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ORIGINAL

ARTICLE

E n d o c r i n e

C a r e

Vitamin D Status and Its Relation to Muscle Mass and

Muscle Fat in Young Women
Vicente Gilsanz, Arye Kremer, Ashley O. Mo, Tishya A. L. Wren,
and Richard Kremer
Departments of Radiology (V.G., A.K., A.O.M., T.A.L.W.) and Orthopaedic Surgery (V.G., T.A.L.W.), Keck
School of Medicine, University of Southern California, Los Angeles, California 90027; and Department of
Medicine (R.K.), McGill University Health Center, McGill University, Montreal, Quebec, Canada H9X 3V9

Context: Vitamin D insufficiency has now reached epidemic proportions and has been linked to
increased body fat and decreased muscle strength. Whether vitamin D insufficiency is also related
to adipose tissue infiltration in muscle is not known.
Objective: The objective of the study was to examine the relationship between serum 25-hydroxyvitamin D (25OHD) and the degree of fat infiltration in muscle.
Design: This was a cross-sectional study.
Outcome Measures and Subjects: Measures were anthropometric measures, serum 25OHD radioimmunoassay values, and computed tomography (CT) values of fat, muscle mass, and percent
muscle fat in 90 postpubertal females, aged 16 22 yr, residing in California.
Results: Approximately 59% of subjects were 25OHD insufficient (29 ng/ml), of which 24% were
deficient (20 ng/ml), whereas 41% were sufficient (30 ng/ml). A strong negative relationship
was present between serum 25OHD and CT measures of percent muscle fat (r 0.37; P 0.001).
In contrast, no relationship was observed between circulating 25OHD concentrations and CT measures
of thigh muscle area (r 0.16; P 0.14). Multiple regression analysis indicated that the relation
between 25OHD and muscle adiposity was independent of body mass or CT measures of sc and visceral
fat. Percent muscle fat was significantly lower in women with normal serum 25OHD concentrations
than in women with insufficient levels and deficient levels (3.15 1.4 vs. 3.90 1.9; P 0.038).
Conclusions: We found that vitamin D insufficiency is associated with increased fat infiltration in
muscle in healthy young women. (J Clin Endocrinol Metab 95: 0000 0000, 2010)

itamin D, a key regulator of bone metabolism, is also

known to be significantly associated with muscle
strength (1). A lack of vitamin D can cause myopathy (2,
3), which tends to be more marked in the proximal muscles
(4). In the elderly, vitamin D deficiency is linked to muscle
weakness, increased body sway, and increased susceptibility to falls and fractures, which are improved by the
administration of vitamin D with calcium (513). Vitamin
D levels have also been shown to be significantly associated with muscle strength in healthy postmenarchal girls,
suggesting that muscle contractility may be affected by

vitamin D status (1). The mechanisms underlying the effect of vitamin D on muscle strength are not fully understood but could be related to an independent effect on
muscle mass, or alternatively, to enhancement of muscle
function mediated through the effect of vitamin D. Nonetheless, vitamin D action requires activation of the vitamin
D receptor, which is widely distributed in various tissues
including skeletal muscle (14, 15).
Available data indicate that a higher muscle lipid content, measured as muscle attenuation with computed tomography (CT), is associated with lower levels of muscle

ISSN Print 0021-972X ISSN Online 1945-7197

Printed in U.S.A.
Copyright 2010 by The Endocrine Society
doi: 10.1210/jc.2009-2309 Received October 29, 2009. Accepted January 21, 2010.

Abbreviations: BMI, Body mass index; CT, computed tomography; CV, coefficient of variation; 25OHD, 25-hydroxyvitamin D; SF, sc fat; VF, visceral fat.

J Clin Endocrinol Metab, April 2010, 95(4):0000 0000

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<zshorttitle>Vitamin D Status and Muscle Fat

jcem.endojournals.org

balt1/zeg-jcem/zeg-jcem/zeg00410/zeg7208-10z xppws S1 2/10/10 4:23 Art: 09-2309 Input-ss

2

Gilsanz et al.

Vitamin D Status and Muscle Fat

strength and physical performance, independent of muscle

mass (16, 17). Additionally, several studies in patients
with neuromuscular disorders have demonstrated skeletal
muscle attenuation determined by CT to be strongly reciprocally associated with muscle lipid content (18 20).
In the current study, we propose that 25-hydroxyvitamin D (25OHD) concentrations are reciprocally related to
adipose tissue infiltration in muscle independently of muscle mass. To test this hypothesis, we examined the relation
between vitamin D and skeletal muscle lipid content and
muscle mass using CT in a cohort of healthy young women
living in California who were recently studied and found
to have a reciprocal relation between vitamin D status and
measures of adiposity (21).

