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MULTI-AUTHOR REVIEW
The response of human skeletal muscle tissue to hypoxia
Carsten Lundby ÆJose A. L. Calbet Æ
Paul Robach
Received: 18 August 2009 / Accepted: 20 August 2009 / Published online: 10 September 2009
ÓBirkha
¨user Verlag, Basel/Switzerland 2009
Abstract Hypoxia refers to environmental or clinical
settings that potentially threaten tissue oxygen homeostasis.
One unique aspect of skeletal muscle is that, in addition to
hypoxia, oxygen balance in this tissue may be further
compromised when exercise is superimposed on hypoxia.
This review focuses on the cellular and molecular responses
of human skeletal muscle to acute and chronic hypoxia, with
emphasis on physical exercise and training. Based on pub-
lished work, it is suggested that hypoxia does not appear to
promote angiogenesis or to greatly alter oxidative enzymes
in skeletal muscle at rest. Although the HIF-1 pathway in
skeletal muscle is still poorly documented, emerging evi-
dence suggests that muscle HIF-1 signaling is only activated
to a minor degree by hypoxia. On the other hand, combining
hypoxia with exercise appears to improve some aspects of
muscle O
2
transport and/or metabolism.
Keywords Altitude Hypoxic Gene Protein
Adaptation Expression Capillary Training
Introduction
Exposure to high altitude refers to an environmental con-
dition associated with whole body and tissue hypoxia,
resulting from a drop in barometric pressure and hence a
concomitant decrease in arterial oxygen availability. Such
a condition is encountered by nearly 140 million people
residing at high altitude worldwide [1] and also by
numerous sea-level dwellers traveling or commuting to
high altitude areas. Healthy humans studied at high altitude
not only contribute to unravel the molecular and systemic
mechanisms involved in O
2
sensing and adaptive responses
to the low oxygen environment, but also help in the
understanding of various pathological situations associated
with hypoxia. These pathologies include anemia and
chronic obstructive pulmonary disease, as well as chronic
heart failure. In addition, tumor growth and development
can be associated with tissue hypoxia, since solid tumors
become hypoxic as they grow larger [2]. Yet another sit-
uation that potentially leads to hypoxia is physical exercise.
Even during normoxic conditions, oxygen availability can
drop in the transition from rest to exercise, implying that
exercising skeletal muscle must operate at a very low
partial pressure of oxygen, estimated to be approximately
*3 mmHg [3].
Whatever the origin of the hypoxic stimulus may be
(environmental or pathological), the ultimate consequence
is an inadequate O
2
delivery/availability at the tissue level,
implying that tissue demand exceeds its O
2
supply. All
nucleated cells in the human body are able to sense O
2
and
to respond to O
2
deficiency in order to maintain homeo-
stasis. The main mediator of cellular hypoxia is the
hypoxia inducible factor (HIF) pathway, discovered by the
group of Semenza [4]. Like other tissues, resting skeletal
muscle homeostasis is challenged during hypoxic
C. Lundby (&)
The Copenhagen Muscle Research Centre,
Rigshospitalet Sect. 7652, 2100 Copenhagen, Denmark
e-mail: Lundby@idraet.au.dk
C. Lundby
Department of Sport Science, University of A
˚rhus,
8200 A
˚rhus N, Denmark
J. A. L. Calbet
Department of Physical Education,
University of Las Palmas de Gran Canaria,
Las Palmas de Gran Canaria, Spain
P. Robach
Ecole Nationale de Ski et d’Alpinisme,
74401 Chamonix, France
Cell. Mol. Life Sci. (2009) 66:3615–3623
DOI 10.1007/s00018-009-0146-8 Cellular and Molecular Life Sciences
exposure, either acutely or chronically. However, differ-
ently from other tissues, muscle function may be further
compromised if exercise is superimposed during hypoxic
exposure.
This review aims to characterize the particular response
of human skeletal muscle tissue to hypoxia, both during
acute and chronic exposure to hypoxia with emphasis on
exercise. Special attention will be given to skeletal muscle
gene expression and protein content, whereas the regula-
tion of metabolism and oxygen delivery will not be
discussed.
