0021-972X/99/$03.00/0
The Journal of Clinical Endocrinology & Metabolism
Copyright © 1999 by The Endocrine Society
Vol. 84, No. 3
Printed in U.S.A.
Tissue Composition Affects Measures of Postabsorptive
Human Skeletal Muscle Metabolism: Comparison
across Genders*
LINDA A. JAHN, EUGENE J. BARRETT, MICHAEL L. GENCO, LIPING WEI,
THOMAS A. SPRAGGINS, AND DAVID A. FRYBURG
Department of Internal Medicine and Diagnostic Radiology and General Clinical Research Center,
University of Virginia Health Sciences Center, Charlottesville, Virginia 22908
ABSTRACT
Despite clear anthropomorphic differences, gender differences in
human skeletal muscle protein and carbohydrate metabolism have
not been carefully examined. We compared postabsorptive forearm
glucose, oxygen, and lactate balances and forearm protein kinetics
between 40 male and 36 female subjects. Forearm composition was
measured in a subset of 17 subjects (8 males and 9 females) using
multislice magnetic resonance imaging. Oxygen uptake, net phenylalanine release, and estimated rates of forearm protein synthesis and
degradation were greater in male than in female subjects when expressed as the rate per 100 mL forearm volume (P , 0.05). In males,
however, muscle accounted for 58% of forearm volume, compared with
S
INCE THE early work by Andres and Zierler (1, 2), the
human forearm has been used extensively to study
human skeletal muscle metabolism. Despite frequent use,
there has been no careful evaluation of gender differences
in the handling of substrates within the human forearm.
The widespread use of anabolic steroids to increase muscle
mass and enhance performance (3) suggests that androgens may particularly alter muscle protein metabolism (4).
If this is true, then we might expect to observe a gender
difference in the metabolism of protein and perhaps other
fuel substrates by the human forearm. To address the issue
of gender differences, we compared forearm muscle protein, glucose, lactate, and oxygen metabolism in healthy
young men and women after an overnight fast. We used
the classical arterial-venous difference measurements together with plethysmographic measurements of forearm
flow. As there is a gender difference in forearm tissue
composition, with men having a greater percentage of
muscle mass per total forearm volume (5, 6), in a subset of
men and women we performed tissue composition studies
of the forearm using magnetic resonance imaging. These
measurements were used to estimate the contribution that
compositional differences might have on estimates of forearm substrate balance.
Received July 23, 1998. Revision received November 24, 1998. Accepted December 4, 1998.
Address all correspondence and requests for reprints to: Linda A.
Jahn, Department of Internal Medicine, MR-4 Box 5116, University of
Virginia Health Sciences Center, Charlottesville, Virginia 22908.
* This work was supported by NIH Grants RO1-DK-38578, RO1-DK54058, and RR-0847
46% in females (P , 0.001). When phenylalanine balance, protein
degradation and synthesis, and glucose and oxygen uptake were expressed per 100 mL forearm muscle, there were no significant differences across gender. Likewise, the extraction fractions for oxygen,
glucose, phenylalanine, and labeled phenylalanine were comparable
in males and females. We conclude that cross-gender comparisons of
metabolic variables must accommodate differences in tissue composition. These data indicate that in the postabsorptive state, skeletal
muscle metabolism of glucose, protein, and oxygen do not differ by
gender in healthy young humans. (J Clin Endocrinol Metab 84: 1007–
1010, 1999)
Subjects and Methods
Subjects
Seventy-six healthy (36 females and 40 males), normal weight (body
mass index, 22.04 6 0.4 in females and 23 6 0.3 in males), young adult
(23 6 1 yr), volunteers were admitted to the University of Virginia
General Clinical Research Center the evening before the study. No
subject was taking any medication, and all female participants had a
negative serum pregnancy test 1–2 days before the study. The study
protocol was approved by the University of Virginia human investigation committee, and each subject gave written consent.
Experimental protocol
After an overnight 12-h fast, a brachial artery and an ipsilateral,
retrograde, median cubital (deep) vein catheter were placed percutaneously. Each subject received a primed (;33 mCi), continuous (0.43
mCi/min) infusion of L-(ring 2, 6)-3H phenylalanine through a catheter
placed in the lower extremity. After a 90-min tracer equilibration period,
quadruplicate, paired arterial and venous samples were taken over 30
min for measurement of phenylalanine concentration and specific activity and of glucose, lactate, and oxygen concentrations. Forearm blood
flow was measured after each set of arterial and venous samples by
capacitance plethysmography.
