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Physical activity and sex hormone levels in postmenopausal women

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Physical activity and sex hormone levels in postmenopausal women

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PHYSICAL ACTIVITY AND SEX HORMONE LEVELS
IN POSTMENOPAUSAL WOMEN

by

Farzana Choudhury

A Thesis Presented to the
FACULTY OF THE GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
MASTER OF SCIENCE
(APPLIED BIOSTATISTICS AND EPIDEMIOLOGY)

December 2008

Copyright 2008 Farzana Choudhury
ii
DEDICATION

Dedicated to my family
iii

ACKNOWLEDGEMENTS

I would like to express my utmost gratitude to Dr. Wendy Mack for her inspiration and
guidance throughout my thesis experience. I am grateful to Drs. Leslie Bernstein,
Howard Hodis and Stanley Azen for their guidance and support in preparing the thesis. I
am also thankful to my friends Towhid, Roksana, Talat and Jessica for their additional
suggestions and encouragement. Special thanks go to my husband and my family for their
unconditional and persistent support.

iv
TABLE OF CONTENTS

Dedication.......................................................................................................................ii

Acknowledgements ........................................................................................................iii

List of Tables……………………………………………………………………………...v

Abstract…………………………………………………………………………………...vi

Introduction and Background ......................................................................................1

Methods .........................................................................................................................7
Study Design...............................................................................................................7
Study Variables ...........................................................................................................8
Sex Steroid Hormones .............................................................................................8
Physical Activity .....................................................................................................9
Other Reproductive and Anthropometric Measures................................................10
Statistical Analysis ....................................................................................................12

Results..........................................................................................................................15
Baseline Characteristics.............................................................................................15
Treatment Group Comparisons on Hormone Levels...................................................15
Associations of Physical Activity with Endogenous Sex Hormone Levels .................17
(Placebo Group) ........................................................................................................17
Associations of Physical Activity with Exogenous Hormone Levels
(Estradiol-Treated Group)..........................................................................................22

Discussion ....................................................................................................................27
Associations in Untreated Women.............................................................................27
Associations in Estradiol-Treated Women .................................................................30
Strengths and Limitations ..........................................................................................31
Conclusion ................................................................................................................32

Bibliography .................................................................................................................33

v

LIST OF TABLES

Table 1: Distribution of Explanatory Variables by Treatment Group .............................16

Table 2: Baseline and On Trial Hormone Levels by Treatment Group ...........................17

Table 3: EPAT Placebo Participants: Total-MET-hours and Sex Hormone
levels (n=104) ...............................................................................................................18

Table 4: EPAT Placebo Participants: Total-MET-hours Quartiles and Sex Hormone
Levels (n=104) ..............................................................................................................19

Table 5: EPAT Placebo Participants: Moderate or More Physical Activity
(hours/week) and Sex Hormone Levels (n=104) ............................................................19

Table 6: EPAT Placebo Participants: Moderate or More Physical Activity Tertiles
(hours/week) and Average Hormone Levels (n= 104)....................................................20

Table 7: EPAT Placebo Participants: Vigorous Physical Activity (hours/week)
and Sex Hormone Levels (n=104) .................................................................................21

Table 8: EPAT Placebo Participants: Vigorous Physical Activity Categories
(hours/week) and Sex Hormone Levels (n= 104) ...........................................................22

Table 9: EPAT Estradiol participants: Total-MET-hours and Sex Hormone
levels (n=90) .................................................................................................................23

Table 10: EPAT Estradiol Participants: Total-MET-hours Quartiles and Sex
Hormone Levels (n=90) ................................................................................................23

Table 11: EPAT Estradiol participants: Moderate or More Physical Activity
(hours/week) and Sex Hormone Levels (n=90) ..............................................................24

Table 12: EPAT Estradiol Participants: Moderate or More Physical Activity
Tertiles (hours/week) and Average Hormone Levels (n=90) ..........................................25

Table 13: EPAT Estradiol participants: Vigorous Physical Activity (hours/week)
and Sex Hormone Levels (n=90) ...................................................................................25

Table 14: EPAT Estradiol Participants: Vigorous Physical Activity Tertiles
(hours/week) and Average Hormone Levels (n=90).......................................................26

vi
ABSTRACT

The crucial yet complex relationships between physical activity and physiologic and
pharmacologic sex hormone levels in postmenopausal women have not been investigated
sufficiently. In this post hoc cohort analysis of data from Estrogen in the Prevention of
Atherosclerosis Trial, physical activity was tested for an association with sex hormone
levels, assessed longitudinally over two years in 194 postmenopausal women, 90
randomized to 17β-estradiol (e2) and 104 to placebo. Physical activity (total energy
expenditure and weekly hours spent in moderate or more physical activity) was
associated with decreased serum levels of estradiol in both groups [Placebo: (p=0.02); e2-
treated group: (p=0.002)]. Physical activity was associated with reduced androgen levels
[testosterone and androstenedione (p<0.001 for both)] and elevated SHBG level
(p<0.001) in the placebo group only.
1
INTRODUCTION AND BACKGROUND