The HIF-1 pathway and its presence within human
skeletal muscle
HIF-1 is a heterodimeric protein belonging to the basic
helix-loop-helix-PAS family of transcription factors. This
protein is composed of two subunits: HIF-1a, which has a
short half-life (*5 min) and is highly sensitive to oxygen,
and HIF-1b(or ARNT: aryl hydrocarbon nuclear receptor),
which is constitutively expressed and remarkably insensi-
tive to oxygen levels. Although the HIF family comprises
two other members, HIF-2 and HIF-3, only HIF-1 is known
to play a very general role in signaling hypoxia, whereas
induction of HIF-2 with hypoxia is restricted to certain
cells and HIF-3 function is still incompletely understood.
During normoxia, HIF-1ais degraded through hydroxy-
lation. This process involves prolyl hydroxylases acting on
HIF-1aoxygen-dependent degradation domain (ODDD).
Once it has been hydroxylated, HIF-1abinds to von Hippel
Lindau (VHL) protein, resulting in proteasomal degradation
of HIF-1a.
Under hypoxic conditions, HIF-1adegradation is
blocked because hydroxylation is inhibited. HIF-1aprotein
therefore accumulates, allowing for its binding to ARNT
(HIF1-b) and hence the formation of a HIF-1 complex that
can recognize hypoxia responsive elements (HRE) located
in the nucleus of target genes (see also Table 1). The
interaction between HIF-1 and HRE ultimately triggers the
transcription of the target genes. To date, more than 100
HIF-1 downstream genes have been identified [5]. Those
genes, which mainly enable the cells to cope with oxygen
stress, are involved in erythropoiesis/iron metabolism,
angiogenesis, glucose metabolism, and cell proliferation/
survival and apoptosis, respectively. While this machinery
helps normal cells to ameliorate local oxygen availability,
it also contributes to the survival/development of tumor
cells, thereby highlighting the dual role played by HIF-1.
The HIF-1 pathway can also be modulated under non-
hypoxic conditions, either at the transcriptional or at the
translational level [6,7]. A variety of non-hypoxic envi-
ronmental factors, including inflammation, reactive oxygen
species, or nitric oxide can induce HIF-1aaccumulation
and target gene expression [5], which may further com-
plicate our understanding of the cellular response to
hypoxia. However, it is worth noting that the contribution
of these regulators is only partially understood and that
their influence on HIF-1 stabilization is far less than the
dramatic induction caused by hypoxia.
HIF-1 in skeletal muscle tissue
Like in virtually all cells, skeletal muscle tissue is able to
respond to hypoxia through the HIF-1 pathway. Animal
studies have demonstrated that 1 h of systemic hypoxia is
sufficient to increase HIF-1aprotein expression in skeletal
muscle [8]. In accordance with this, when leg oxygen
delivery is impaired following blood flow restriction in
humans, HIF-1aprotein levels are known to be increased
[9]. One particular aspect of skeletal muscle is that HIF-1a
protein is highly expressed in this tissue even in normoxic
conditions, suggesting that HIF-1 could have potential
function in muscle homeostasis in normoxia [9,10].
Of note is the fact that the HIF-1 response to hypoxia is
time dependent. Previous data obtained in brain, liver, and
kidney show that HIF-1 protein levels peak within the first
hours of hypoxic exposure then progressively decline
toward basal levels [8,11]. The reason for HIF-1 to be
down-modulated with ‘‘acclimatization’’ could be that
local or systemic responses to hypoxia may attenuate the
degree of cellular hypoxia in a tissue-dependent manner.
That a similar response may occur in skeletal muscle tissue
during sustained hypoxia could be hypothesized, but has
not so far been verified. Our ongoing work, investigating
the alterations of human skeletal muscle HIF-1aacross
time in hypoxia, provides the first indication that HIF-1a
protein levels are barely modified in human skeletal muscle
during environmental hypoxia (4,559 m of altitude), either
acutely (0.5–8 h) or chronically (7–9 days) [12]. This
preliminary finding questions the actual role of HIF-1ain
vivo on the control of muscle adaptations during sustained
hypoxia. In the same subjects, we concurrently observed
permanently high levels of HIF-1amRNAs at high altitude
[13], suggesting that, in skeletal muscle, (1) factors other
than hypoxia may be involved in the increase in HIF-1a
mRNA, and (2) HIF-1aprotein expression is probably not
regulated at the transcriptional level. In summary, although
HIF-1 has long been recognized as the master regulator of
the cellular responses to hypoxia in numerous cell types,
the few available data obtained from skeletal muscle tissue
in vivo suggest that HIF-1 protein expression is only
marginally altered during hypoxia. One reason could be the
high pre-existing level of HIF-1 expression in skeletal
muscle (aimed at maintaining homeostasis in a tissue
operating at low physiological oxygen tension) that would
3616 C. Lundby et al.
leave only little room for additional changes when oxy-
genation is decreased.