Subjects and Methods

Study subjects
The sample of this study was the same 90 postpubertal
healthy females who have been more thoroughly described in a
previous investigation on the relation of vitamin D to body fat
and bone (21). This study was approved by the institutional
review board at our institution, and informed consent was obtained from all parents and/or subjects.
Candidates were excluded if they had a diagnosis of any underlying disease or chronic illness, had been ill for longer than wk
during the 6 months before examination, had been admitted to
the hospital at any time during the prior 3 yr, or were taking any
medications including oral contraceptives. Candidates who were
pregnant, had ever been pregnant, or had absence of menses for
more than 4 consecutive months were also excluded from the
study. To decrease the seasonal variability in biochemical determinations, all appointments were scheduled between May and
October. In addition, all subjects had normal kidney function
and normal liver function tests, and there was no evidence of liver
abnormalities detected by CT. All participants had reached sexual maturity, defined as Tanner V of breast development (22),
and skeletal maturity, defined as epiphyseal closure in the phalanges and metacarpals using the radiographic atlas of Greulich
and Pyle (23).
Levels of physical activity in 83 participants were examined
using a 7-d physical activity recall questionnaire. Participants
were asked to indicate the number of times in the past week that
they had engaged in strenuous, moderate, and mild forms of
physical activity for more than 15 min and the daily time spent
watching television and/or on the computer. This measure represents frequency, intensity, and duration elements of physical
activity and inactivity with a test-retest reliability coefficient of
0.81 (24, 25).

Muscle and fat measurements

CT measurements of muscle and sc fat were determined using
a Hilite Advantage scanner (General Electric Healthcare, Milwaukee, WI) with a standardized reference phantom for simultaneous calibration. At the level of the umbilicus, measurements
of the abdominal visceral fat (VF; square centimeters) and sc fat
(SF; square centimeters) were obtained. For the purpose of this

J Clin Endocrinol Metab, April 2010, 95(4):0000 0000

study, SF was defined as the amount of adipose tissue located

between the skin and rectus muscles of the abdomen, the external
oblique muscles, the broadest muscles of the back, and the erector muscles of the spine at the level of the umbilicus. VF was
defined as the intraabdominal adipose tissue surrounded by the
rectus muscles of the abdomen, the external oblique muscles, the
lumbar quadrate muscle, the psoas muscles, and the lumbar
spine at the same level. The coefficient of variation (CV) for
repeated measures of VF and SF has been reported to range from
1.5 to 3.5% (26). Additionally, measures of sc thigh fat and thigh
muscle area were obtained at the midshafts of the femurs, and
values from both the right and left leg measurements were averaged for analysis; the CVs for repeated CT measurements of sc
fat and muscle in the thigh were previously reported to fall between 1 and 2% (27). Measures of thigh muscle area included the
vastus muscles, adductors, rectus femoris, sartorius, gracilis, and
semitendinosis and semimembranosus muscles.
The tissue densities of muscle and fat were determined by
converting CT values expressed as Hounsfield units into measures of tissue density using an external phantom (28). All muscle
and fat measures were obtained from a 2-cm2 region of the rectus
femoris and a 2-cm2 region of adjacent sc fat. Based on the tissue
densities of fat and muscle, the percentage of adipose tissue in the
muscle was calculated according to the formula: (Mx)/
(My) 100, where M represents the density of muscle without
fat, x represents the density of the muscle, and y represents the
density of fat. For the purpose of this study, the highest muscle
density was used as the reference value representing muscle tissue
with no fat infiltration. The CV for muscle attenuation from
single-slice CT scans of the midthigh have previously been reported to be 0.51% for test-retest variability and 3.3% for within-subject variance (18). The inter- and intraclass correlation
coefficients for measurements of adiposity in soft tissues using
CT and a reference phantom are outstanding at greater than 0.98
(29). The time taken to complete the CT scans was approximately 10 min and the effective radiation dose was approximately 0.1 mSv (30).

Biochemical determinations
Serum levels of 25OHD were assayed using a RIA as described by Hollis et al. (31). The lower limit of detection was 5
ng/ml (12.5 nmol/liter). Goat anti-25OHD was a gift from Dr.
Bruce Hollis. 125I-25-(OH)D3 and donkey antigoat secondary
antibody were purchased from Diasorin (Stillwater, MN). This
assay recognizes 25OHD2 and 25OHD3 equally and shows no
bias when compared with HPLC (32). Calculated assay precision
for within-assay variation averages of 6% and interassay of
16%. For the purpose of this study and according to the current
consensus, subjects were divided into a 25OHD sufficient, or
normal, group (30 ng/ml) and an insufficient group (29 ng/
ml). Intact PTH (1 84) was measured with an electrochemiluminescent assay (33). The sensitivity of the assay is 1.2 pg/ml
(0.127 pmol/liter) and intra- and interassay variations are 1.9 4
and 2.6 6.5%, respectively. To minimize interassay variability,
all samples were analyzed simultaneously.

