| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
* Departments of Surgery and
* Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati, Ohio 45267-0558, USA; and
Shriners Hospital for Children, Cincinnati, Ohio 45267-0558, USA
1Correspondence: Department of Surgery, University of Cincinnati College of Medicine, 231 Bethesda Ave., Mail Location 0558, Cincinnati, OH 45267-0558, USA. E-mail: hasselp{at}uc.edu
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Key Words: muscle catabolism • actin • myosin • ubiquitin • proteasome • calpain
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
2
3
4)
. This metabolic response may be
deleterious to patients with sepsis because it can prevent ambulation,
thereby increasing the risk for life-threatening thrombo-embolic
complications, and may affect respiratory muscles (5)
,
increasing the risk for pneumonia and other pulmonary complications.
Understanding the mechanisms of sepsis-induced muscle catabolism,
therefore, has important clinical implications.
In previous studies we found evidence that muscle breakdown during
sepsis mainly reflects degradation of the myofibrillar proteins actin
and myosin (6)
and that the myofibrillar proteolysis is
regulated by the ubiquitin-proteasome pathway, both in experimental
animals (7
8
9)
and patients with sepsis (2)
.
An apparently conflicting observation reported recently is that the
ubiquitin-proteasome pathway does not degrade intact myofibrils. For
example, incubation of ovine skeletal muscle with proteasome did not
result in morphological or biochemical changes indicative of
myofibrillar breakdown (10)
. In other studies, isolated
muscle proteasomes degraded free actin and myosin but not intact
myofibrils (11)
. These studies suggest that actin and
myosin need to be released from the myofibrils before they can be
ubiquitinated and degraded by the 26S proteasome.
Evidence
for Z-band disruption and release of myofilaments in skeletal muscle has
been reported previously in certain catabolic conditions, including
fasting and treatment with glucocorticoids (12)
. There is
evidence that release of myofilaments from the myofibrils reflects
calcium-dependent calpain activity (13
14
15)
. The influence
of sepsis on Z-band integrity and release of myofilaments is not known.
In the present study we tested the hypothesis that sepsis results in
increased expression of calpains, disruption of Z-bands, and
calcium-dependent release of myofilaments in skeletal muscle.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
7
8
9)
. Control animals were sham operated,
i.e., they underwent laparotomy and manipulation, but no ligation or
puncture of the cecum. All animals were resuscitated with 10 ml/100 g
b.w. sterile saline administered subcutaneously (s.c.) on the back at
the time of surgery. The rats had free access to drinking water, but
food was withheld after the surgical procedure to avoid the influence
of reduced food intake (as noted in septic rats) on metabolic changes.
Groups of rats were studied at intervals up to 16 h after sham
operation or CLP.
The experimental model used here resembles the situation in many
surgical patients with sepsis caused by intra-abdominal abscess and
devitalized tissue. In previous studies, cultures of blood and
peritoneal fluid 16 h after CLP showed a bacterial flora commonly
seen in patients with septic peritonitis, i.e., a flora of aerobic and
anaerobic bacteria (16
, 17)
. The model has been
characterized in previous studies from our and other laboratories with
regard to survival rates, bacteriology, and hemodynamic changes
(16
, 17)
. In several previous reports we found that
ubiquitin-proteasome-dependent muscle protein breakdown was increased
after CLP in rats (7
8
9)
. Small rats were used because
they possess muscles that are thin enough to allow for metabolic
studies during incubation in vitro (18
, 19)
and
rats of this size were used in previous studies to examine the
influence of sepsis on muscle protein breakdown (6
7
8
9)
.
Four series of experiments were performed. In the first series of
experiments, extensor digitorum longus (EDL) and soleus muscles were
harvested from rats 16 h after sham operation or CLP for
morphological studies using electron microscopy. These muscles were
studied because in previous reports (6
, 8)
we found
evidence that the effect of sepsis on protein degradation is more
pronounced in white, fast-twitch muscle (such as the EDL muscle) than
in red, slow-twitch muscle (such as the soleus muscle). In the second
series of experiments, release of myofilaments was measured in the same
muscles at intervals of up to 16 h after CLP or sham operation. In
the third series of experiments, the gene expression of µ-calpain,
m-calpain, and p94 (20)
was examined in EDL muscles
16 h after CLP or sham operation.