There were an estimated 600 million postmenopausal women in the world in 2000
(Hill 1996). About 25 million women pass through menopause each year and by 2030,
the world population of menopausal and postmenopausal women is projected to increase
to 1.2 billion, with 47 million new entrants each year (Hill 1996). Many of these women
will spend one-third of their life in menopause (Cutson and Meuleman 2000).
Menopause results in considerable physical and psychological changes.
Reproductive hormonal fluctuations in menopause have been associated with most of the
disorders and symptoms associated with menopause, although the nature of the
relationships is still inconclusive (McTiernan, Kooperberg et al. 2003). Almost all
women suffer to some extent due to menopause. The most common short-term problems
are vasomotor symptoms (hot flushes and night sweats) (Warren 2007). Mood changes,
sleep disturbances, deceased libido and somatic complaints such as breathing difficulty,
palpitation, sleepiness and vaginal dryness are common (Warren 2007). Menopause is
also associated with increased risk of chronic diseases including cardiovascular diseases,
breast cancer and osteoporosis. Less serious problems include dyspareunia and urinary
incontinence (Cutson and Meuleman 2000).
A large body of observational evidence suggests a positive association between
cardiovascular disease risk and menopause varying by ethnic background and geographic
region (Witteman, Grobbee et al. 1989; Hill 1996). This association is hypothesized to be
related to menopause-associated metabolic and hormonal changes (Witteman, Grobbee et
al. 1989; Zarate, Saucedo et al. 2007). Some studies suggest that an increase in androgens
2
relative to estrogen in healthy postmenopausal women is associated with an unfavorable
cardiovascular risk profile (Rexrode, Manson et al. 2003). A study using data from the
Estrogen in the Prevention of Atherosclerosis (EPAT) trial demonstrated an inverse
association of estrogen and sex-hormone binding globulin (SHBG) with subclinical
atherosclerosis progression in healthy postmenopausal women with or without hormone
therapy, while total testosterone had a borderline significant inverse association (Karim,
Hodis et al. 2008). Other studies failed to find any association between physical activity
and sex hormones (Barrett-Connor and Goodman-Gruen 1995).
While hormone therapy (HT) has been the focus of intervention in menopause, its
risk-benefit ratio is still debated (Tavani and La Vecchia 1999). Although the
heterogeneity of hormonal regimen and timing of initiation relative to menopause makes
generalization difficult, HT has been shown to have favorable effects on bone
metabolism and osteoporosis as well as cardiovascular disease risk (Hodis and Mack
2007). Numerous observational studies including the Framingham Heart Study and the
Nurses' Health Study showed that HT use is associated with protection from
cardiovascular disease (McPherson 2000).) Results from EPAT suggest that estrogen
therapy slows subclinical atherosclerosis progression in post-menopausal women (Hodis,
Mack et al. 2001; Karim, Hodis et al. 2008). Results from the Women’s Health Initiative
(WHI) study suggest women who were randomized to HT closer to menopause tended to
have a reduced risk of cardiovascular disease compared to the placebo group, whereas
women who were randomized to HT distant from menopause tended to have an increased
risk of CHD compared to the placebo group (Rossouw, Prentice et al. 2007). Compared
to the placebo group, women randomized within 10 years of menopause had a hazard
3
ratio of 0.76 (95% confidence interval [CI], 0.50-1.16); for those randomized within10 to
19 years, the hazard ratio was 1.10 (95% CI, 0.84-1.45); and for those randomized at 20
or more years, the hazard ratio was 1.28 (95% CI, 1.03-1.58). There was a statistically
significant trend in the change of impact based on time from menopause (P-trend = 0.02)
(Rossouw, Prentice et al. 2007).
HT has also been associated with a number of adverse events, primarily elevated
risk of estrogen-dependent cancers including endometrial cancer and breast cancer, with
risk increasing with increasing duration of use (Tavani and La Vecchia 1999; Persson
2000; Beral, Banks et al. 2002). The increase in the risk of for endometrial cancer is due
to estrogen alone therapy and for breast cancer it appears to be combined therapy (Beral,
Banks et al. 2002). Among women participating in observational studies who are current
or recent users of HT, risk of breast cancer is modestly elevated with combined estrogen-
progestin therapy (Bernstein 2006). Results from one component of the WHI trial, which
included postmenopausal women with intact uterus, indicate an increased risk of breast
cancer in participants randomized to combined conjugated equine estrogens (CEE) and
progestin therapy compared to participants receiving placebo in women with(HR=2.19,
95% CI, 1.56, 3.09) (Prentice, Chlebowski et al. 2008). Results from another component
involving postmenopausal women with history of hysterectomy suggest a lower breast
cancer risk in those receiving unopposed CEE alone (HR=0.72, 95% CI, 0.42, 0.84)
(Prentice, Chlebowski et al. 2008). High endogenous estrogen levels have also been
associated with increased breast cancer risk in postmenopausal women, albeit
inconsistently (Bernstein and Ross 1993). Cumulative exposure to estrogen seems
particularly important and may be reflected by risk factors such as age at menarche, age
4
at menopause, pregnancy, use of oral contraception and lactation (Bernstein 2002). In a
case-control study, post menopausal breast cancer cases had 15% higher serum estradiol
levels (P = 0.02), 40% higher urinary estradiol (P = 0.03) and 44% higher urinary estriol
(P = 0.04) levels, compared to the controls (Bernstein, Ross et al. 1990).
Low level of physical activity has been associated with increased cardiovascular
disease in postmenopausal women in many studies (Dubnov, Brzezinski et al. 2003).
Physically active postmenopausal women have a lower incidence of coronary heart
disease (CHD) compared to sedentary women (Stevenson, Davy et al. 1995). Numerous
studies have investigated the relationship of physical activity and breast cancer risk; 9 of
the 10 studies involving postmenopausal women reported an inverse association
(Monninkhof, Peeters et al. 2007).
The mechanisms by which physical activity exerts its beneficial effects are still
being explored and debated. The facts that, in postmenopausal women, high sex steroid
levels, especially estrogen, elevate breast cancer risk and physical activity reduces breast
cancer risk, implicate a possible relationship between physical activity and sex hormone
levels (McTiernan, Tworoger et al. 2004). A plausible mechanism by which physical
activity may modulate disease risk is through hormone-related pathways (Chan, Dowsett
et al. 2007). However, observational studies have yielded somewhat inconsistent
associations between physical activity and sex hormone levels. The effect of physical
activity separately on endogenous hormone levels in women not exposed to exogenous
hormones and endogenous levels of hormone in women exposed to exogenous sex
hormones in postmenopause remains largely unexplored (Villareal, Steger-May et al.
2004)
5
In a recent large cross-sectional study involving 2082 postmenopausal women,
physical activity levels were significantly associated with lower circulating levels of
estradiol and testosterone, whereas SHBG followed a U-shaped relationship to physical
activity (Chan, Dowsett et al. 2007). These findings are supported by another
observational study which found an inverse relationship between estrone and estradiol
levels and physical activity in postmenopausal women (Cauley, Gutai et al. 1989). In
another study, compared to sedentary postmenopausal women, endurance-trained
postmenopausal women had lower estrogen, specifically estrone, levels (Nelson, Sammel
et al. 2008). In 125 postmenopausal women, androstenedione, estrone, and estrone sulfate
were inversely associated with non-recreational, but not with recreational, physical
activity (Madigan, Troisi et al. 1998). Other studies, however, failed to find any
association between physical activity and sex hormones in postmenopausal women
(Bjornerem, Straume et al. 2004; Copeland, Chu et al. 2004).
To date, only one randomized controlled trial has been published that investigated
the effects of physical activity on endogenous sex hormone levels in postmenopausal
women. In the “Physical Activity for Total Health” study (PATH study), exercise
intervention resulted in a significant decrease in female sex hormone levels at 3 months
(McTiernan, Tworoger et al. 2004; McTiernan, Tworoger et al. 2004). At 12 months, the
direction of effect remained the same, although the differences were no longer
statistically significant. Also at 12 months, women in the exercise intervention group
showed reductions in total testosterone and free testosterone, compared to the controls.
However, the associations were statistically significant only in women who lost more
than 2% of body fat (p=0.02). Three other physical activity intervention trials in
6
postmenopausal women are under way. These are the Sex Hormones and Physical
Activity Study (SHAPE study), Alberta Physical Activity and Breast Cancer Prevention
Trial (ALPHA trial) and The Nutrition and Exercise for Women (NEW) Trial