Physical exercise per se may also challenge muscle
oxygen homeostasis [3]. Accordingly, an alteration in
HIF-1 signaling is expected. In support of this is the recent
finding of increased HIF-1aprotein expression in human
skeletal muscle following an acute bout of normoxic
exercise [9]. However, the exact pathway by which HIF-1
acts on muscle tissue in response to exercise-induced
hypoxia and ultimately on exercise tolerance remains a
Table 1 Gene expression and
protein activation in skeletal
muscle in response to hypoxia
Lundby u.o. indicates a personal
unpublished observation in
muscle biopsies obtained from
human subjects after up to 8 h
exposure to a barometric
pressure equivalent to 4,559 m
altitude. This is only used when
no other data are available
Gene HIF-1
target
Main function Response to hypoxia
mRNA Protein
VEGF Yes Angiogenesis :[50]
:[9]
?
Leptin Yes Angiogenesis ? ?
Transforming growth
factor-b3
Yes Angiogenesis ? ?
Nitric oxide synthase Yes Vascular tone ? ?
Heme oxygenase 1 Yes Vascular tone ?(Lundby u.o.) ?
Adrenomedullin Yes Vascular tone :[50]?
a
1B
-Adrenergic receptor Yes Vascular tone ? ?
Adenylate kinase-3 Yes Glucose metabolism ? ?
Aldolase-A,C Yes Glucose metabolism ?(Lundby u.o.) ?
Carbonic anhydrase-9 Yes Glucose metabolism ? IV ?[30]
XIV ;[30]
Enolase-1 Yes Glucose metabolism ? ?
Glucose transporter 1-3 Yes Glucose metabolism ? ?
Glyceraldehyde phosphate
dehydrogenase
Yes Glucose metabolism :[51]?
Hexokinase Yes Glucose metabolism ?(Lundby u.o.) ?[52]
:?[26]
:[53]
Lactate dehydrogenase A Yes Glucose metabolism :[50]?[54]
?[30]
?[52]
?[26]
Pyruvate dehydrogenase M Yes Glucose metabolism ? ?
Phosphofructokinase L Yes Glucose metabolism ?[50];[55]
Phosphoglycerate kinase-1 Yes Glucose metabolism :[50]?[26]
Glucose transporter 4 ? Glucose metabolism :(Lundby u.o.) :[56]
Transferrin receptor Yes Iron metabolism ;[13];[13]
Ferritin ? Iron metabolism ? ;[13]
Ferroportin ? Iron metabolism :[13]?
Insulin-like growth factor-2 Yes Cell proliferation ?(Lundby u.o.) ?
Monocarboxylate
transporter 1-4
? Lactate transport ? ?(;)[54]
?[30]
Citrate synthase ? Oxidative metabolism ? ?[31,55]
?[24]
;[26]
?[53]
3 hydroxyacyl CoA
dehydrogenase
? Oxidative metabolism ? ?[24]
?[53]
Myoglobin ? Oxygen storage and diffusion ?(Robach u.o.) ;[13]
Myosin heavy chains ? Muscle contraction ? ?[23]
?[55]
Skeletal muscle and hypoxia 3617
complex issue. Recent studies using mice lacking skeletal
muscle HIF-1areveal that, surprisingly, endurance capac-
ity is increased in these animals through an increase in
oxidative metabolism [14]. However, these HIF-null mice
are otherwise subject to increased muscle damage because
of an impeded glycolytic metabolism, highlighting that
HIF-1 is nevertheless essential in the metabolic control of
muscle function.