Finally, the role of calcium in sepsis-induced release of
myofilaments was assessed by treating rats with dantrolene, which is a
substance that blocks the release of calcium from intracellular stores
(21)
. Rats were injected s.c. on the back with 10 mg/kg
dantrolene or corresponding volume of solvent (7.5% mannitol/NaOH
solution, pH 9.0) 2 h before and 8 h after sham operation or
CLP, and release of myofilaments was determined in EDL muscles 16 h after CLP or sham operation. Four experimental groups were included:
1) sham operation + vehicle; 2) sham operation +
dantrolene; 3) CLP + vehicle; and 4) CLP +
dantrolene. The dose of dantrolene used here was based on previous
reports in which this treatment influenced sepsis-induced metabolic
changes (22)
.
Electron microscopy
Sixteen hours after sham operation or CLP, EDL and soleus
muscles were fixed at 4°C in 2% glutaraldehyde for 2 h,
followed by 1 h in ice-cold 1% OsO4. Both
fixatives were made up in 0.1 M cacodylate buffer (pH 7.2) with 0.1 M
sucrose. The tissue was bloc-stained with 2% uracyl acetate in 10%
ethanol. The specimens were dehydrated in increasing concentrations of
ethanol (50–100%), passed through propylene oxide, and embedded in
Spurr (Electron Microscopy Sciences, Fort Washington, Pa.). Tissue
sections were prepared with a Reichert ultramicrotome (Reichert,
Vienna, Austria). The sections were contrasted with lead citrate and
studied in a Hitachi H-600 transmission electron microscope (Hitachi,
Tokyo, Japan) operated at 60 kV.
In addition to providing an overall picture of sepsis-induced changes
in the ultrastructure of the muscles, the electron microscopy studies
focused on Z-band morphology. To assess Z-band thickness, the ratio
between Z-band and A-band thickness in the same sarcomere was
determined in 10 random fields from 4 control and 4 septic soleus and
EDL muscles. These measurements were performed in a blinded fashion,
i.e., the person who performed the determinations was unaware from
which group of rats the muscles originated. Assessing changes in Z-band
thickness by determining the ratio between the Z-band and A-band
provides a measure that is unaffected by the state of muscle
contraction or orientation of the section, thereby standardizing the
measurements of Z-band thickness (23
, 24)
.
Release of myofilaments
Release of myofilaments from the myofibrils was determined by
measuring the fraction of `easily releasable myofilaments' as
described in detail elsewhere (12
, 25
26
27)
, with minor
modifications. First, the myofibrillar proteins were isolated by
soaking the muscles at 4°C in low-salt buffer (LSB; 0.1 M KCl, 2 mM
MgCl2, 2 mM EGTA, 0.5 mM dithiothreitol, 10 mM
Tris-maleate, pH 7.0) containing 1% Triton X-100 for 90 min with three
changes of the soaking solution. After soaking for 90 min, the muscles
were homogenized at 4°C in the same solution using a Polytron
homogenizer (Brinkman, Westbury, N.Y.). The homogenate was centrifuged
at 1500 x g for 10 min and the pellet was resuspended with a
Pasteur pipette in 10 ml of LSB containing 1% Triton X-100, filtered
through two layers of gauze cloth, and recentrifuged. The resulting
myofibrillar pellet was washed once in LSB containing 1% Triton X-100
and three times in LSB.
The easily releasable myofilaments were extracted from the myofibrillar
fraction by repeated pipetting (10 passages through a Pasteur pipette)
in 1.5 ml of LSB containing 5 mM ATP. The suspension was layered over
0.75 ml LSB containing 20% glycerol in a conical tube and centrifuged
at 1,500 g for 10 min. The supernatant, including the
glycerol-containing layer, was collected with a Pasteur pipette and was
centrifuged through 0.5 ml LSB containing 20% glycerol. The final
supernatant contained the released myofilaments and the pellet the
residual myofibrillar fraction. Protein was determined in both
fractions according to Lowry et al. (28)
and the easily
releasable myofilaments were expressed as a percentage of the combined
amount of protein in the two fractions.
To further characterize the proteins in the myofibrillar and
myofilament fractions, proteins were separated by sodium dodecyl
sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as described by
Laemmli (29)
. Electrophoresis was carried out at 25
mA/slab (12 x 15 cm). Gels were stained overnight in 0.05%
Coomassie blue, 25% isopropyl alcohol, and 10% acetic acid and
destained by soaking in 10% acetic acid.
