function changes in
ASM are sensitive to strain profile
As described above and listed in Table 1, mechanical
strain of ASM cells in culture is generally reported to cause
maladaptive changes in ASM culture. However, early evi-
dence from those studies and a recent report by Wang et al.
(2004b) suggest that the type of oscillatory strain is critically
important in determining changes in contractile function of
cultured ASM cells. The early studies on the responses of
cultured cells to several days of mechanical strain were con-
ducted using a commercial system from Flexcell (Flexcell
International Corp., Hillsborough, N.C.). In this system,
cells are grown in 6-well plates on matrix (usually collagen
I)- coated silastic membranes, and oscillatory negative pres-
sure is provided under each well causing the membranes to
stretch downwards. However, the stretch is anisotropic over
the membrane, varying from purely biaxial and uniform in
the centre to approximately uniaxial in a circumferential di-
rection at the well edge. Strain in the radial direction is
roughly constant over the membrane, but strain in the cir-
cumferential direction decreases in a strain gradient to 0 at

© 2005 NRC Canada

Maksym et al.
Fig. 6. Cells stained to dectect myosin light chain kinase
(MLCK; green) using a 1:100 primary antibody dilution (Sigma
Chemical Co., St. Louis, Mo.), with secondary 1:300 dilution us-
ing Alexa 488 molecular probes with nuclei stained using 4′ ,
6-diamidino-2-phenylindole (DAPI) (blue). The nuclei have been
over-exposed for cell counting purposes, while the procedure for
staining for MLCK was identical for (A) and (B). (A) Cells near
the centre of the Flexcell silastic membranes have very little
MLCK protein, as do control cells that did not receive any strain
(not shown), while (B) cells stained near the periphery (2 mm
from edge) show much more positive staining for MLCK.
the edge (Fig. 5). Cells grown in media containing 10% se-
rum and exposed to 10% strain (measured in the centre in
any direction) organize themselves circumferentially around
the periphery of the wells in the Flexcell strain unit (Smith
et al. 1997a) but are not organized in the centre of the well
(Fig. 6). Cell alignment begins as early as day 4 of mechani-
cal strain and continues towards the centre until about day
11. The reasons for the alignment near the periphery are not
known. However, the region where cells become aligned is
also the location where the strain is highly anisotropic to the
point of being approximately uniaxial, and there is also a
strain gradient through this region. Since the contractile ap-
paratus of the ASM has a defined orientation, changes in
cell orientation with respect to the predominant strain direc-
tion could have important consequences for cell signalling.
Cyclic loading of the contractile lattice along the actin and
myosin filaments would likely be lessened as cells re-
oriented perpendicular to the strain while loading through
focal adhesions would be carried predominantly through
non-contractile cytoskeletal components, but these differ-
ences have not been studied.
In any case, most of the results from Table 1 (but not all)
were from cells that were aligned, and thus the maladaptive
pro-contractile changes reported are more likely due to
strain anisotropy or strain gradient than strain per se, as had
been sometimes previously reported. When differences in re-
sponse were examined with respect to position within the
membrane, myosin heavy chain (MHC) increased more at
the edge of the wells than in the centre, with the amount of
MHC matching that found in control cells, in agreement with
the changes largely arising because of strain anisotropy or
gradient effects. Conversely, MLCK showed less difference
with position in early measures using immunoblot techniques
919
(Smith et al. 1997b). However, in preliminary experiments
measuring MLCK as a function of position using immuno-
histochemical fluorescent imaging sensitive to either the short
or long MLCK, we found dramatic differences in MLCK
content. Following 11 d of strain, MLCK was increased only
in a narrow band approximately 2 mm wide near the edge.