In a subset of 17 subjects (8 males and 9 females), magnetic resonance
imaging (MRI) of the forearm was performed using a 1.5 T Magneton
63SP (Siemens, Erlangen, Germany). The image time was 6.52 min, and
the resolution time was 1.2 mm in plane with a 4.7-mm effective slice
thickness. The sequence was repeated with an overlapping slice to cover
the full distance between the olecranon and the styloid processes. Images
were evaluated using Sigma Scan (version 1.20, Jandel Scientific, Chicago, IL). A total of 16 slices/subject were used to reconstruct a 3-dimensional image of the forearm. Tissue volumes between slices were
estimated as a truncated cone and then summed.
Analytic methods
Whole blood glucose and lactate concentrations were measured by a
combined glucose/lactate analyzer (Yellow Springs Instruments, Yel-
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low Springs, OH). Blood oxygen content was measured spectrophotometrically using an OSM2 hemoximeter (Radiometer, Copenhagen,
Denmark). Phenylalanine concentration and specific activity were measured as previously described (7).
Calculations of forearm phenylalanine kinetics. The net forearm balances for
glucose, lactate, oxygen, and phenylalanine were calculated using the
Fick principle: net forearm balance 5 ([A] 2 [V]) 3 F (Eq I), where [A]
and [V] are arterial and venous substrate concentrations, and F is forearm blood flow in milliliters per min/100 mL forearm volume. The rates
of protein synthesis and degradation were estimated from the kinetics
of exchange of labeled phenylalanine across the forearm using the specific activity of phenylalanine in venous plasma to reflect the precursor
pool used for protein synthesis as previously described (8): synthesis 5
([dpmart 2 dpmvein] 3 flow)/SAvein (Eq II), and muscle protein breakdown (B) as: breakdown 5 S 2 net balance (Eq III).
As the deep forearm venous catheter used in our forearm balance
study drains nearly exclusively muscle (1) we used two approaches to
calculate muscle balance and protein kinetics using Eq I–III. In the first
approach, in each of the subjects whose forearm composition was measured we simply divided the balance or flux by the fractional contribution of muscle to forearm volume. This result, expressed as mass per
100 mL forearm muscle, assumes that blood flow to muscle, skin, bone,
and adipose distributes in proportion to the volume that they each
contribute to the forearm. The second approach to correcting forearm
balances and fluxes relies on the results of recent positron emission
tomographic studies (9), indicating that human leg total blood flow can
be related to muscle blood flow by the equation: blood flow/100 mL leg
muscle 5 [1.41 3 total leg blood flow (mL/100 mL limb volume)] 2 0.43.
This estimate of flow distribution, together with the areterio-venous
difference (across muscle) and the estimated fraction of the limb occupied by muscle, can be used to calculate the total substrate exchange by
muscle.
For the entire study group, the extraction ratios ({[X]artery 2 [X]vein }/
[X]artery) for glucose, oxygen, lactate, phenylalanine, and [3H]phenylalanine were also calculated to assess the handling of these substrates
by muscle without blood flow as a multiplier.
Data presentation and statistical analysis. All data are presented as the
mean 6 sem. Comparisons between males and females, before as well
as after correcting for muscle mass, were made using Student’s t test.
Results
Forearm phenylalanine kinetics; glucose, lactate, and
oxygen balances; and blood flows
Forearm phenylalanine balance was negative in these
postabsorptive subjects. Moreover, net phenylalanine release
was significantly greater in males than in females (222 6 2
vs. 217 6 1; P , 0.05). In addition, in males, the rates of both
protein degradation (Eq III; 71 6 6 vs. 54 6 3; P , 0.005) and
protein synthesis (49 6 4 vs. 37 6 3; P , 0.005) were greater
than those in females, suggesting a higher rate of muscle
protein turnover and a greater net catabolism in men than in
women (Table 1). There were no significant gender differences in glucose uptake, lactate release, or blood flow per 100
mL forearm (Table 1). Oxygen uptake was greater in male
than in female subjects (Table 1).
The subjects in whom MRI measurements were obtained
were of a similar age and weight to the larger study group,
and the balances for phenylalanine, glucose, lactate, oxygen,
and the kinetics for tracer phenylalanine exchange (expressed per 100 mL forearm volume) were similar to those
seen in the entire study group (Table 1). The gender differences in phenylalanine and oxygen balance seen in the subgroup studied using MRI were similar to those seen in the
entire study population; although they were not statistically
significant in this subgroup, presumably due to the sample
size. All these data were expressed per 100 mL forearm
volume, as has been done in virtually all studies of forearm
metabolism (1, 2, 14, 18).
Forearm tissue composition
Table 2 indicates the forearm composition determined by
the reconstruction of the 16 cross-sectional images in each of
the 17 subjects. The relative contribution of adipose tissue to
total forearm volume was greater (42% vs. 30%) and the
contribution of muscle was less (46% vs. 58%) in the female
subjects (P , 0.001 for each). Interestingly, the fractional
content of bone was similar in both sexes (12% vs. 12%).