Calpain mRNA levels
mRNA levels for µ- and m-calpain and p94 were measured by dot
blot hybridization. The hybridization was performed under stringent
conditions in 50% formamide and 5x SSC (1xSSC = 0.15 M NaCl,
0.015 M Na-citrate) at 56°C. The cDNA probes were randomly labeled
with digoxigenin (DIG)-11-dUTP (Boehringer Mannheim, Indianapolis,
Ind.). RNA was extracted from EDL muscles by the guanidinium
thiocyanate-phenol-chloroform method (30)
using an RNA
STAT-60 kit (Tel-Test `B', Inc., Friendship, Tex.). Different amounts
(20, 10, and 5 µg) of RNA were loaded onto a nylon membrane
(Boehringer Mannheim) using a Minifold II slot-blot filtration manifold
(Bio-Rad, Hercules, Calif.) and fixed to the membrane by UV
cross-linking for 5 min. Prehybridization was performed at 56° for
4 h in a buffer consisting of 50% formamide, 7% SDS, 50 mM
Na-phosphate (pH 7.0), 2% blocking reagent (Boehringer Mannheim), 5x
SSC, and 0.1% N-lauroylsarcosine. Hybridization was carried out
overnight at 56°C in the same buffer containing 25 ng/ml of
DIG-labeled cDNA probe. After two posthybridization washes in 2xSSC
and 0.1% SDS for 5 min at room temperature and in 0.1x SSC and 0.1%
SDS for 15 min at 68°C, chemiluminescent detection of bound probe was
carried out using 
-DIG alkaline phosphatase-conjugated Fab fragment
(37.5 mU/ml; Boehringer Mannheim) and the substrate CDP-Star (0.25 mM;
Boehringer Mannheim). The membranes were then exposed to X-ray film
(X-Omat, Eastman Kodak, New Haven, Conn.) and the signal intensities
were determined by densitometry.
cDNA probes for rat µ- and m-calpain and p94 were generated by
performing reverse transcriptase polymerase chain reaction as described
previously (31)
. The specificity of the probes was
determined by Northern blotting. All probes hybridized a single band
corresponding to the expected size (not shown). A rat 18S ribosomal
probe (31)
was used to normalize the mRNA levels.
Statistics
Results are given as means ± SE. Statistical
significance was determined by Student's t test or analysis
of variance, followed by Tukey's test.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
). In some sections, there was a loss of registry between neighboring
sarcomeres. Z-band streaming and thickening and complete loss of some
Z-bands were seen in some sections of septic EDL muscles (Fig. 1)
. The
ratio between Z-band and A-band thickness in the same sarcomere was
determined in 10 random fields per muscle in EDL muscles from 4 control
and 4 septic rats by an observer who was unaware of whether the muscles
were from septic or control rats. The ratio was 6.1 ± 0.2 x
10-2 and 8.4 ± 0.4 x
10-2 in control and septic muscle, respectively
(P<0.05). A large number of the mitochondria appeared
swollen in EDL muscles from septic rats. In contrast, no apparent
morphological changes were seen in soleus muscles from sham-operated or
septic rats (Fig. 2
). The Z-band to A-band ratio was 8.1 ± 0.4 x
10-2 and 8.0 ± 0.4 x
10-2 in control and septic soleus muscle,
respectively (N.S.).
|
|
Sixteen hours after CLP, the fraction of easily releasable myofilaments
was increased by ~50% in EDL muscles but was not significantly
altered in soleus muscles (Fig. 3
). To determine how soon after induction of sepsis the release of
myofilaments increased, measurements were made in EDL muscles from
groups of rats 4 h, 8 h, and 16 h after sham operation
or CLP. Results showed that the fraction of easily releasable
myofilaments was increased 8 h after induction of sepsis and
remained elevated throughout the remainder of the experimental period
(Fig. 4
).
|
|
Because the easily releasable myofilaments were isolated from the
myofibrillar proteins in a medium containing ATP, it is possible that
muscles from septic rats were more sensitive in vitro to ATP
than control muscles, and that the difference between control and
septic muscle merely reflected a difference in sensitivity to ATP. We
therefore tested the effect of different ATP concentrations in the
relaxing buffer on the amount of myofilaments released in control and
septic muscles. The release of myofilaments increased with increasing
ATP concentrations in the relaxing buffer in muscles from both
sham-operated and septic rats and the difference between septic and
control muscles was not dependent on the ATP concentration (Fig. 5
).