The increase was more than 5 times the amount from cells in
other parts of the membrane and also about 5 times more
than that of control cells that did not receive any strain. In-
terestingly, even though most of the cells were aligned cir-
cumferentially after 11 d of strain over a region extending
more than 10 mm inwards from the edge, the increase in
MLCK was highly specific to a ring corresponding to ap-
proximately 10% radial strain and about 1% circumferential
strain. The reason for this specific location of enhanced con-
tractile protein expression is unknown, but could be impor-
tant given that remodelling of the airway in asthma likely
alters the strain profile experienced by the ASM cell and
that increases in MLCK protein are reported to occur in sub-
jects with asthma (Table 1).
It has been observed for many cell types that cells align in
response to anisotropic or uniaxial strain, including fibro-
blasts (Wang et al. 2004a) and vascular endothelial cells
(Wang et al. 2001; Yamada et al. 2000). Interestingly, the
cellular response to strain also depends on the orientation of
the applied strain relative to the cells. For example, in re-
sponse to uniaxial strain, fibroblasts forced to align in
grooves with the applied strain increase their expression of
alpha-actin, while fibroblasts forced to align in grooves per-
pendicular to the applied strain increase their secretory be-
haviour (Wang et al. 2004a). In response to biaxial strain, no
orientation behaviour is observed in fibroblast or ASM cells
(Smith et al. 2003a; Wang et al. 2004a). Thus, strain profile
has important morphological and functional consequences in
many cell systems.
This leads to the question “what strain profile is experi-
enced by ASM cells in vivo?” The ASM is largely oriented
circumferentially with an average slight angle near 12° (Lei
et al. 1997). However, Mead and colleagues showed that air-
way deformation was highly variable, but generally the air-
ways deform largely isotropically, increasing both in
diameter and length similarly with inspiration (Hughes et al.
1972). Thus in healthy normal lungs, strain on the ASM is
likely to be largely biaxial regardless of ASM orientation
around the airways. However, with constriction or elevated
tone that occurs in asthma, airway diameters become limited
(Brown and Mitzner 1996). Although the effect of tone on
bronchial length has not been clearly determined, as ASM
increases force and stiffness, lung inflation would tend to
preferentially lengthen airways rather than dilate them be-
cause of the primarily circumferential orientation of ASM.
Thus, more stretch would be conferred across the waists of
the ASM then the lengths. This closely matches the elevated
tone, anisotropic strain conditions, and cell alignment that
occur within the Flexcell system.
As mentioned, the complexity of the applied strain profile
was not fully appreciated in the early studies on ASM (Smith
et al. 1994, 1997a, 1997b, 1999, 2000). Indeed, since control
cells experienced no strain, the pro-contractile structure-
function changes (Table 1) were attributed only to changes
in strain magnitude. Last year, as we introduced above, quite
© 2005 NRC Canada
920
contrasting results were reported in ASM cells subjected to
purely biaxial strain (Wang et al. 2004b). Wang et al. found
that in response to uniform biaxial strain there was no align-
ment, no increase in proliferation (unlike in Table 1), an ap-
proximately 50% decrease in SM22 and smMHC promoter
activity, and a decreased filamentous to globular (monomeric)
actin ratio. These results are consistent with decreased Rho
activation, which is in contrast to the increased RhoA activa-
tion observed in response to complex anisotropic strain
(Smith et al. 2003b), and would appear to represent anti-
contractile effects of strain rather than pro-contractile effects
of strain as reported in the left column of Table 1. Thus, the
effects of mechanical strain in culture appear to be exqui-
sitely sensitive to strain profile. If sensitivity to strain profile
occurs in vivo, then the more biaxial strain present in healthy
lungs (Hughes et al. 1972) would tend to decrease contractile
function of ASM, while increased tone that likely limits bron-
chial dilation relative to lengthening causing strain anisotropy
would promote increased contractile function.
The sensitivity of cultured ASM cell structure to strain pro-
file was also evident in the narrow ring of increased MLCK at
roughly 10% anisotropic strain as described above (Fig. 6).