(Monninkhof, Peeters et al. 2007).
In summary, the crucial yet complex relationship between physical activity and
sex hormone levels in postmenopause has not been investigated sufficiently. Moreover,
the few pertinent studies investigating endogenous sex hormone modulation by physical
activity have yielded somewhat inconsistent results. The effects of physical activity on
endogenous levels of sex hormones in women exposed to exogenous hormones remain
unexplored.
Therefore, we utilized the data from the Estrogen in the Prevention of
Atherosclerosis trial (EPAT) to study the relationship of physical activity and sex
hormone levels in postmenopausal women. The objectives of this analysis were:
1. To determine the association between physical activity and sex hormone levels in
postmenopausal women not randomized to ET.
2. To determine the association between physical activity and sex hormone levels in
postmenopausal women randomized to ET.
7
METHODS
This study uses a post hoc cohort analysis of data collected from a randomized
clinical trial, the Estrogen in the Prevention of Atherosclerosis Trial (EPAT). We utilized
these data to assess the cross-sectional associations between self-reported levels of
physical activity and serum levels of sex steroid hormones.
Study Design
EPAT was a randomized, double-blind, placebo-controlled trial involving 222
postmenopausal women, aged 45 and older, who had no preexisting cardiovascular
disease. The trial tested the primary hypothesis that unopposed ET reduces the
progression of subclinical atherosclerosis in this population (Hodis, Mack et al. 2001).
Potential participants were prescreened by telephone and then seen at three screening
visits for collection of baseline data and determination of study eligibility.
Women were eligible if they were postmenopausal (serum estradiol less than 20 pg/mL),
45 years of age or older, and had a low-density lipoprotein (LDL) cholesterol level of 130
mg/dL or greater. Women with diabetes mellitus were included if their fasting blood
glucose level was less than 200 mg/dL.
The exclusion criteria were: breast or any gynecologic cancer diagnosed in the
past 5 years or identified during screening, previous use of HT for more than 10 years or
within 1 month of the first screening visit, history of five or more hot flushes daily that
interfered with daily activity, diastolic blood pressure greater than 110 mm Hg, untreated
thyroid disease, life-threatening disease with a survival prognosis of less than 5 years,
total triglyceride level of 400 mg/dL or greater, high-density lipoprotein (HDL)
8
cholesterol level less than 30 mg/dL, serum creatinine concentration greater than 2.5
mg/dL, and current smokers.
The participants were followed every month for the first 6 months and every other
month thereafter for a total of 2 years. In addition to the assigned treatment, participants
received dietary counseling and lipid-lowering medication if their LDL cholesterol level
exceeded 160 mg/dl. At each visit, medication adherence, vital signs, clinical events and
use of non-study medications and dietary supplements were ascertained. Blood was
drawn to assess lipid and hormone levels. Physical activity was assessed by a seven-day
recall questionnaire at baseline and every six months.
To address the original hypothesis of the study, a sample size of 200 (100
participants per treatment group) was needed to detect a treatment effect size of 0.40 or
greater with 80% power. Considering anticipated dropout rate, 222 women were recruited
(111 per group). Written informed consent was obtained from all participants. The study
protocol was approved by the Institutional Review Board of the University of Southern
California.
Study Variables
Sex Steroid Hormones
Prior to randomization and at every 6 months during trial follow-up, fasting blood
was drawn and stored at –70
0
C. At the end of the trial, sex hormone levels were
measured from stored samples. All samples for a given participant were processed in the
same batch. Radio-immuno assay (RIA) was used to quantify serum levels of
androstenedione, DHEA, testosterone, estrone and estradiol. Prior to RIA, steroids were
extracted from serum with hexane:ethyl acetate (3:2). Androstenedione, DHEA, and total
9
testosterone were then separated by Celite column partition chromatography (Karim,
Hodis et al. 2008). Estrone and estradiol were separated by ethyl acetate in
trimethylpentane. SHBG was quantified by direct immunoassay using the Immulite
analyzer (Diagnostic Products Corporation, Inglewood, CA). Free testosterone and free
estradiol were calculated using total testosterone and total estradiol, respectively, as well
as SHBG concentrations and an assumed constant for albumin in a validated algorithm
(Sodergard, Backstrom et al. 1982). Intra-assay and inter-assay coefficients of variation
ranged from 4 to 8% and 8 to 13%, respectively.
We computed the average value of sex hormone, SHBG and physical activity
levels for each participant. Thus, each participant contributed one observation to the
analysis. We checked for the variations across time, outliers and errors of all
measurements. All measurements (baseline and on-trial) were used to calculate the means
of sex hormones and SHBG for the placebo group. For the estradiol-treated group, only
the on-trial measurements were used to calculate these means.
Physical Activity
The Seven-Day Physical Activity Recall Questionnaire was used at baseline and
every 6 months. This questionnaire is a self-report recall instrument designed to assess
physical activity during the previous week (Blair, Haskell et al. 1985). Respondents are
asked about the number of hours spent in sleep, and specific activities and duration of
moderate, hard, and very hard activities during the preceding week. Examples of the
types of activities in each category are provided, and the week is separated into weekend
10
days and weekdays. The remaining amount of time was presumed to have been spent in
light activities.
Each reported activity was coded for the metabolic cost of each activity expressed
in metabolic equivalents (METs), using the coding scheme provided by “Compendium of
Physical Activities: Classification of energy costs of human physical activities”
(Ainsworth, Haskell et al. 1993; Ainsworth, Haskell et al. 2000). One MET is the ratio of
the energy expenditure of a given activity to the resting metabolic rate, which is the
energy used as one sits quietly or reads a book. For each 7-day period recalled, the energy
expenditure of each activity was calculated by multiplying participation (hours/week) by
the metabolic cost of each activity expressed in METs. Any self-reported activity that
required 3-6 METs was considered moderate-intensity physical activity and any that
required more than 6 METs was considered vigorous-intensity physical activity. Energy
expenditure in sleep was calculated by multiplying sleep time with 1 MET. Time spent
in light activity was computed by subtracting the sum of time spent in sleep, moderate
and vigorous activity from the total 168 weekly hours. The metabolic cost of light
activity was computed by multiplying the number of hours spent in light activities times
1.5 MET (CDC 2008). Total energy expenditure was the sum of energy expenditure that
summed over sleep, light, moderate, hard and very hard activities. It was the total average
activity of the multiple measures of physical activity collected at baseline and every 6
months.
11
Other Reproductive and Anthropometric Measures
Reproductive history and demographic factors including age, race, income, and
education were assessed by structured questionnaires completed at baseline prior to
randomization. Menstrual status was assessed by self-report and validated by levels of
serum estradiol (less than 20pg/ml). To determine age at menarche, participants were
asked: ‘‘How old were you when you had your first menstrual period?’’ Age at first birth
was ascertained based on their response to ‘‘Have you had any children?’’, and if ‘‘yes’’,
the year of birth of each child was recorded. Total number of pregnancies, abortions and
still births were similarly assessed. Parity was calculated based on the number of
childbirths recorded. History of oral contraceptive use and use of hormone therapy were
also obtained.
Information regarding age at menopause, history of hysterectomy and
oophorectomy was collected on the reproductive history questionnaire. For women
without a hysterectomy/oophorectomy, age at last menstrual period was considered their
age at natural menopause. For those women with a prior bilateral oophorectomy, age at
surgery was considered as the age of menopause. For 15 participants with unilateral
oophorectomy and 26 participants with hysterectomy without oophorectomy, the mean
menopausal age of the sample was imputed as their menopause during analyses.
Smoking history was also obtained by structured questionnaire. Fasting plasma total
cholesterol (TC), total triglyceride (TG) levels, HDL-cholesterol (HDL-C), and fasting
insulin and glucose levels were also measured.
The participants’ height and weight in light clothing without shoes were measured
with a free-standing stadiometer and calibrated digital scale (Salter) to the nearest 0.1 cm
12
and 100 g, respectively. Body mass index (BMI) was calculated as weight (kg) divided
by the square of height (m
2
).