Statistical analysis
Statistical analysis was carried out using Statview (version
5.0.1; SAS Institute Inc., Cary, NC). Data were analyzed using
simple linear regression analysis, multiple regression analysis,
and unpaired t tests. All values are expressed as mean SD.

AQ: A

balt1/zeg-jcem/zeg-jcem/zeg00410/zeg7208-10z xppws S1 2/10/10 4:23 Art: 09-2309 Input-ss

J Clin Endocrinol Metab, April 2010, 95(4):0000 0000

jcem.endojournals.org

P values
0.001
0.408
0.046
0.048
0.014
0.029
0.009
0.202
0.067
0.038
0.161
Insufficient
(n 53)
21.5 5.9 (6.729.6)
19.5 1.4 (17.0 22.87)
71.3 20.0 (45.5126.0)
162.1 5.1 (153.9 171.8)
27.1 7.1 (17.6 44.5)
288.1 174.0 (63.9 799.7)
44.81 46.83 (2.8 240.8)
83.66 34.01 (33.5177.4)
128.7 69.4 (14.4 324.0)
3.90 1.85 (0.0 9.5)
8.4 36.5 (1.531.8)
Measures of Inactivity were obtained in 83 subjects: 34 in the sufficient and 49 in the insufficient group.

In this study, we used CT to investigate whether muscle

mass and muscle adiposity in healthy young adult females
are associated with vitamin D. Our data demonstrated
that serum 25OHD was inversely related to the percent fat
of skeletal muscle, a relation that was independent of body
mass or CT measures of sc and visceral fat. Compared with
women with normal 25OHD values, vitamin D-insuffi-

Discussion

25OHD (ng/ml)
Age (yr)
Weight (kg)
Height (cm)
BMI
SF (cm2)
VF (cm2)
Thigh musculature (cm2)
Subcutaneous thigh fat (cm2)
Muscle fat (%)
Inactivity (h/wk)a

T3

Sufficient
(n 37)
42.4 10.1 (30.0 69.6)
19.2 1.6 (16.322.8)
63.9 11.9 (45.6 113.0)
164.1 3.9 (156.8 170.3)
23.7 4.6 (16.7 43.9)
203.3 98.9 (59.1 431.9)
24.74 33.88 (5.6 218.2)
92.98 33.50 (36.5151.9)
104.8 45.5 (26.6 219.5)
3.15 1.37 (0.4 5.8)
6.8 10.4 (3.0 16.0)

T2,F1

All
(n 90)
30.1 13.0 (6.7 69.6)
19.4 1.5 (16.322.8)
68.3 17.5 (45.5126.0)
162.9 4.7 (153.9 171.8)
25.7 6.3 (16.7 44.5)
252.8 152.7 (59.1799.7)
36.46 42.89 (2.8 240.8)
87.49 33.92 (33.5177.4)
118.9 61.0 (14.4 324.0)
3.59 1.70 (0.0 9.5)
7.8 5.1 (1.531.8)

T1

Age, anthropometric characteristics, 25OHD, and CT

measures of muscle and muscle fat, separated based on
25OHD concentrations, are described in Table 1. Thirtyseven women (41%) had 25OHD concentrations 30
ng/ml or greater, whereas 57 women (59%) had insufficient 25OHD concentrations (29 ng/ml), of which 22
women (24%) were vitamin D deficient (20 ng/ml).
Compared with women with normal 25OHD values, vitamin D-insufficient subjects were significantly shorter
and heavier and had greater body mass index (BMI) and
abdominal SF and VF.
CT measures of muscle attenuation were significantly
lower in the 25OHD-insufficient group when compared
with women with normal 25OHD values. The percentage
of fat infiltration of the muscle derived from the attenuation values was also greater in women with lower
25OHD concentrations. In contrast, there were no differences in CT values for muscle area in the thighs between
women in the two vitamin D groups. This was true
whether the right or left thighs were analyzed independently (data not shown) or both extremities analyzed
together.
Significant inverse correlations were observed between
25OHD and BMI, SF, VF, sc thigh fat, the degree of inactivity, and percent muscle fat; the strongest with percent
muscle fat (Table 2 and Fig. 1). In contrast, neither age nor
CT measures of muscle area were associated with 25OHD
levels. Measures of muscle fat were unrelated to BMI, SF,
VF, sc thigh fat, and measures of inactivity but inversely
related to muscle area (Table 2).
Multiple regression analysis indicated that the relation
between 25OHD levels and percent muscle fat was independent of body mass and measures of inactivity (Table 3).
Similar results were found when measures of sc and visceral adiposity replaced BMI as an independent variable in
this model (data not shown).
A significant inverse correlation was found between
25OHD and PTH (r 0.27; P 0.01), and PTH values
were higher in the insufficient than in the sufficient group
(2.28 0.88 and 1.92 0.90, respectively; P 0.025).