If a single bout of exercise is associated with muscle
hypoxia, it can be suggested that the repetition of exercise
potentially challenges muscle oxygen homeostasis even
more. The effect of chronic exercise on skeletal muscle
function has been extensively investigated. Angiogenesis
and alterations of metabolic control are well-known adap-
tations to endurance training [15]. It is thought that HIF-1
plays a central role in this setting, since HIF-1 modulates
vascular endothelial growth factor, as well as several gly-
colytic enzymes. However, recent evidence shows that
endurance training actually reduces muscle HIF-1asig-
naling [16]. Furthermore, HIF-1 null and wild-type muscles
are shown to respond similarly to endurance training
[17,18], suggesting that the HIF-1 pathway is not essential
for endurance training, while the latter could be useful
during acute exercise.
Finally, how does skeletal muscle respond to hypoxia at
the molecular level if exercise is superimposed and hence
combines the effects of exercise and hypoxia on HIF-1
signaling. Although relevant because extremely challeng-
ing for local oxygen homeostasis, this issue is not well
documented, making it difficult to understand the interac-
tion between HIF-1 and its target genes during this
condition. As mentioned above, combining acute hypoxia
and acute exercise results in higher HIF-1 protein expres-
sion than with exercise alone [9]. If hypoxia is associated
with endurance training, HIF-1amRNA levels are found to
be higher than after endurance training alone [19,20].
Taken together, these data indicate that HIF-1 can be
activated in skeletal muscle during exercise in hypoxia,
highlighting its possible functional role on target genes
involved in angiogenesis and/or energy metabolism. The
early (acute) response of hypoxia on gene expression in
human skeletal muscle is at present poorly described, and
for this reason and the limited space available, this step will
limited to that shown in Table 1.
Morphological and enzymatical adaptations in human
skeletal muscle exposed to prolonged hypoxia
The first report describing the potential for human skeletal
muscle to adapt to hypoxic exposure was published by
Reynafarje in 1962 [21]. He showed that in miners per-
manently exposed to high altitude as compared to
lowlanders cytocrome creductase activity was increased
by 78% and myoglobin content by 16%. Subsequently, it
was for a long time believed that these adaptations had
occurred to compensate for the lack of oxygen. Later, it has
been argued that hypoxic stimulus is not sufficient in order
to induce these responses alone, and that hypoxia has to be
combined with either cold or exercise [22]. More recent
data from our research group showed that capillary density
was not increased in sea level residents exposed to 4,100 m
in La Paz for 8 weeks [23], and also that 75 days exposure
to 5,300 m in the base camp of Mount Everest did not
cause the capillary number per muscle fiber to be altered
(i.e., no neo-formation) in samples obtained from arm and
leg skeletal muscle tissue [24]. In the later study, however,
a decrease in fiber size resulted in more capillaries per
area—a phenomenon often observed at this altitude or
above [25,26]. It has been speculated that the decrease in
fiber size at altitude is due to a hypoxia-induced down-
regulation of protein synthesis, because COPD patients
have a reduced protein synthesis, and this has been
hypothesized to be a direct effect of hypoxia (reviewed in
[27]). While the muscle atrophy could be a consequence of
hypoxic exposure per se, high altitude expeditions are also
frequented by gastroenteritis, malnutrition, low physical
activity levels, and low temperatures, and may very well be
the reason for muscle atrophy [28].
Selected marker enzymes for oxidative metabolism (CS
and HAD activity) were unchanged after 75 days at
5,300 m in the leg and arm muscles of active climbers and
inactive base camp personnel [24]. Using the proteomic
approach, it has recently been demonstrated that high
altitude Sherpas have a slightly reduced HAD and lactate
dehydrogenase protein content. Also, it seemed that, at
least to some extent, high altitude residing Sherpas are
protected from ROS-induced tissue damage and possess
specific metabolic adaptations [29]. The protein density of
lactate transport proteins MCT 1 and 4 do not seem to
change in skeletal muscle in humans exposed to altitude,
whereas proteins involved in acid–base regulation are
increased [30]. Also, buffer capacity has been shown to
increase in both arm and leg extremities following altitude
exposure [24]. After a climbing expedition to Mt. Denali, a
13.8% down-regulation in muscle Na
?
/K
?