Statistical Analysis
We did not have baseline and follow-up data for physical activity as well as all
hormones on all of the 222 participants. Therefore, the post hoc analysis included 104 of
111 participants in the placebo group and 90 of the 111 participants in the estradiol
group.
The demographic characteristics between the estradiol and placebo groups were
compared using t-tests for independent samples for comparison of means or a chi-square
test for comparison of proportions. Baseline and average on-trial levels of sex hormones
were compared between the treatment groups using independent t-tests. In both groups,
total estradiol and estrone levels were not normally distributed. Therefore, these variables
were log-transformed to approximate the normal distribution during analysis and back
transformed for reporting the results. The physical activity measures were very stable
over time in both groups [co-efficient of variation: 2.6 (placebo) 3.7 (e2-treated)].
Estradiol and estrone levels showed significant variability, especially in estradiol-treated
group [co-efficient of variation: 34.2 (estradiol) 26.3 (estrone)].
Each sex hormone was modeled separately for its association with physical
activity. We used general linear models to assess the continuous relationship between
average physical activity (independent variable) and average sex hormone levels
(dependent variables). Analysis of variance was used to compare the means of hormone
levels across categorical levels of different physical activity variables. Initially, hormones
13
were modeled unadjusted for any covariates. Possible confounding by a factor was
addressed by noting the change in the beta-estimate of the main effect after adding the
covariate to the main effect model. Age at randomization, race, BMI, age at menopause,
type of menopause (natural, surgical), smoking status (ever, never smoked regularly) and
years since menopause were tested as possible confounders, in light of literature review.
Factors that resulted in more than a 15% change in the regression coefficient were
considered as confounders of the relationship between physical activity and sex hormone
levels. Wald’s tests were used to assess possible effect modification by these covariates
after incorporating proper interaction terms in the multivariate model.
Age at randomization, race, BMI and type of menopause resulted in a significant
(more than 15%) change in beta-coefficients. None of the interaction tests were
significant (P>0.10 for all interactions). Therefore, final models included age at
randomization, race, BMI and type of menopause (natural vs surgical). Due to high
correlation between age at randomization and age at menopause (r=0.86), we only kept
age at randomization in the multivariate analyses. Past smoking status and years since
menopause were not significantly associated with hormone levels and including them in
the multivariate model did not alter the impact of physical activity on sex hormones.
Therefore, these variables were not included in the multivariate model.
The serum levels of the different sex hormones and SHBG were modeled as
continuous variables in all analyses. We modeled physical activity in three ways to assess
the association with hormone levels. Total weekly MET expenditure (total MET-hours)
was the sum of energy expenditure as detailed above. Participants were also divided into
14
four quartiles of total MET-hours with the first group (quartile 1 of total MET-hours)
being the least physically active and hence, the referent group.
Physical activity was also modeled as total time (in hours per week) spent in
moderate or higher intensity physical activity (i.e., total hours spent in activities with
MET estimates 3 or greater) and hours per week spent in vigorous-intensity physical
activity (total hours spent in activities with MET estimates greater than 6.0). Time in
moderate or higher physical activity was categorized into three levels based on the tertile
distribution of the total sample. Time in vigorous physical activity was also categorized
into three levels; the lowest levels represented women reporting no vigorous activity,
while the upper two levels were determined by a median split of women who reported
more than 0 weekly hours of vigorous activity. All statistical analysis used SAS software
9.13 (SAS, Inc, Cary, North Carolina) and all tests for significance were conducted at a
2-sided 0.05 level.