The heterogeneity in MLCK protein expression is reminis-
cent of the behaviour of ASM cells in response to long peri-
ods (14 d) of serum deprivation. With serum deprivation, a
subset of cells becomes locally aligned and has a 7-fold in-
crease in MLCK like the 5-fold increase we observed in pre-
liminary experiments (Halayko et al. 1999). Also, in our
case after 11 d of strain, the increase in MLCK was hetero-
geneous appearing as a patchy ring near 10% strain, but not
in all the aligned cells. What are the mechanisms driving
this change? Can pro-contractile phenotypic changes be re-
produced over a wider region of a controlled magnitude of
anisotropic strain? Furthermore, it is not understood whether
the differing responses to mechanical strain are due to aniso-
tropy alone or to the strain gradient present in the complex
anisotropic strain field (Fig. 5). These differing responses
need to be further examined by measuring responses to uni-
form strain, anisotropic strain, and uniaxial strain together
with measurements of contractile function to clarify the spe-
cific strain conditions contributing to maladaptive changes in
contractile structure and function.
Summary
This report highlights a current controversy in the role of
strain to the function of ASM. Strain is beneficial in the
healthy individual where there is no elevated contractile tone
or other humeral effects associated with asthma such as in-
flammation or matrix remodelling. However, strain is mal-
adaptive in cultured cells, promoting pro-contractile changes
in structure and function that mimic some of the functional
and structural changes that occur in asthma. These changes
occurred without most humeral factors associated with
asthma in vivo, but did occur in the presence of elevated
contractile tone present in cell culture. These changes also
required that the strain be either highly anisotropic or in-
clude a gradient of strain in one direction, which might have
important implications with tone and with the airway remod-
elling that is a characteristic feature of asthma. Localized
mechanical strain via twisting beads attached to the CSK
Can. J. Physiol. Pharmacol. Vol. 83, 2005
causes pronounced changes in actin structure and mechani-
cal stiffness after 1 h of twist. We have preliminary data
showing that inhibiting tone in culture via relaxant agonists
prevents mechanically induced changes in the cytoskeleton
within 2 h of stimulation. It remains to be seen whether
maladaptive changes that occur over many days can be like-
wise inhibited. However, the possibility that tone turns me-
chanical stress into a tool for maladaptive phenotypic
change in the ASM cell introduces a novel mechanism for
worsening asthma. If tone and stress lead to ASM cell re-
modelling, and this remodelling leads to further stiffening of
the ASM cells, this only contributes to increased stress, fur-
ther worsening asthma (Deng et al. 2005). It is interesting to
note that long-acting beta-agonists such as formoterol ad-
ministered twice daily improve airway function more than
the administration of corticosteroids alone (Sin et al. 2004).
Perhaps by relaxing the ASM, reducing the elevated tone in
asthma, these bronchodilators act to reduce ASM remodel-
ling in vivo.
Acknowledgements
The authors wish to thank Darren J. Cole for technical as-
sistance, Dr. Paul G. Smith for assistance with experimental
methods, and Dr. Jeffrey J. Fredberg, Harvard School of
Public Health, for providing the ferrimagnetic beads.
References
Alcaraz, J., Buscemi, L., Grabulosa, M., Trepat, X., Fabry, B.,
Farre, R., and Navajas, D. 2003. Microrheology of human lung
epithelial cells measured by atomic force microscopy. Biophys.
J. 84: 2071–2079.
An, S.S., Fabry, B., Mellema, M., Bursac, P., Gerthoffer, W.T.,
Kayyali, U.S., Gaestel, M., Shore, S.A., and Fredberg, J.J. 2004.
Role of heat shock protein 27 in cytoskeletal remodeling of the
airway smooth muscle cell. J. Appl. Physiol. 96: 1701–1713.
Bao, J., Oishi, K., Yamada, T., Liu, L., Nakamura, A., Uchida,
M.K., and Kohama, K. 2002. Role of the short isoform of myo-
sin light chain kinase in the contraction of cultured smooth mus-
cle cells as examined by its down-regulation. Proc. Natl. Acad.