TABLE 1. 25OHD values, age, anthropometric characteristics, and CT measures of muscle for 90 women separated by 25OHD concentration groups

Results

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4

Gilsanz et al.

Vitamin D Status and Muscle Fat

J Clin Endocrinol Metab, April 2010, 95(4):0000 0000

TABLE 2. Relations between 25OHD, anthropometric

characteristics, and CT measures of muscle and fat in 90
women
25OHD (ng/ml)
Age (yr)
BMI (kg/m2)
SF (cm2)
VF (cm2)
Thigh muscle area (cm2)
Subcutaneous thigh fat (cm2)
Inactivity (h/wk)a
Muscle fat (%)
Age (yr)
BMI (kg/m2)
SF (cm2)
VF (cm2)
Thigh muscle area (cm2)
Subcutaneous thigh fat (cm2)
Inactivity (h/wk)a
a

0.17
0.35
0.36
0.28
0.16
0.32
0.27

0.101
0.001
0.001
0.007
0.138
0.002
0.013

0.28
0.09
0.08
0.04
0.24
0.05
0.17

0.007
0.411
0.459
0.690
0.024
0.634
0.122

Measures of inactivity were obtained in 83 subjects.

cient women had approximately 24% greater muscle fat

infiltration than women with normal serum 25OHD concentration. In contrast, we found no relation between
thigh muscle area and serum levels of vitamin D and no
significant differences in the cross-sectional area of thigh
muscles between women with and without vitamin D
insufficiency.
It should be noted that none of the young women had
symptoms of overt vitamin D insufficiency or complained
of muscle weakness, yet we were able to show the association of 25OHD with fat accumulation in skeletal muscle. However, muscle adiposity is known to strongly influence muscle strength, and available data indicate that
im fat accumulation increases with reduced physical activity and decreases with exercise (16, 34, 35). It has also
been shown that exercise training in both lean and obese
subjects significantly improved the ability to oxidize lipids
in skeletal muscle (34, 36). Although we did not measure

FIG. 1. Relation between vitamin D concentrations and CT measures

of muscle fat infiltration.

TABLE 3. Multiple regression analysis with serum levels

of vitamin D as the dependent variable in 83 women
25OHD (ng/ml)
Muscle fat (%)
BMI (kg/m2)
Inactivity (h/wk)

SE

P value

R2

2.94
0.65
0.29

0.72
0.20
0.25

0.001
0.001
0.254

0.29

muscle strength, a recent study by Ward et al. in a group

of healthy postmenarchal adolescents (1) demonstrated
serum 25OHD to be positively related to muscle power,
force, velocity, and jump height. Because these associations, like those found in the current study, were independent of BMI, they cannot be ascribed to overall body
adiposity.
In the current study, we found no relation between vitamin D levels and the cross-sectional area of the muscles
in the thigh. However, vitamin D supplementation has
been reported to enhance muscle strength without any
apparent changes in muscle mass (11, 37, 38). Indeed,
whereas muscle mass and strength are often associated,
this association is relatively weak (39), and muscle
strength can significantly improve without significant
changes in muscle mass (40 42). Whereas measures of
inactivity were positively related to weight and reciprocally related to vitamin D, neither CT measures of muscle
area nor muscle adiposity was associated with time spent
watching television and on the computer. Hence, it is possible that the muscle fat content in our subjects was a direct
consequence of vitamin D insufficiency rather than the
result of reduced vitamin D associated with increased adiposity due to inactivity.
Although the mechanism(s) underlying the vitamin
D-muscle fat link is unknown, it is possible that that im
adipose tissue accumulation could regulate muscle metabolism. Vitamin D has been shown to mediate protein
synthesis and cellular ATP accumulation (43), increase
troponin C (44), and increase actin and sarcoplasmic protein expression (45) in striated muscles; whether these
and/or other cellular events are influenced by im fat requires further study. Regardless of the mechanism by
which vitamin D influences skeletal muscle composition,
im lipids play an important role in the function of myocytes. Skeletal muscle fat infiltration impairs skeletal muscle mitochondrial function (46) and decreases the rate of
mitochondrial oxidative phosphorylation (47). Furthermore, muscle triglyceride content reduces insulin-stimulated glucose uptake within muscle (48, 49). Other studies
indicate that im adipose tissue deposition is associated
with insulin resistance and diabetes (50 52) and accompanied by decreased glucose use, decreased signaling
through the phosphatidylinositol 3-kinase/Akt pathway,