15
RESULTS
Baseline Characteristics
The baseline characteristics of the 194 participants in the analysis stratified by
treatment group are summarized in Table 1. Except for type of menopause and
oophorectomy status, the groups did not significantly differ on demographic and
reproductive characteristics. Compared to the placebo recipients, more estradiol
recipients had a history of hysterectomy or surgical menopause (p=0.03). Among those
with surgical menopause, proportionately more estradiol recipients had a history of
complete oophorectomy than the placebo group, but the difference was not statistically
significant (p=0.15). In our subsequent analyses, we adjusted for type of menopause. The
physical activity levels were generally comparable between treatment groups. However,
women in the estradiol group spent significantly more time in vigorous physical activity
per week than placebo-treated women.
Treatment Group Comparisons on Hormone Levels
Table 2 summarizes the comparison of baseline and on-trial hormone levels
between the treatment groups. The groups differed in the level of estrone at baseline
(p=0.02) but were comparable on all other hormone levels. During the trial, the estradiol
group had significantly higher mean levels of estradiol, estrone, bioavailable estradiol,
free estradiol and SHBG and significantly lower mean levels of bioavailable
testosterone, and free testosterone (p<0.001 for all) and DHEA (p=0.02) compared to
placebo participants.

16

Table 1: Distribution of Explanatory Variables by Treatment Group

Placebo group
(N=104)
Estradiol group
(N=90)
P-value*
Race
Non-Hispanic White
Black
Hispanic
Asian
Others

65 (59.1%)
10 (9.1%)
24 (21.8%)
11 (10.0%)
0 (0.0%)

56 (52.8%)
15 (14.2%)
23 (21.7%)
11 (10.4%)
1 (0.9%)

0.62
Annual family Income
<29,900
30,000-59,900
>60,000

45 (45.4%)
33 (33.3%)
21 (21.2%)

47 (49.0%)
34 (35.4%)
15 (15.6%)

0.60
Education level
High school or Less
Trade or Business school, Some college
Bachelors degree or more

4 (3.6%)
69 (62.7%)
37 (33.6%)

2 (1.9 %)
19 (59.4 %)
85 (38.7%)

0.60
Parity
0
1-3
>3

18 (16.4%)
47 (42.7%)
45 (40.9%)

12 (11.3%)
42 (39.6%)
52 (49.1%)

0.38
Age at Menarche
<= 13
>13

59 (53.6%)
51 (46.4%)

56 (52.8%)
50 (47.2%)

0.91
Type of menopause
Natural
Surgical

74 (67.3%)
36 (32.7%)

56 (52.8%)
50 (47.2%)

0.03
Proportion of surgical menopause with bilateral
oophorectomy

19 (57.1%)

31 (72%)

0.15
Age at randomization 61.5 (7.2) 60.5 (6.6) 0.28
BMI (kg/m
2
) 29.0 (5.4) 28.9 (5.9) 0.76
Years since menopause 13.9 (9.7) 13.5 (8.9) 0.84
Total energy expenditure (MET-hours/ week) 237.5 (25.4) 238.0 (16.2) 0.90
Moderate (>3 MET)physical activity (Hrs/Week) 4.1 (6.9) 4.3 (4.1) 0.90
Vigorous (>6 MET ) physical activity (Hrs/Week) 0.2 (0.7) 0.6 (1.2) 0.04
* Chi-square test (categorical variables) and independent t-test (continuous variables)
N (%) for categorical and mean (SD) for continuous variables; MET: Metabolic Equivalent

17

Table 2: Baseline and On Trial Hormone Levels by Treatment Group

Placebo group
(N=104)
Estradiol group
(N=90)
Hormone
Mean (SD) Mean (SD)
P-value*
Estradiol (pg/ml)
Baseline
Ontrial

19.1 ( 8.4)
17.2 ( 12.0)

20.3 ( 12.2)
61.5 (16.2 )

0.53
<0.001
Estrone (pg/ml)
Baseline
Ontrial

44.3 ( 16.5)
46.2 ( 39.2)

48.9 ( 44.6)
296.6 ( 198.1)

0.02
<0.001
Bioavailable Estradiol (pg/ml)
Baseline
Ontrial

14.1 ( 4.7)
14.7 ( 6.7)

16.8 ( 14.2)
39.2 ( 20.2)

0.08
<0.001
Free Estradiol (pg/ml)
Baseline
Ontrial

0.6 ( 0.2)
0.6 ( 0.3)

0.7 ( 0.6)
1.5 ( 0.8)

0.08
<0.001
Testosterone (ng/dl)
Baseline
Ontrial

22.9 ( 9.5)
22.6 ( 9.6)

21.7 ( 10.4)
23.3 ( 11.9)

0.38
0.46
DHEA (ng/ml)
Baseline
Ontrial

2.3 (1.5)
2.1 ( 1.3)

2.2 ( 1.4)
1.9 ( 1.0)

0.55
0.02
Androstenedione (pg/ml)
Baseline
Ontrial

515.1 ( 221.3)
508.4 ( 194.3)

522.9 ( 247.7)
515.7 ( 214.4)

0.82
0.64
Bioavailable testosterone (pg/ml)
Baseline
Ontrial

9.4 ( 4.5)
9.1 ( 4.0)

8.9 ( 4.9)
7.1 ( 4.0)

0.50
<0.001
Free Testosterone (pg/ml)
Baseline
Ontrial

4.3 ( 2.0)
4.1 ( 1.8)

4.1 ( 2.3)
3.2 ( 1.8)

0.50
<0.001
SHBG (nmol/l)
Baseline
Ontrial

35.2 ( 15.1)
36.3 ( 19.3)

36.4 ( 21.6)
57.1 ( 27.3)

0.64
<0.001
*P-value from independent t-test;
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Associations of Physical Activity with Endogenous Sex Hormone Levels
(Placebo Group)
A total of 104 placebo-treated women were available for this analysis. Table 3
summarizes the relationships between continuous measures of average total weekly MET
expenditure (Total MET-hours/week) and hormone and SHBG levels. After adjusting for
age at randomization, race, BMI and type of menopause, total energy expenditure was
18
positively associated with SHBG level (p<0.001) and inversely associated with
testosterone (p<0.001), bioavailable testosterone (p<0.001), free testosterone (p<0.001),
androstenedione (p<0.001), bioavailable estradiol (p=0.02) and free estradiol (p=0.02).

Table 3: EPAT Placebo Participants: Total-MET-hours and Sex Hormone levels
(n=104)

Hormone (mean) Beta** SE P-value*
Estradiol (pg/ml) -2.84 2.70 0.30
Estrone (pg/ml) -9.78 9.11 0.30
Bioavailable Estradiol (pg/ml) -3.94 1.66 0.02
Free Estradiol (pg/ml) -0.15 0.07 0.02
Testosterone (ng/dl) -8.65 3.20 0.007
DHEA (ng/ml) 0.11 0.43 0.80
Androstenedione (pg/ml) -270.23 66.64 <0.001
Bioavailable testosterone (pg/ml) -6.51 1.37 <0.001
Free Testosterone (pg/ml) -2.99 0.63 <0.001
SHBG (nmol/l) 26.52 5.70 <0.001
* Adjusted for age, race, BMI, type of menopause; ** Reported per 100 MET-hours
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

When analyzed by quartile of total energy expenditure (Table 4), results were
consistent with the continuous analysis. Mean SHBG level increased (p<0.001), while
mean levels of testosterone (p<0.001), androstenedione (p<0.001), bioavailable
testosterone (p<0.001), free testosterone (p<0.001), bioavailable estradiol (p=0.03) and
free estradiol decreased (p=0.03) with increasing quartiles of energy expenditure.