|
To further characterize the proteins in the fraction of easily
releasable myofilaments, proteins were separated by SDS-PAGE. Proteins
with molecular weights corresponding to the molecular weights of myosin
heavy chain and actin were present at higher concentrations among the
easily releasable myofilaments from septic EDL muscles than from
control muscles; these proteins were the predominant, although not
exclusive, proteins among the easily releasable myofilaments
(Fig. 6
). In addition to increased release of myosin heavy chain and actin,
SDS-PAGE indicated that titin and several breakdown products of titin
as well as desmin and -actinin were present in the easily releasable
myofilament fraction at increased concentrations in muscles from septic
rats.
|
Because previous studies suggest that release of myofilaments from the
myofibrils is regulated by calcium-dependent calpain activity in
certain other catabolic conditions (13
14
15)
, we next
determined mRNA levels for µ- and m-calpain and p94 in EDL muscles
from sham-operated and septic rats. Sixteen hours after induction of
sepsis, the expression of all three calpains was substantially
increased, with the most marked increase (threefold) noted for
µ-calpain (Figs. 7
8
9
).
|
|
|
To further test the role of calcium in sepsis-induced release of
myofilaments, the effect of the calcium antagonist dantrolene on the
release of myofilaments was examined. Treatment of rats with dantrolene
prevented the sepsis-induced increase in myofilament release in EDL
muscles (Fig. 10
). Dantrolene had no effect on the fraction of easily releasable
myofilaments in muscles from sham-operated rats.
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Sepsis-induced changes in skeletal muscle morphology were reported
previously in experimental animals (32
, 33)
and humans
(34)
, and consisted mainly of mitochondrial swelling,
atrophy, and segmental necrosis. Changes in Z-band morphology were not
commented on in those studies, but close inspection of figures suggest
that Z-band disintegration probably occurred as well.
Current concepts of the architecture and function of the muscle
sarcomere were reviewed recently (35)
. The Z-disks serve
to anchor and organize the myofilaments and to mechanically link actin
from one sarcomere to the next sarcomere along the myofibril. There is
evidence that -actinin is involved in the anchorage of actin to the
Z-disk and in the cross-linking of one actin molecule to the next actin
molecule in the adjacent sarcomere (36)
. Actin and
-actinin account for a large percentage of the proteins in the
Z-disk, but other proteins are present as well. For example, myosin is
anchored to the Z-disk by titin and the titin filaments extend to the
center of the Z-disk (37)
. It is easy to understand,
therefore, that disintegration/disruption of the Z-disk can result in
the release of actin and myosin.
The changes in Z-disk morphology noted in EDL muscles from septic rats
in the present study were similar to those reported previously in
muscle atrophy due to denervation or muscular dystrophy (38
, 39)
. Intense exercise has also been reported to result in muscle
damage characterized by Z-band streaming, disintegration, and
disappearance, with loss of the transverse register of the Z-bands
(40
, 41)
. Muscle catabolism caused by denervation was
associated with early degradation and gradual disappearance of Z-disks
whereafter actin and myosin filaments became disordered and eventually
dispersed, leaving large vacant areas in the sarcomere
(38)
. Thus, Z-disk disintegration and disruption of the
anchorage of myofilaments to the Z-disk may be a common response in
skeletal muscle to different catabolic conditions, including sepsis.
In addition to morphological changes, there is also biochemical
evidence that myofilaments are released from myofibrils at an increased
rate during certain catabolic conditions. In previous work, gentle
treatment of isolated myofibrils from rat skeletal muscle with an
ATP-containing relaxing solution released a small amount of
myofilaments constituting ~5–10% of total myofibrillar proteins
(25
, 26)
. Using this method, Dahlmann et al.
(12)
studied the influence of starvation or treatment with
corticosterone on the amount of easily releasable myofilaments in rat
skeletal muscle. Both experimental conditions increased the amount of
easily releasable myofilaments in gastrocnemius and EDL muscles
(fast-twitch muscles) but not in soleus muscles (a slow-twitch muscle).
The selective increase in myofilament release in fast-twitch muscle
noted during starvation, after treatment with corticosterone
(12)
, and during sepsis (present study) parallels the
predominant increase in protein breakdown in fast-twitch muscle during
sepsis (6
, 8)
and in other catabolic conditions (42
, 43)
.