Moreover, the variances in the total volumes and percent
composition within each sex were narrow.
Estimated forearm muscle glucose, lactate, and oxygen
balances and phenylalanine kinetics
Given the narrow variance of forearm percent composition
within genders, we used the forearm tissue composition
given in Table 2 to recalculate the balances for phenylalanine,
glucose, lactate, and oxygen (calculated as (arterio 2 venous)
3 F) for the entire study population, now normalized per 100
mL forearm muscle (Table 3). Calculating balances or fluxes
in this manner is equivalent to assuming that all blood flow
to the forearm is directed to tissue in proportion to its contribution to forearm volume. In an effort to more precisely
estimate muscle’s contribution to forearm metabolism, fluxes
were estimated assuming that blood flow in the forearm of
the present study’s subjects is partitioned between tissues as
described by recent positron emission tomographic mea-
TABLE 1. Forearm muscle metabolism in postabsorptive men and women
Entire cohort
Phenylalanine balance
Breakdown
Synthesis
Glucose balance
Lactate balance
Oxygen balance
Flow
MRI cohort
Men (n 5 40)
Women (n 5 36)
Men (n 5 8)
Women (n 5 9)
222 6 2
71 6 6
49 6 4
0.73 6 0.1
20.14 6 0.1
11 6 1
3.5 6 0.2
217 6 1a
54 6 3b
37 6 3b
0.65 6 0.1
20.25 6 0.1
9 6 1a
3.5 6 0.2
222 6 3
73 6 17
51 6 16
0.61 6 0.1
20.00 6 0.1
11 6 2
4.0 6 1
217 6 2
54 6 7
39 6 6
0.66 6 0.2
20.35 6 0.1a
961
3.3 6 0.3
Data are expressed as the mean 6 SEM. Phenylalanine balance, synthesis, and degradation in nanomoles per min/100 mL of forearm volume.
Glucose, lactate, and oxygen balances in micromoles per min/100 mL forearm volume. Blood flow in milliliters per min/100 mL forearm volume.
a
P , 0.05.
b
P , 0.005.
TISSUE COMPOSITION AND SKELETAL MUSCLE METABOLISM
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TABLE 2. Gender comparison of forearm volume as measured by MRI
Men (n 5 8)
Total
Muscle
Bone
Fat 1 skin
1044 6 50
607 6 36
125 6 5
312 6 16
Data are expressed as the mean 6
of absolute volumes between sexes.
SEM
% of total
Women (n 5 9)
58 6 1.2
12 6 0.5
30 6 1.1
703 6 29
323 6 16
85 6 7
295 6 24
% of total
P
46 6 2.6
12 6 0.6
42 6 2.5
0.00001
0.0000008
0.0002
0.55
in milliliters and as a percentage of the total volume for each tissue. P values refer to comparison
TABLE 3. Forearm muscle metabolism in postabsorptive men and women normalized per 100 mL/forearm muscle
Vol corrected
Phenylalanine balance
Breakdown
Synthesis
Glucose balance
Lactate balance
Oxygen balance
Vol and flow corrected
Men (n 5 40)
Women (n 5 36)
P
Men (n 5 40)
Women (n 5 36)
P
237 6 3
125 6 9
87 6 7
1.3 6 0.1
20.24 6 0.1
19 6 1
237 6 3
117 6 7
79 6 6
1.4 6 0.1
20.6 6 0.1
19 6 1
0.98
0.48
0.35
0.49
0.06
1.00
248 6 4
160 6 12
112 6 9
1.6 6 0.2
20.31 6 0.1
24 6 2
248 6 3
150 6 9
102 6 8
1.8 6 0.2
20.7 6 0.2
24 6 1
0.97
0.49
0.37
0.52
0.06
0.98
Data are expressed as the mean 6 SEM. Phenylalanine balance, synthesis, and degradation in nanomoles per min/100 mL forearm muscle.
Glucose, lactate, and oxygen balances in micromoles per min/100 mL forearm muscle. P was determined using Student’s t test.
surements (see Materials and Methods) (9). These results are
also shown in Table 3. Regardless of which approach was
used, the balances for each of the metabolites were closely
matched between genders (Table 3).
Extraction fractions for phenylalanine, phenylalanine
disintegrations per min, glucose, lactate, and oxygen
The forearm extraction fraction for each substrate was
calculated as previously described. These results are given in
Table 4. Again, there was no significant difference noted
between genders, with the exception of a greater fractional
lactate release in females.