Table 5 summarizes the continuous relationship between the total weekly hours
spent in moderate or more intense physical activity (activities rated as 3 or more MET)
and hormone and SHBG levels. The SHBG level was significantly positively associated
with time spent in moderate or higher physical activity (p<0.001). Total hours per week
spent in moderate or higher physical activity was inversely associated with serum levels
of total testosterone (p=0.006), bioavailable testosterone (p<0.001) and free testosterone
(p<0.001).
19

Table 4: EPAT Placebo Participants: Total-MET-hours Quartiles and Sex Hormone
Levels (n=104)

Total MET-hours Quartiles
Mean (SE)
Hormone (mean)
<230
(n=24)
230-236
(n=28)
237-242
(n=25)
>242
(n=27)
P-value*
(linear trend)
Estradiol (pg/ml) 20.63
(1.03)
18.69
(1.04)
16.99
(1.03)
19.00
(1.03)
0.91
Estrone (pg/ml) 47.48
(1.04)
40.32
(1.04)
36.41
(1.04)
40.66
(1.04)
0.35
Bioavailable Estradiol (pg/ml) 16.14
(0.48)
14.51
(0.52)
12.63
(0.47)
13.54
(0.48)
0.03
Free Estradiol (pg/ml) 0.64
(0.02)
0.57
(0.02)
0.50
(0.02)
0.53
(0.02)
0.03
Testosterone (ng/dl) 23.97
(0.95)
20.32
(1.03)
23.32
(0.93)
19.50
(0.95)
<0.001
DHEA (ng/ml) 2.24
(0.13)
1.91
(0.14)
1.94
(0.13)
2.08
(0.13)
0.80
Androstenedione (pg/ml) 595.42
(19.27)
465.24
(21.00)
459.20
(18.97)
443.01
(19.35)
<0.001
Bioavailable testosterone (pg/ml) 10.60
(0.40)
8.39
(0.43)
9.01
(0.39)
6.4
(0.39)
<0.001
Free Testosterone (pg/ml) 4.85
(0.18)
3.83
(0.20)
4.12
(0.18)
3.08
(0.18)
<0.001
SHBG (nmol/l) 29.03
(1.65)
32.25
(1.80)
34.71
(1.63)
43.29
(1.66)
<0.001
*Adjusted for age, race, BMI, type of menopause;
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Table 5: EPAT Placebo Participants: Moderate or More Physical Activity
(hours/week) and Sex Hormone Levels (n=104)

Hormone (mean) Beta SE P-value*
Estradiol (pg/ml) -0.002 0.004 0.54
Estrone (pg/ml) -0.002 0.005 0.60
Bioavailable Estradiol (pg/ml) -0.10 0.06 0.09
Free Estradiol (pg/ml) -0.004 0.002 0.09
Testosterone (ng/dl) -0.31 0.11 0.006
DHEA (ng/ml) 0.03 0.01 0.07
Androstenedione (pg/ml) -1.91 2.46 0.44
Bioavailable testosterone (pg/ml) -0.21 0.05 <0.001
Free Testosterone (pg/ml) -0.10 0.02 <0.001
SHBG (nmol/l) 0.82 0.21 <0.001
*Adjusted for age, race, BMI, type of menopause;
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

20

Table 6: EPAT Placebo Participants: Moderate or More Physical Activity Tertiles
(hours/week) and Average Hormone Levels (n= 104)

Tertiles of Moderate or Vigorous
activities (hours/week)
Mean (SE)
Hormone (mean)
<1.7
(n=32)
1.7-3.8
(n=32)
> 3.8
(n=34)
P-value*
(linear trend)
Estradiol (pg/ml) 19.51
(1.03)
17.96
(1.03)
18.91
(1.03)
0.99
Estrone (pg/ml) 44.65
(1.03)
37.79
(1.04)
41.04
(1.04)
0.61
Bioavailable Estradiol (pg/ml) 15.04
(0.45)
13.45
(0.50)
13.62
(0.46)
0.09
Free Estradiol (pg/ml) 0.59
(0.02)
0.53
(0.02)
0.54
(0.02)
0.09
Testosterone (ng/dl) 23.23
(0.84)
23.62
(0.94)
20.62
(0.86)
0.004
DHEA (ng/ml) 1.83
(0.11)
2.29
(0.13)
2.22
(0.11)
0.11
Androstenedione (pg/ml) 491.34
(18.24)
503.05
(20.41)
499.20
(18.63)
0.88
Bioavailable testosterone (pg/ml) 9.87
(0.36)
9.32
(0.40)
7.44
(0.36)
<0.001
Free Testosterone (pg/ml) 4.51
(0.16)
4.26
(0.18)
3.40
(0.17)
<0.001
SHBG (nmol/l) 31.85
(1.55)
33.30
(1.73)
40.50
(1.58)
<0.001
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Table 6 summarizes the mean hormone and SHBG levels by weekly hours of
moderate or more intense physical activity categorized into tertiles. Overall, there is a
statistically significant increasing trend in serum SHBG level (p<0.001) and statistically
significant decreasing trends in levels of total testosterone (p=0.004), bioavailable
testosterone (p<0.001) and free testosterone (p<0.001) with increasing tertiles of time per
week spent in moderate or higher activity.
The linear relationships between the hormone and SHBG levels with the total
weekly hours spent in vigorous physical activity (activities rated as >6 MET) are
summarized in Table 7. Only 36% of the participants in the placebo group reported some
time spent in vigorous physical activity and only 9% reported more than one hour of
21
weekly vigorous physical activity. DHEA level (p=0.002) shows a statistically significant
positive association with time spent in vigorous physical activity, while time spent in
vigorous physical activity was inversely associated with serum levels of total testosterone
(p=008), bioavailable testosterone (p<0.001) and free testosterone (p<0.001).
Table 8 summarizes the results of analyses by three categorical levels of time
spent in vigorous physical activity: 0 hours in vigorous physical activity per week, more
than 0 to 0.3 (median) hours per week and more than 0.3 hours per week. SHBG levels
show an overall increasing trend with increasing categories of vigorous physical activity
in hours per week (p=0.04). All testosterone measures (total, bioavailable and free)
showed statistically significant decreasing trends (p<0.001 for all) with time spent in
vigorous physical activity.