Sci. U.S.A. 99: 9556–9561.
Benayoun, L., Druilhe, A., Dombret, M.C., Aubier, M., and
Pretolani, M. 2003. Airway structural alterations selectively as-
sociated with severe asthma. Am. J. Respir. Crit Care Med. 167:
1360–1368.
Blue, E.K., Goeckeler, Z.M., Jin, Y., Hou, L., Dixon, S.A., Her-
ring, B.P., Wysolmerski, R.B., and Gallagher, P.J. 2002. 220-
and 130-kDa MLCKs have distinct tissue distributions and
intracellular localization patterns. Am. J. Physiol. Cell Physiol.
282: C451–C460.
Brackel, H.J., Pedersen, O.F., Mulder, P.G., Overbeek, S.E.,
Kerrebijn, K.F., and Bogaard, J.M. 2000. Central airways be-
have more stiffly during forced expiration in patients with
asthma. Am. J. Respir. Crit. Care Med. 162: 896–904.
Brown, R.H., and Mitzner, W. 1996. Effect of lung inflation and
airway muscle tone on airway diameter in vivo. J. Appl. Physiol.
80: 1581–1588.
Brown, R.H., Scichilone, N., Mudge, B., Diemer, F.B., Permutt, S.,
and Togias, A. 2001. High-resolution computed tomographic
evaluation of airway distensibility and the effects of lung infla-
tion on airway caliber in healthy subjects and individuals with
asthma. Am. J. Respir. Crit. Care Med. 163: 994–1001.
© 2005 NRC Canada
Maksym et al.
Brusasco, V., and Pellegrino, R. 2003. Complexity of factors mod-
ulating airway narrowing in vivo: relevance to assessment of air-
way hyperresponsiveness. J. Appl. Physiol. 95: 1305–1313.
Chicurel, M.E., Chen, C.S., and Ingber, D.E. 1998. Cellular control
lies in the balance of forces. Curr. Opin. Cell Biol. 10: 232–239.
Deng, L., Fairbank, N.J., Connolly, S.C., Cole, D.J., Smith, P.G.,
and Maksym, G.N. 2004a. Long acting beta-adrenoceptor ago-
nist, formoterol, inhibits stiffening of cultured airway Smooth
muscle cells induced by mechanical stress. Am. J. Respir. Crit.
Care Med. 169: A448.
Deng, L., Fairbank, N.J., Fabry, B., Smith, P.G., and Maksym,
G.N. 2004b. Localized mechanical stress induces time-
dependent actin cytoskeletal remodeling and stiffening in cul-
tured airway smooth muscle cells. Am. J. Physiol. Cell Physiol.
287: C440–C448.
Deng, L., Fairbank, N.J., Cole, D.J., Fredberg, J.J., and Maksym,
G.N. 2005. Airway smooth muscle tone modulates mechanically
induced cytoskeletal stiffening and remodeling. J Appl. Physiol.
99: 334–341.
Fabry, B., Maksym, G.N., Butler, J.P., and Fredberg, J.J. 2000.
Contractile response of human airway smooth muscle cells in
culture. Am. J. Respir. Crit. Care Med. 161: A472.
Fabry, B., Maksym, G.N., Butler, J.P., Glogauer, M., Navajas, D.,
and Fredberg, J.J. 2001. Scaling the microrheology of living
cells. Phys. Rev. Lett. 87: 148102.
Fredberg, J.J. 2004. Bronchospasm and its biophysical basis in air-
way smooth muscle. Respir. Res. 5: 2.
Fredberg, J.J., Jones, K.A., Nathan, M., Raboudi, S., Prakash, Y.S.,
Shore, S.A., Butler, J.P., and Sieck, G.C. 1996. Friction in air-
way smooth muscle: mechanism, latch, and implications in
asthma. J. Appl. Physiol. 81: 2703–2712.