Discussion
In comparing males and females, there was an apparent
gender difference in rates of forearm protein synthesis, degradation, and net balance and metabolic rate (O2 consumption) when fluxes were expressed in the conventional manner as the rate per 100 mL forearm volume. Previous studies
had demonstrated that subcutaneous adipose makes a
greater contribution to total extremity volume in both the
arm and the leg of healthy young females compared to males
(5, 6). This led us to estimate the metabolic rates per 100 mL
forearm muscle as well. We used two methods. First, we
simply divided the total forearm balance measurements by
the fraction of forearm that is muscle. Second, we estimated
the blood flow distribution to muscle using a mathematical
relationship recently derived from positron emission tomography imaging studies (9) of the human leg where total limb
flow and muscle flow were both measured. Using either
approach, the rates of muscle metabolism of glucose, oxygen,
and protein were not significantly different between genders.
This was confirmed by comparison of the extraction ratios for
oxygen, glucose, phenylalanine, and labeled phenylalanine,
a measurement that is not dependent upon blood flow as a
multiplier. Previously, the fractional rate of protein synthesis
in leg muscle was determined using stable isotopic methods
combined with muscle biopsy. No gender difference was
TABLE 4. Extraction fractions of substrates across forearm
muscle
Phenylalanine
Phenylalanine (dpm)
Glucose
Lactate
Oxygen
Men (n 5 40)
Women (n 5 36)
P
212 6 0.6
21 6 1.0
5 6 0.5
27 6 3.0
38 6 1.0
212 6 0.7
18 6 1.0
4 6 0.4
218 6 3.0
37 6 2.0
0.59
0.08
0.61
0.01
0.73
Data are expressed as the mean 6 SEM, expressed as a percentage.
P was determined using Student’s t test.
noted in fractional synthesis rate for either myosin heavy
chain, sarcoplasmic proteins, or mixed muscle protein in that
study (10). Of these variables, the latter would be expected
to correspond to the rate of protein synthesis measured in the
current study.
These considerations underscore the importance of recognizing some inherent assumptions of the forearm technique and how “correcting” for muscle’s contribution to
forearm composition is affected by these assumptions. In
forearm studies, the venous catheter is placed retrograde in
the antecubital fossa and advanced into the deep forearm
veins. Therefore, the arterio-venous difference measures metabolites or tracer differences between the systemic arterial
inflow and the venous effluent that drains almost exclusively
muscle and bone. Thus, the metabolic activity measured with
the forearm technique is largely confined to those tissues.
However, the forearm volume (measured by volume displacement) and forearm blood flow (measured by plethysmographic or dye dilution methods) includes contributions
from all tissues in the forearm. The MRI measurements provide accurate estimates of forearm composition. However,
there is no generally accepted method available to estimate
the distribution of blood flow between muscle and sc adipose
tissue and skin. The common practice of virtually all laboratories that use the forearm model (11–16) involves calculating the balance by multiplying the arterio-venous difference across muscle by the total forearm flow. This tacitly
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assumes that muscle, bone, skin and fat behave in a quantitatively similar fashion. For many substrates this seems
unlikely.
By contrast, in leg balance studies, the venous catheter
(placed in the femoral vein) drains venous effluent from skin,
subcutaneous adipose, bone, and muscle. Therefore, for the
leg, venous sampling, blood flow, and volume (measured by
volume displacement) reflect the contributions of leg tissues.
This has obvious advantages, vis-á-vis the above discussion.
However, it does not allow dissection of the metabolic activity of muscle (or muscle and bone) per se as can be performed in the forearm.
Several studies have demonstrated that replacement of
androgens in hypoandrogenic men increases lean body and
muscle mass (4, 17, 18). Likewise, androgen excess in women
increases lean body mass and decreases fat mass. As this
androgen effect must result from an increased rate of protein
synthesis relative to breakdown (17, 19), it is at first surprising that there was not a higher rate of protein synthesis or
more positive protein balance in the male subjects. However,
two caveats must be kept in mind. First, these were postabsorptive subjects, and in the postabsorptive state there is a net
loss of muscle protein in all subjects. Whether males would
show a greater postprandial response to amino acid supply,
insulin, or other anabolic factors cannot be ascertained from
these studies. Second, the methods used have approximately
80% power (a , 0.05) to detect a decrease in protein synthesis
or a balance of 20% in the women. Lesser decrements could
contribute to a lower mass but not be evident by these kinetic
measures. These caveats aside, the current results suggest
that under the conditions studied, the greater concentration
of androgens in these healthy males does not have a major
influence on postabsorptive muscle oxygen, glucose, or protein metabolism.
In summary, despite proportionately greater muscle mass
in males, the rates of glucose uptake, oxygen consumption,
and protein metabolism per unit mass of muscle are similar
in male and female subjects. These findings suggest that sex
hormones are not major regulators, in postabsorptive humans, of skeletal muscle metabolism of glucose and protein
or of the overall metabolic rate.
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