Table 7: EPAT Placebo Participants: Vigorous Physical Activity (hours/week) and
Sex Hormone Levels (n=104)

Hormone (mean) Beta SE P-value*
Estradiol (pg/ml) -0.005 0.02 0.75
Estrone (pg/ml) -0.003 0.02 0.88
Bioavailable Estradiol (pg/ml) -0.25 0.23 0.27
Free Estradiol (pg/ml) -0.01 0.009 0.27
Testosterone (ng/dl) -1.13 0.42 0.008
DHEA (ng/ml) 0.18 0.06 0.002
Androstenedione (pg/ml) -9.39 9.07 0.30
Bioavailable testosterone (pg/ml) -0.66 0.18 <0.001
Free Testosterone (pg/ml) -0.30 0.08 <0.001
SHBG (nmol/l) 0.96 0.79 0.22
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

22
Table 8: EPAT Placebo Participants: Vigorous Physical Activity Categories
(hours/week) and Sex Hormone Levels (n= 104)

Hormone (mean) Categories of Vigorous
Physical activity
Mean (SE)
P-value*
(linear trend)
No Vig. PA
(n=71)
>0-0.3 (n=17) > 0.3 (n=16)
Estradiol (pg/ml) 19.09
(1.02)
17.63
(1.04)
19.98
(1.04)
0.05
Estrone (pg/ml) 41.71
(1.03)
39.40
(1.04)
40.06
(1.05)
0.62
Bioavailable Estradiol (pg/ml) 14.50
(0.37)
12.99
(0.55)
14.44
(0.61)
0.45
Free Estradiol (pg/ml) 0.57
(0.01)
0.51
(0.02)
0.57
(0.02)
0.45
Testosterone (ng/dl) 22.14
(0.71)
23.62
(1.06)
18.47
(1.17)
<0.001
DHEA (ng/ml) 2.04
(0.09)
1.87
(0.14)
2.47
(0.16)
0.001
Androstenedione (pg/ml) 509.12
(15.05)
461.43
(22.55)
473.66
(24.90)
0.36
Bioavailable testosterone (pg/ml) 9.19
(0.30)
8.59
(0.45)
6.64
(0.50)
<0.001
Free Testosterone (pg/ml) 4.20
(0.14)
3.92
(0.20)
3.03
(0.23)
<0.001
SHBG (nmol/l) 32.95
(1.28)
39.40
(1.92)
38.72
(2.12)
0.04
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Associations of Physical Activity with Exogenous Hormone Levels
(Estradiol-Treated Group)
The available sample size for this analysis was 90 women. Table 9 summarizes
the results of continuous analysis of the relationship between the total weekly MET
expenditure (Total-MET) and hormone and SHBG levels in the estradiol-treated group.
None of the hormones or SHBG was significantly associated with total weekly energy
expenditure after adjusting for age at randomization, race, BMI and type of menopause.

23

Table 9: EPAT Estradiol participants: Total-MET-hours and Sex Hormone levels
(n=90)

Hormone (mean) Beta** SE P-value*
Estradiol (pg/ml) -15.56 10.51 0.14
Estrone (pg/ml) -44.21 64.70 0.49
Bioavailable Estradiol (pg/ml) -8.98 6.29 0.15
Free Estradiol (pg/ml) -0.35 0.25 0.15
Testosterone (ng/dl) -2.88 4.37 0.51
DHEA (ng/ml) -0.11 0.32 0.73
Androstenedione (pg/ml) -31.40 76.65 0.68
Bioavailable testosterone (pg/ml) 0.35 1.43 0.81
Free Testosterone (pg/ml) 0.14 0.65 0.83
SHBG (nmol/l) -11.09 9.12 0.22
*Adjusted for age, race, BMI, type of menopause; ** Reported per 100 MET
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Table 10: EPAT Estradiol Participants: Total-MET-hours Quartiles and Sex
Hormone Levels (n=90)

Hormone (mean) Total MET-hours Quartiles
Mean (SE)
P-value*
(linear trend)
<232
(n=23)
232-237
(n=22)
238-244
(n=22)
>244
(n=23)

Estradiol (pg/ml) 68.94
(4.5)
70.66
(4.4)
65.27
(4.4)
63.02
(4.2)
0.04
Estrone (pg/ml) 309.29
(27.7)
321.82
(26.7)
288.44
(27.1)
301.52
(26.7)
0.40
Bioavailable Estradiol (pg/ml) 42.44
(2.7)
46.61
(2.6)
41.78
(2.6)
39.65
(2.5)
0.06
Free Estradiol (pg/ml) 1.67
(0.1)
1.83
(0.1)
1.64
(0.1)
1.56
(0.1)
0.06
Testosterone (ng/dl) 18.27
(1.9)
21.93
(1.8)
17.41
(1.8)
17.78
(1.7)
0.09
DHEA (ng/ml) 2.01
(0.1)
1.79
(0.1)
1.71
(0.1)
1.97
(0.1)
0.86
Androstenedione (pg/ml) 488.56
(32.8)
498.02
(31.6)
460.13
(32.1)
510.81
(30.4)
0.76
Bioavailable testosterone (pg/ml) 5.43
(0.6)
7.09
(0.6)
6.10
(0.6)
5.83
(0.6)
0.60
Free Testosterone (pg/ml) 2.47
(0.3)
3.23
(0.3)
2.78
(0.3)
2.65
(0.3)
0.59
SHBG (nmol/l) 54.1
(3.9)
45.35
(3.7)
45.29
(3.8)
49.13
(3.6)
0.15
*Adjusted for age, race, BMI, type of menopause;
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

We then analyzed the mean hormone levels by quartiles of total MET-hours of
energy expenditure (Table 10). Total estradiol level decreased with increasing quartiles of
24
energy expenditure (p=0.04). There was evidence of decreasing trends in bioavailable
estadiol and free estradiol levels, although these associations were only marginally
significant (p=0.06 for both). Associations of total MET quartiles with other hormones
were consistent with the continuous analysis.
Table 11 summarizes the linear relationship between the total weekly hours spent
in moderate or more physical activity (activities rated as 3 or more METs) and hormone
and SHBG levels. Total estradiol (p=0.001), bioavailable estradiol (p=0.01) and free
estradiol (p=0.01) levels were inversely associated with time spent in moderate or higher
physical activity. Androgen levels were not significantly associated with hours spent in
moderate or higher activities.

Table 11: EPAT Estradiol participants: Moderate or More Physical Activity
(hours/week) and Sex Hormone Levels (n=90)

Hormone (mean) Beta SE P-value*
Estradiol (pg/ml) -1.38 0.39 <0.001
Estrone (pg/ml) -4.02 2.45 0.10
Bioavailable Estradiol (pg/ml) -0.80 0.28 <0.001
Free Estradiol (pg/ml) -0.03 0.009 <0.001
Testosterone (ng/dl) -0.04 0.17 0.81
DHEA (ng/ml) 0.01 0.01 0.33
Androstenedione (pg/ml) 1.50 2.92 0.61
Bioavailable testosterone (pg/ml) -0.006 0.05 0.92
Free Testosterone (pg/ml) -0.002 0.02 0.92
SHBG (nmol/l) 0.10 0.34 0.80
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Table 12 summarizes the mean levels in hormone and SHBG levels with tertiles
of hours spent in moderate or more physical activity. Consistent with the continuous
analysis, estradiol (p=0.03), bioavailable estradiol (p=0.05) and free estradiol (p=0.05)
levels showed decreasing trends with increasing tertiles of moderate physical activity.
Serum DHEA (p=0.001) and androstenedione (p=0.005) levels increased with increasing
tertiles of time spent in moderate or higher activity.
25
Table 12: EPAT Estradiol Participants: Moderate or More Physical Activity
Tertiles (hours/week) and Average Hormone Levels (n=90)