Fredberg, J.J., Inouye, D., Miller, B., Nathan, M., Jafari, S.,
Raboudi, S.H., Butler, J.P., and Shore, S.A. 1997. Airway
smooth muscle, tidal stretches, and dynamically determined
contractile states. Am. J. Resp. Crit. Care Med. 156: 1752–1759.
Galbraith, C.G., Yamada, K.M., and Sheetz, M.P. 2002. The rela-
tionship between force and focal complex development. J. Cell
Biol. 159: 695–705.
Glogauer, M., and Ferrier, J. 1998. A new method for application
of force to cells via ferric oxide beads. Pflueg. Arch. Eur. J.
Physiol. 435: 320–327.
Glogauer, M., Arora, P., Chou, D., Janmey, P.A., Downey, G.P.,
and McCulloch, C.A. 1998. The role of actin-binding protein
280 in integrin-dependent mechanoprotection. J. Biol. Chem.
273: 1689–1698.
Halayko, A.J., Camoretti-Mercado, B., Forsythe, S.M., Vieira, J.E.,
Mitchell, R.W., Wylam, M.E., Hershenson, M.B., and Solway, J.
1999. Divergent differentiation paths in airway smooth muscle
culture: induction of functionally contractile myocytes. Am. J.
Physiol. Lung Cell. Mol. Physiol. 20: L197–L206.
Hirshman, C.A., and Emala, C.W. 1999. Actin reorganization in
airway smooth muscle cells involves Gq and Gi-2 activation of
Rho. Am. J. Physiol. 277: 653–661.
Hirshman, C.A., Zhu, D.F., Panettieri, R.A., and Emala, C.W.
2001. Actin depolymerization via the beta-adrenoceptor in air-
way smooth muscle cells: a novel PKA-independent pathway.
Am. J. Physiol. Cell Physiol. 281: C1468–C1476.
Hubmayr, R.D., Shore, S.A., Fredberg, J.J., Planus, E., Panettieri,
R.A., Jr., Moller, W., Heyder, J., and Wang, N. 1996. Pharmaco-
logical activation changes stiffness of cultured human airway
smooth muscle cells. Am. J. Physiol. 271: 1660–1668.
Hughes, J.M., Hoppin, F.G., Jr., and Mead, J. 1972. Effect of lung
inflation on bronchial length and diameter in excised lungs. J.
Appl. Physiol. 32: 25–35.
921
Jensen, A., Atileh, H., Suki, B., Ingenito, E.P., and Lutchen, K.R.
2001. Selected contribution: airway caliber in healthy and asth-
matic subjects: effects of bronchial challenge and deep inspira-
tions. J. Appl. Physiol. 91: 506–515.
Johnson, P.R., Roth, M., Tamm, M., Hughes, M., Ge, Q., King, G.,
Burgess, J.K., and Black, J.L. 2001. Airway smooth muscle cell
proliferation is increased in asthma. Am. J. Respir. Crit. Care
Med. 164: 474–477.
Kapsali, T., Permutt, S., Laube, B., Scichilone, N., and Togias, A.
2000. Potent bronchoprotective effect of deep inspiration and its
absence in asthma. J. Appl. Physiol. 89: 711–720.
King, G.G., Pare, P.D., and Seow, C.Y. 1999. The mechanics of ex-
aggerated airway narrowing in asthma: the role of smooth mus-
cle. Respir. Physiol. 118: 1–13.
Laporte, J.D., Moore, P.E., Abraham, J.H., Maksym, G.N., Fabry,
B., Panettieri, R.A., Jr., and Shore, S.A. 1999. Role of ERK
MAP kinases in responses of cultured human airway smooth
muscle cells to IL-1beta. Am. J. Physiol. 277: L943–L951.
Latourelle, J., Fabry, B., and Fredberg, J.J. 2002. Dynamic equili-
bration of airway smooth muscle contraction during physiologi-
cal loading. J. Appl. Physiol. 92: 771–779.