-ATPase has
been reported [31]. In contrast to the climbing expedition
to Mt. Denali, eight sea level natives exposed to 4,100 m
altitude in the outskirts of La Paz (living in a modern
apartment) did not experience any change in the three
Na
?
/K
?
pump subunits in muscle biopsies obtained after 2
and 8 weeks of high altitude exposure [30]. A decrease in
Na
?
/K
?
-ATPase has been hypothesized to allow a given
amount of work to be performed at lower ATP costs, and
would thus seem a favorable adaptive response to high
altitude exposure [32]. Since O
2
consumption for a given
3618 C. Lundby et al.
workload seems unchanged following altitude exposure
[33], this also argues against an altitude-induced down-
regulation of skeletal muscle Na
?
/K
?
-ATPase. In sum-
mary, the current knowledge indicates that prolonged
hypoxic exposure would not induce skeletal muscle angi-
ogenesis, and also that oxidative enzymes in human
skeletal muscle respond only marginally to long-term
altitude exposure. This may come as a surprise, since one
would think that both responses (angiogenesis and oxida-
tive enzymes) should be increased knowing the HIF-1
pathway.
Human skeletal muscle gene response to exercise
in hypoxia
As stated in the ‘‘Introduction’’, exercise may superimpose
an already present hypoxic stimulus, and the gene response
to hypoxic exercise has been investigated in at least two
studies (and more if studies using ischemia are also
regarded). In both studies, human subjects underwent a
muscle biopsy before and after a 6-week training period,
either involving two [20] or five [19] training sessions per
week. The biopsies were obtained 24 h [19]or48h[20]
following the last training session, and therefore it can
unfortunately not be assessed whether the reported gene
response is associated to the total 6-week stimulus or to the
last hypoxic training session. Following exercise training,
most genes peak their expression 2–8 h into recovery [34].
Since the biopsies in the above mentioned studies were
obtained 24 and 48 h following the last exercise bout, (1)
the potential increase in genes augmented as a consequence
of the last exercise bout may not be represented in biopsies
obtained at these time points, or (2) the augmented genes
may be the result of the total training regimen. In order to
be able to distinguish between the acute and chronic
response, an additional biopsy should have been obtained
following the very first training session. Regardless of
origin, however, it may be assumed that the augmented
mRNA levels may also induce increased protein contents.
Although the degree of hypoxia, and also the training
intensity was quite similar in both studies, the mRNA
response was not similar. While mRNA levels of HIF-1
and myoglobin were augmented in both studies, VEGF was
increased in [19] but not in [20]. On the other hand, the
mRNA content of COX-1 and PFK did not increase more
than in the normoxic control group in [19], whereas this
was the case in [20]. In addition, Glut-4, PGC-1a, CS,
MCT-1, and a few other genes were augmented in [20], but
not investigated in [19]. To draw clear-cut conclusions
regarding the effect of hypoxic exercise on gene expres-
sion, future studies could be conducted—if possible with
subjects performing one-legged kicking in normoxia and
hypoxia, with biopsies obtained in the early recovery per-
iod following the single exercise bout.
Morphological and enzymatical adaptations following
hypoxic exercise training
The rationale of hypoxic exercise training relies on the
hypothesis that such regimen may induce muscle adapta-
tions that are beyond the responses triggered by chronic
exercise alone. Different training models of hypoxic
exercise may be used: living at sea level and training at
altitude (as mentioned above) or training and living at
altitude. Studies employing either model are discussed
below.
In the above-mentioned studies (living at sea level and
training in hypoxia), Vogt et al. [19] reported that total
mitochondrial density was increased more in the hypoxic
training groups as compared to those performing the
training in normoxia. In addition, capillary length density
was also increased as a consequence of the training per-
formed in hypoxia (i.e., both training responses are
different from those reported to occur with chronic altitude
exposure). In agreement herewith, Ponsot et al. [35]
reported that hypoxic training did not alter mitochondrial
function but led to a better coupling between the energy
utilization and production sites. They also reported that
hypoxic training did not alter skeletal muscle fiber com-
position or the content of selected oxidative enzymes
(as with chronic altitude exposure). Using a similar
approach, Masuda and coworkers [36] investigated the
effects of 8 weeks training in hypoxia (2,500 m) as com-
pared to normoxia. Following training, there was no
difference between groups in myoglobin, muscle fiber
composition, capillarity, or citrate synthase activity. In yet
another study, eight subjects were assigned to altitude
(2,300 m) or sea level training (all living at sea level).