Although inhibited myofilament release after treatment of septic rats
with dantrolene suggests that calcium is involved in the regulation of
sepsis-induced release of actin and myosin from the myofibrils, the
mechanism by which calcium regulates the catabolic response in muscle
is not known from the present study. It is likely, however, that the
result reflected involvement of one or several of the
calcium-activated calpains (20
, 41
, 44)
. The
concentration of calpains is high in the vicinity of Z-disks in
skeletal muscle. The present result of increased mRNA levels for µ-
and m-calpain suggests that increased activity of these proteases
during sepsis, as reported previously (45)
, may at least
in part reflect increased amounts of the enzymes. The muscle-specific
calpain, p94 (46)
, has binding sites on titin, raising the
possibility that titin may be subjected to calcium-dependent
proteolysis, thereby disrupting the anchorage of myosin to the Z-disk
(37)
. The present study provides the first evidence that
sepsis may be associated with increased expression of p94.
Increased calcium uptake and content in skeletal muscle during sepsis,
as reported by us (47)
and others (48)
, as
well as stimulated total protein breakdown after treatment of incubated
muscles in vitro with calcium or the ionophore A23187
(47
, 49
, 50)
are consistent with a role of calcium in
sepsis-induced muscle catabolism. Because treatment of muscles in
vitro with calcium antagonists in other studies did not inhibit
the sepsis-induced increase in total or myofibrillar protein breakdown,
the role of calcium in the regulation of muscle proteolysis during
sepsis was questioned (47)
. It should be noted, however,
that the lack of effect of dantrolene in vitro on
proteolysis in incubated muscles from septic rats (47)
does not rule out a role of calcium in sepsis-induced muscle breakdown.
If calcium regulates the release of myofilaments, as suggested by the
present results, rather than the actual degradation of myofilaments, it
is not surprising that calcium antagonists do not inhibit protein
breakdown in muscles in which the release of myofilaments has already
taken place.
The present study provides the first evidence of Z-band disintegration
and release of myofilaments from myofibrils in skeletal muscle during
sepsis. Taken together with previous evidence that the proteasome does
not degrade intact myofibrils (10
, 11)
, the results are
consistent with the concept that actin and myosin are released from the
myofibrils before they are ubiquitinated and degraded by the 26S
proteasome. Thus, it is possible that the increased expression and
activity of the ubiquitin-proteasome proteolytic pathway may be the
result of an increased amount of substrates available rather than the
cause of sepsis-induced muscle breakdown. This would put the
ubiquitin-proteasome pathway in a different perspective and may suggest
that this mechanism is important for the `clean-up' in septic muscle,
and perhaps other catabolic muscles as well, rather than being the
cause of muscle breakdown. This is interesting considering the role
that was initially ascribed to the ubiquitin-proteasome pathway,
namely, a mechanism by which cells can degrade and dispose of abnormal
proteins (51)
.
In previous studies, there was a relationship between the half-life of
a protein and the identity of its amino-terminal residue, the `N-end
rule' (52)
. According to this rule, proteins with basic
amino-terminal residues (Arg, Lys, His), bulky hydrophobic
amino-terminal residues (Phe, Leu, Trp, Tyr, Ile), or with acidic
NH2 termini that have undergone
arginyl-tRNA-dependent amino-terminal arginylation are recognized by
the ubiquitin-protein ligase E3, which is the predominant E3 enzyme
in skeletal muscle. Since 14 kDa ubiquitin-conjugating enzyme,
E214 k, is associated with E3
(53)
, a recent report of increased expression of
E214 k in septic muscle (54)
supports the concept that the N-end rule pathway accounts at least in
part for sepsis-induced protein breakdown in skeletal muscle.