Lei, M., Ghezzo, H., Chen, M.F., and Eidelman, D.H. 1997. Air-
way smooth muscle orientation in intraparenchymal airways. J.
Appl. Physiol. 82: 70–77.
Ma, X., Cheng, Z., Kong, H., Wang, Y., Unruh, H., Stephens, N.L.,
and Laviolette, M. 2002. Changes in biophysical and biochemi-
cal properties of single bronchial smooth muscle cells from
asthmatic subjects. Am. J. Physiol. Lung Cell. Mol. Physiol.
283: L1181–L1189.
Maksym, G.N., Fabry, B., Butler, J.P., Navajas, D., Tschumperlin,
D.J., Laporte, J.D., and Fredberg, J.J. 2000. Mechanical proper-
ties of cultured human airway smooth muscle cells from 0.05 to
0.4 Hz. J. Appl. Physiol. 89: 1619–1632.
Maksym, G.N., Fabry, B., Fredberg, J.J., Shue, G., and Smith, P.G.
2001. Fluctuation driven softening and plasticity in the isolated
smooth muscle cell. Am. J. Respir. Crit. Care Med. 163: A539.
McParland, B.E., Pare, P.D., Johnson, P.R., Armour, C.L., and
Black, J.L. 2004. Airway basement membrane perimeter in hu-
man airways is not a constant; potential implications for airway
remodeling in asthma. J. Appl. Physiol. 97: 556–563.
Mehta, D., and Gunst, S.J. 1999. Actin polymerization stimulated
by contractile activation regulates force development in canine
tracheal smooth muscle. J. Physiol. (Lond.) 519: 829–840.
Mitzner, W. 2004. Airway smooth muscle: the appendix of the
lung. Am. J. Respir. Crit. Care Med. 169: 787–790.
Mitzner, W., and Brown, R.H. 2000. Potential mechanism of
hyperresponsive airways. Am. J. Respir. Crit. Care Med. 161:
1619–1623.
Pellegrino, R., Violante, B., and Brusasco, V. 1996. Maximal
bronchoconstriction in humans. Relationship to deep inhalation
and airway sensitivity. Am. J. Respir. Crit. Care Med. 153: 115–
121.
Raboudi, S.H., Miller, B., Butler, J.P., Shore, S.A., and Fredberg,
J.J. 1998. Dynamically determined contractile states of airway
smooth muscle. Am. J. Respir. Crit. Care Med. 158: 176–178.
Salome, C.M., Thorpe, C.W., Diba, C., Brown, N.J., Berend, N.,
and King, G.G. 2003. Airway re-narrowing following deep in-
spiration in asthmatic and nonasthmatic subjects. Eur. Respir. J.
22: 62–68.
Scichilone, N., Kapsali, T., Permutt, S., and Togias, A. 2000. Deep
inspiration-induced bronchoprotection is stronger than broncho-
dilation. Am. J. Respir. Crit. Care Med. 162: 910–916.
Sin, D.D., Man, J., Sharpe, H., Gan, W.Q., and Man, S.F. 2004.
Pharmacological management to reduce exacerbations in adults
© 2005 NRC Canada
922
with asthma: a systematic review and meta-analysis. JAMA,
292: 367–376.
Skloot, G., and Togias, A. 2003. Bronchodilation and broncho-
protection by deep inspiration and their relationship to bronchial
hyperresponsiveness. Clin. Rev. Allergy Immunol. 24: 55–72.
Smith, P.G., Janiga, K.E., and Bruce, M.C. 1994. Strain increases
airway smooth muscle cell proliferation. Am. J. Respir. Cell
Mol. Biol. 10: 85–90.
Smith, P.G., Garcia, R., and Kogerman, L. 1997a. Strain reorga-
nizes focal adhesions and cytoskeleton in cultured airway
smooth muscle cells. Exp. Cell Res. 232: 127–136.