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J Clin Endocrinol Metab, April 2010, 95(4):0000 0000

and decreased insulin-stimulated glycogen synthesis (52,

53). Additionally, low levels of circulating 25OHD have
also been associated with glucose intolerance (54, 55) and
insulin resistance (56) in adults, suggesting a possible link
between vitamin D status, muscle fat content, and glucose
homeostasis.
The high prevalence of vitamin D insufficiency in this
young population, living in a sun-rich area, even during
spring and summer months when vitamin D synthesis is
greatest, is surprising and likely multifactorial. Close to
60% of women studied had insufficient 25OHD concentrations (29 ng/ml), and close to 25% were vitamin D
deficient (20 ng/ml). These findings are, however, consistent with international data suggesting that even in the
sunniest areas of the world, vitamin D deficiency is common. Studies in Turkey, Lebanon, Saudi Arabia, United
Arab Emirates, India, and Australia show 30 50% of children and young adults to have 25OHD levels less than 20
ng/ml (57). Moreover, a recent study found that a substantial proportion (60%) of healthy adolescents living in
a sunny Brazilian climate had vitamin D insufficiency (58).
There are limitations to our study, including its crosssectional design and the inability to establish a causal relation between skeletal muscle adipose tissue infiltration
and vitamin D. We used a single CT measure obtained at
the midthigh as a surrogate of muscle volume to minimize
radiation exposure and were able to measure only a relatively small depot of skeletal muscle adipose tissue. However, previous studies noted a relatively strong correlation
between the skeletal muscle fat content at different sites
(59). Moreover, our study subjects were not recruited
from the community at large and were limited to young
females. Future studies are needed to determine whether
vitamin D influences fat infiltration in the muscle in males
and the elderly. Nevertheless, serum 25OHD is related
to overall physical fitness in postmenopausal women
(60), and ample data indicate that inadequate vitamin D
status is related to muscle weakness and the risk of falls
in older persons, regardless of gender (35). Lastly, studies are needed to determine whether the increased muscular adiposity (17, 31, 49) and decreased muscle
strength and mobility (57 60) with aging are associated
with vitamin D status. This is particularly relevant because vitamin D insufficiency is highly prevalent in the
elderly population (61, 62).
In conclusion, our data show that vitamin D insufficiency is significantly and inversely associated with the
degree of fat infiltration in skeletal muscle, a relation that
is independent of body mass. This reciprocal association
between vitamin D status and muscle fat was not previously reported and is unexplained and intriguing. Further

jcem.endojournals.org

studies are needed to characterize the possible mechanistic

relationship between muscle fat and vitamin D.

Acknowledgments
The authors express their gratitude to Dr. Timothy Reinhardt for
providing the measures of serum 25OHD.
Address all correspondence and requests for reprints to:
Vicente Gilsanz, M.D., Ph.D., Department of Radiology, Childrens
Hospital Los Angeles, MS no. 81, 4650 Sunset Boulevard, Los
Angeles, California 90027. E-mail: vgilsanz@chla.usc.edu.
This work was supported by National Institutes of Health
Grant 1R01 AR052744-01, Department of the Army Grant
DAMD17-01-1-0817, Canadian Institutes of Health Research
Grant MT-10839, NSERC/Dairy Farmers of Canada, and Natural Sciences and Engineering Research Council and Dimensional Fund Advisors Canada.
Disclosure Summary: V.G., A.K., A.O.M., T.A.L.W., and
R.K. have nothing to declare.

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JOBNAME: AUTHOR QUERIES PAGE: 1 SESS: 3 OUTPUT: Wed Feb 10 04:25:28 2010
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AUTHOR QUERIES
AUTHOR PLEASE ANSWER ALL QUERIES
APlease provide affiliation and location for Hollis.
BPlease confirm that phosphatidylinositol 3-kinase is the correct expansion of PIK3.
CPlease write out NSERC.
DPlease provide exact title of book (Physical growth and development or Textbook of
pediatrics), and provide city of publication.
EPlease verify volume number and page range added to Reference 42.