In a recent study using specific inhibitors of the ubiquitin
ligase E3, evidence was found that a major fraction of
non-myofibrillar protein degradation in skeletal muscle was catalyzed
by the N-end rule pathway, whereas intact actin and myosin were not
degraded by this mechanism (55)
. In a subsequent report
from the same laboratory, results suggested that ubiquitin conjugation
increases in cachectic muscle due to activation of the N-end rule
pathway (56)
. It was speculated that a rate-limiting step
in the degradation of long-lived proteins (mainly actin and myosin)
is an exo- or endoproteolytic cleavage that exposes a destabilizing
amino-terminal residue (55
, 56)
. Based on the results in
the present report, we propose that calcium-dependent, calpain-mediated
release of myofilaments from the Z-disks may be such a rate-limiting
step in sepsis-induced muscle protein degradation. This could have
important clinical implications because it would suggest that
inhibition of myofilament release may be a more logical approach in
preventing and treating muscle catabolism than inhibition of the
proteasome pathway. It will be important in future studies to establish
the link between calcium-dependent release of myofilaments and
ubiquitin-proteasome-dependent proteolysis in skeletal muscle during
sepsis.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
Received for publication December 16, 1998.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
This article has been cited by other articles:
|
|
 |
|
  D. E. Goll, G. Neti, S. W. Mares, and V. F. Thompson Myofibrillar protein turnover: The proteasome and the calpains J Anim Sci, April 1, 2008; 86(14_suppl): E19 - E35. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. A. Harrigan, J. Piotrowski, L. Di Noto, R. L. Levine, and V. A. Bohr Metal-catalyzed Oxidation of the Werner Syndrome Protein Causes Loss of Catalytic Activities and Impaired Protein-Protein Interactions J. Biol. Chem., December 14, 2007; 282(50): 36403 - 36411. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. W. H. van Hees, H. F. M. van der Heijden, C. A. C. Ottenheijm, L. M. A. Heunks, C. J. C. Pigmans, F. W. A. Verheugt, R. M. H. J. Brouwer, and P. N. R. Dekhuijzen Diaphragm single-fiber weakness and loss of myosin in congestive heart failure rats Am J Physiol Heart Circ Physiol, July 1, 2007; 293(1): H819 - H828. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. A. C. Ottenheijm, L. M. A. Heunks, Y.-P. Li, B. Jin, R. Minnaard, H. W. H. van Hees, and P. N. R. Dekhuijzen Activation of the Ubiquitin-Proteasome Pathway in the Diaphragm in Chronic Obstructive Pulmonary Disease Am. J. Respir. Crit. Care Med., November 1, 2006; 174(9): 997 - 1002. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. U. Fareed, A. R. Evenson, W. Wei, M. Menconi, V. Poylin, V. Petkova, B. Pignol, and P.-O. Hasselgren Treatment of rats with calpain inhibitors prevents sepsis-induced muscle proteolysis independent of atrogin-1/MAFbx and MuRF1 expression Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2006; 290(6): R1589 - R1597. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 O. Zolk, C. Schenke, and A. Sarikas The ubiquitin-proteasome system: Focus on the heart Cardiovasc Res, June 1, 2006; 70(3): 410 - 421. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. GISSEL The Role of Ca2+ in Muscle Cell Damage Ann. N.Y. Acad. Sci., December 1, 2005; 1066(1): 166 - 180. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. N. Mehiri, E. Barreiro, M. Hayot, M. Voyer, A. S. Comtois, A. E. Grassino, and G. Czaika Time-based gene expression programme following diaphragm injury in a rat model Eur. Respir. J., March 1, 2005; 25(3): 422 - 430. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. Wei, M. U. Fareed, A. Evenson, M. J. Menconi, H. Yang, V. Petkova, and P.-O. Hasselgren Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3 Am J Physiol Regulatory Integrative Comp Physiol, March 1, 2005; 288(3): R580 - R590. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. J. Durham, S. L. Miller, C. W. Yeckel, D. L. Chinkes, K. D. Tipton, B. B. Rasmussen, and R. R. Wolfe Leg glucose and protein metabolism during an acute bout of resistance exercise in humans J Appl Physiol, October 1, 2004; 97(4): 1379 - 1386. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. W. Jackman and S. C. Kandarian The molecular basis of skeletal muscle atrophy Am J Physiol Cell Physiol, October 1, 2004; 287(4): C834 - C843. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. BAR-SHAI, E. CARMELI, R. COLEMAN, and A. Z. REZNICK Mechanisms in Muscle Atrophy in Immobilization and Aging Ann. N.Y. Acad. Sci., June 1, 2004; 1019(1): 475 - 478. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. S. Criswell, R. A. Shanely, J. J. Betters, M. J. McKenzie, J. E. Sellman, D. L. Van Gammeren, and S. K. Powers Cumulative Effects of Aging and Mechanical Ventilation on In Vitro Diaphragm Function Chest, December 1, 2003; 124(6): 2302 - 2308. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. E. Helman, E. Huff-Lonergan, G. M. Davenport, and S. M. Lonergan Effect of dietary protein on calpastatin in canine skeletal muscle J Anim Sci, September 1, 2003; 81(9): 2199 - 2205. [Abstract] [Full Text] [PDF]
|
|
|
|
<
7 for each
data point. *P<0.05 vs. sham at corresponding time
point.