Compared to the normoxic training group, the moderate
altitude group experienced a decrease in muscle PFK
activity but an increase in muscle capillary density [37].
Interestingly, the same research group conducted a study
using one-legged training [38]. This model has the
advantage that two situations with similar magnitudes of
mitochondrial substrate flux but different blood oxygen
contents can be compared. Ten subjects trained one leg
under normoxic conditions and the other under hypoxic
conditions. There was a greater increase of citrate synthase
activity under hypoxic conditions than under normoxic
conditions. In addition, the myoglobin content increased in
the leg trained under hypobaric conditions, whereas it
tended to decrease in the normoxia-trained leg. Capillary
density did not respond to the addition of hypoxia. Using a
somewhat similar model, Melissa and coworkers studied
Skeletal muscle and hypoxia 3619
ten males before and after 8 weeks of unilateral cycle
ergometry training so that one leg was trained while
breathing an inspirate of 13.5% O2 and the other while
breathing normal ambient air. Biopsies from quadriceps
revealed an increase in CS, whereas succinate dehydroge-
nase and PFK activity, capillary density, fiber area, % fiber
type, and mitochondrial and lipid volume density all
remained unaltered between groups [39]. In line with this,
six Scandinavian runners were taken to either Portugal (sea
level) or Kenya (2,000 m) for a 14-day-long training camp.
No differences were found between groups with regard to
muscle fiber size, composition, or capillarization. Also, CS
and HAD activity did not differ between the groups. In
contrast to Scandinavian runners, local Kenyan runners
(predominantly living and training at 2,000 m) were
reported to have higher HAD activity levels, but the other
characteristics were similar to those found in the Scandi-
navians [40]. Using an interesting model, Desplanches and
coworkers [41] investigated the effects of supplementing
high altitude natives with oxygen during training at alti-
tude. The rationale for this was the belief that reduced
muscle stress during endurance training in hypoxia could
limit muscle adaptations. Compared to the non-oxygen-
supplemented training group, the 6-week training program
(5 weeks, 30 min/session at *70% of max) with oxygen
supplementation did not lead to differences in capillary-to-
fiber ratio between groups, capillary density, volume den-
sity of total mitochondria, or CS-, PKF-, and HAD-activity.
In summary, it would seem that hypoxic training may
increases CS more than training in normoxic conditions.
Although such an adaptation theoretically improves muscle
function and therefore exercise tolerance, the physiological
significance of this enzymatic adaptation deserves further
investigation. In addition, it appears that the response
hereof likely depends on the degree of hypoxia and training
duration. In contrast, it seems that structural changes—in
addition to those already occurring with normoxic train-
ing—are less likely to occur when hypoxia is superimposed
on chronic exercise.
New insights: interactions between iron metabolism,
myoglobin and muscle function at high altitude
Iron plays a central role in a large number of essential
cellular functions. Its pivotal role in oxygen transport has
in recent years generated a considerable body of scientific
work that is greatly improving our understanding of the
interactions between tissue hypoxia and iron metabolism.
However, reviewing the response of skeletal muscle tissue
to hypoxia shows that the role of iron remains globally
unknown while other cellular/molecular aspects have been
documented.
The central role for HIF-1 signaling in oxygen homeo-
stasis by regulating the glycoprotein hormone erythro-
poieitin (Epo) is well established. Erythropoiesis is a
complex process that concurrently induces dramatic changes
in iron metabolism (also influenced by HIF-1) in order to
fulfill the high demand for iron within the bone marrow to
synthesize hemoglobin. In response to prolonged hypoxia,
the up-regulation of red blood cell production is associated
with progressive systemic iron deficiency [42]. We recently
investigated whether this high need for iron during enhanced
erythropoiesis at high altitude could interact on skeletal
muscle iron stores and ultimately on muscle oxygen homo-
eostasis in humans [13]. Our results indeed demonstrated
that prolonged exposure to hypoxia induces a down-modu-
lation in several iron proteins in skeletal muscle, and hence
indicating muscle iron loss. The consequence of such muscle
iron loss is a decrease in myoglobin protein expression at
high altitude, suggesting an altered muscle oxygen homeo-
stasis. Our results do not support previous evidence showing
that prolonged hypoxia may enhance the synthesis of myo-
globin [21,38,43]. Beyond the differences between the
experimental approaches that may account for the divergent
results, an important clue to our understanding of myoglobin
biology is still lacking, namely the evidence that the
transcriptional regulation of myoglobin is mediated by a
HIF-1-dependent mechanism [44].