Smith, P.G., Moreno, R., and Ikebe, M. 1997b. Strain increases air-
way smooth muscle contractile and cytoskeletal proteins in vi-
tro. Am. J. Physiol. 272: L20–L27.
Smith, P.G., Roy, C., Dreger, J., and Brozovich, F. 1999. Mechani-
cal strain increases velocity and extent of shortening in cultured
airway smooth muscle cells. Am. J. Physiol. Lung Cell. Mol.
Physiol. 21: L343–L348.
Smith, P.G., Roy, C., Fisher, S., Huang, Q.Q., and Brozovich, F.
2000. Selected contribution: mechanical strain increases force
production and calcium sensitivity in cultured airway smooth
muscle cells. J. Appl. Physiol. 89: 2092–2098.
Smith, P.G., Deng, L., Fredberg, J.J., and Maksym, G.N. 2003a.
Mechanical strain increases cell stiffness through cytoskeletal
filament reorganization. Am. J. Physiol. Lung Cell. Mol.
Physiol. 285: L456–L463.
Smith, P.G., Roy, C., Zhang, Y.N., and Chauduri, S. 2003b. Me-
chanical stress increases RhoA activation in airway smooth mus-
cle cells. Am. J. Respir. Cell Mol. Biol. 28: 436–442.
Spector, A.A., Brownell, W.E., and Popel, A.S. 1996. A model for
cochlear outer hair cell deformations in micropipette aspiration
experiments: an analytical solution. Ann. Biomed. Eng. 24:
241–249.
Can. J. Physiol. Pharmacol. Vol. 83, 2005
Wang, J.H., Goldschmidt-Clermont, P., Wille, J., and Yin, F.C.
2001. Specificity of endothelial cell reorientation in response to
cyclic mechanical stretching. J. Biomech. 34: 1563–1572.
Wang, N., Tolic-Norrelykke, I.M., Chen, J., Mijailovich, S.M.,
Butler, J.P., Fredberg, J.J., and Stamenovic, D. 2002. Cell pre-
stress. I. Stiffness and prestress are closely associated in adher-
ent contractile cells. Am. J. Physiol. Cell Physiol. 282: C606–
C616.
Wang, J.H., Yang, G., Li, Z., and Shen, W. 2004a. Fibroblast re-
sponses to cyclic mechanical stretching depend on cell orienta-
tion to the stretching direction. J. Biomech. 37: 573–576.
Wang, L., Liu, H.W., McNeill, K.D., Stelmack, G., Scott, J.E., and
Halayko, A.J. 2004b. Mechanical strain inhibits airway smooth
muscle gene transcription via protein kinase C signaling. Am. J.
Respir. Cell Mol. Biol. 31: 54–61.
Williams, J.L., Chen, J.H., and Belloli, D.M. 1992. Strain fields on
cell stressing devices employing clamped circular elastic dia-
phragms as substrates. J. Biomech. Eng. 114: 377–384.
Woodruff, P.G., Dolganov, G.M., Ferrando, R.E., Donnelly, S.,
Hays, S.R., Solberg, O.D., Carter, R., Wong, H.H., Cadbury,
P.S., and Fahy, J.V. 2004. Hyperplasia of smooth muscle in mild
to moderate asthma without changes in cell size or gene expres-
sion. Am. J. Respir. Crit. Care Med. 169: 1001–1006.
Woolcock, A.J., Salome, C.M., and Yan, K. 1984. The shape of the
dose-response curve to histamine in asthmatic and normal sub-
jects. Am. Rev. Respir. Dis. 130: 71–75.
Yamada, T., Naruse, K., and Sokabe, M. 2000. Stretch-induced
morphological changes of human endothelial cells depend on
the intracellular level of Ca2+ rather than of cAMP. Life Sci. 67:
2605–2613.
© 2005 NRC Canada