A question that remained unanswered is the physiolog-
ical importance of myoglobin for exercise tolerance at high
altitude. Although myoglobin is basically known as an
oxygen-storage protein and a diffusion facilitator, its
physiological role has been challenged by studies showing
that myoglobin knockout mice are able to exercise nor-
mally, suggesting that muscle function can be preserved
even in the absence of myoglobin [45]. Of note, however,
is the fact that the myoglobin-null mice demonstrate a
number of molecular adaptations/compensations that may
explain their unexpected normal exercise capacity. Insights
into the functional role of myoglobin may also be gained
from previous physiological studies on altitude acclimati-
zation indicating that exercise capacity at high altitude
(which is permanently decreased in comparison to sea
level) is immediately restored to sea level values when
acute reoxygenation is applied to acclimatized lowlanders
[46–48]. Such an observation leads to the speculation that
myoglobin would not be a major determinant of exercise
capacity in normoxia, otherwise any decrease in myoglobin
content following altitude acclimatization [13] would be
expected to impair the recovery of exercise capacity upon
acute reoxygenation. Hence, the process of O
2
diffusion
from the capillaries to muscle mitochondria would be, to
some extent, independent of myoglobin during normoxic
conditions. During chronic hypoxia, leg O
2
conductance
(which is a measure of oxygen diffusing capacity in muscle
3620 C. Lundby et al.
tissue) is found to be reduced [47], suggesting that O
2
conductance is a limiting factor for exercise tolerance at
high altitude. Although such observation raises the possi-
bility of a significant role played by myoglobin in
maintaining muscle O
2
homeostasis when O
2
supply is
restricted, to date, no evidence supports nor opposes this
contention.
In summary, iron can be considered as an emerging
issue for skeletal muscle function in hypoxia. However,
how does high altitude exposure exactly alter muscle iron
metabolism remains a complex issue since two stimuli, i.e.,
hypoxia and accelerated erythropoiesis, coexist during high
altitude studies. In order to gain further insights into the
interactions between oxygen, erythropoiesis, and iron, we
recently investigated the effect of enhanced erythropoiesis
on muscle iron metabolism under non-hypoxic conditions,
by injecting healthy humans with recombinant erythro-
poietin. Surprisingly, we found that, under normoxic
erythropoietin stimulation (1 month), muscle was not a
source of iron for erythropoiesis. On the contrary, the
changes in the expression of muscle iron proteins were
indicative of skeletal muscle iron accumulation [49]. Such
response differs from muscle iron loss occurring at high
altitude [13] and highlights the potential role of hypoxia in
triggering muscle iron mobilization. Accordingly, it can be
speculated that skeletal muscle tissue would serve as an
iron storage compartment only under massive stress, such
as prolonged hypoxia.
Conclusion
The current published data suggest that skeletal muscle
tissue is not remarkably altered with chronic exposure to
hypoxia in resting humans. This contention is supported by
two major aspects, which are first the absence of muscle
angiogenesis, and second the marginal response of oxi-
dative enzymes to long-term altitude exposure. Our
understanding of the molecular events mediating muscle
morphological and enzymatic responses to hypoxia is still
fragmented. Nevertheless, emerging evidence suggests that
HIF-1 signaling in skeletal muscle tissue displays a rather
modest response to hypoxia, and thereby explains why
muscle function is only barely modified in hypoxia. In
contrast, the few data obtained in humans exercising in
hypoxia raise the possibility that the combination of these
two stimuli might result in structural and functional adap-
tations in skeletal muscle.
In conclusion, skeletal muscle tissue appears to be rather
well adapted to the hypoxic environment, at least in healthy
humans exposed to terrestrial hypoxia. That exercise
training associated with hypoxia may improve muscle
oxygen homoeostasis remains a relevant issue that deserves
future work.
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