Hormone (mean) Tertiles of Moderate or Vigorous activities
(hours/week)
Mean (SE)
P-value*
(linear trend)
<2.20
(n=29)
2.20-5.10
(n=30)
> 5.10
(n=29)

Estradiol (pg/ml) 73.16
(4.3)
64.71
(4.1)
64.17
(4.0)
0.03
Estrone (pg/ml) 298.82
(26.9)
317.62
(25.9)
301.02
(25.0)
0.88
Bioavailable Estradiol (pg/ml) 45.99
(2.6)
41.51
(2.5)
41.11
(2.4)
0.05
Free Estradiol (pg/ml) 1.81
(0.1)
1.63
(0.1)
1.62
(0.1)
0.05
Testosterone (ng/dl) 20.13
(1.8)
15.91
(1.7)
20.74
(1.7)
0.18
DHEA (ng/ml) 2.00
(0.1)
1.49
(0.1)
2.15
(0.1)
0.001
Androstenedione (pg/ml) 508.91
(30.9)
423.62
(29.7)
547.27
(28.7)
0.005
Bioavailable testosterone (pg/ml) 6.48
(0.6)
5.62
(0.6)
6.41
(0.6)
0.67
Free Testosterone (pg/ml) 2.95
(0.3)
2.56
(0.3)
2.92
(0.3)
0.66
SHBG (nmol/l) 47.44
(3.8)
47.43
(3.6)
49.50
(3.5)
0.33
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

Table 13: EPAT Estradiol participants: Vigorous Physical Activity (hours/week)
and Sex Hormone Levels (n=90)

Hormone (mean) Beta SE p-value*
Estradiol (pg/ml) -2.83 1.05 0.007
Estrone (pg/ml) -17.00 6.51 0.01
Bioavailable Estradiol (pg/ml) -2.41 0.63 <0.001
Free Estradiol (pg/ml) -0.09 0.02 <0.001
Testosterone (ng/dl) 0.76 0.45 0.09
DHEA (ng/ml) 0.14 0.03 <0.001
Androstenedione (pg/ml) 11.45 7.77 0.14
Bioavailable testosterone (pg/ml) 0.07 0.15 0.61
Free Testosterone (pg/ml) 0.03 0.07 0.64
SHBG (nmol/l) 0.33 0.92 0.72
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone

The linear relationship between the hormone levels and SHBG levels with the
total weekly hours spent in vigorous physical activity (activities rated as >6 MET) are
summarized in Table 13. DHEA levels show a statistically significant positive association
26
ith time spent in vigorous physical activity (p<0.001). Time spent in vigorous physical
activity was inversely associated with estradiol (p=0.007), estrone (p=0.01), bioavailable
estradiol (p<0.001) and free estradiol (p<0.001).
Table 14 summarizes the results of analyses by three categories of time spent in
vigorous physical activity per week: 0 hours, more than 0 to 0.8 (median) hours and more
than 0.8 hours. Consistent with the continuous analysis, all estrogen measures [estradiol
(p=0.001), estrone (p=0.003), bioavailable estradiol (p<0.001) and free estradiol
(p<0.001)] show an overall deceasing trend with increasing categories of vigorous
physical activity, while mean DHEA (p<0.001) and androstenedione (p=0.01) levels
show a statistically significant increase with time spent in vigorous physical activity.

Table 14: EPAT Estradiol Participants: Vigorous Physical Activity Tertiles
(hours/week) and Average Hormone Levels (n=90)

Hormone (mean) Categories of Vigorous
Physical activity
Mean (SE)
P-value*
(linear trend)
No Vig. PA
(n=50)
0-0.83 (n=22) > 0.83 (n=18)
Estradiol (pg/ml) 67.73
(3.8)
67.26
(4.3)
57.21
(4.6)
0.001
Estrone (pg/ml) 319.50
(23.1)
289.37
(26.6)
254.01
(28.5)
0.003
Bioavailable Estradiol (pg/ml) 43.23
(2.2)
43.77
(2.6)
34.52
(2.7)
<0.001
Free Estradiol (pg/ml) 1.70
(0.1)
1.72
(0.1)
1.36
(0.1)
<0.001
Testosterone (ng/dl) 18.32
(1.6)
19.55
(1.8)
20.66
(1.9)
0.17
DHEA (ng/ml) 1.80
(0.1)
1.91
(0.1)
2.23
(0.1)
<0.001
Androstenedione (pg/ml) 480.19
(27.5)
504.09
(31.6)
548.62
(33.8)
0.01
Bioavailable testosterone (pg/ml) 5.91
(0.5)
6.84
(0.6)
6.19
(0.6)
0.87
Free Testosterone (pg/ml) 2.69
(0.2)
3.11
(0.3)
2.82
(0.3)
0.88
SHBG (nmol/l) 48.85
(3.3)
44.35
(3.8)
51.34
(4.0)
0.24
*Adjusted for age, race, BMI, type of menopause
SHBG: Sex hormone-binding globulin; DHEA: Dehydroepiandrosterone
27
DISCUSSION

In this cohort of post-menopausal women participating in a randomized
clinical trial, physical activity levels were associated with sex steroid hormone levels in
both placebo-treated and estradiol-treated women.
Associations in Untreated Women
In the sample of placebo-treated postmenopausal women, total energy expenditure
was inversely associated with circulating concentrations of total testosterone, bioavailable
testosterone, free testosterone, androstenedione, bioavailable estradiol and free estradiol
after adjusting for age at randomization, race, BMI and type of menopause. SHBG was
positively associated with total energy expenditure. These associations were evident in
both continuous and categorical analysis by quartiles of physical activity. Total hours
spent in moderate or more physical activity was negatively associated with serum levels
of total testosterone, bioavailable testosterone and free testosterone in continuous and
categorical analysis. Similarly, time spent in vigorous physical activity was inversely
associated with serum levels of testosterone, bioavailable testosterone and free
testosterone. Time spent in moderate and vigorous activity was positively associated with
SHBG. The majority of our findings are consistent with the results from other studies
(Cauley, Gutai et al. 1989; McTiernan, Tworoger et al. 2004; McTiernan, Tworoger et al.
2004; Chan, Dowsett et al. 2007).
The first randomized clinical trial to address the relationship of physical activity
and endogenous sex hormone levels in postmenopausal women investigated the effects of
a moderate-intensity physical activity intervention, comprising outdoor walking,
28
treadmill, and stationary bicycling, on endogenous hormones, compared with a control
group of stretching exercises (McTiernan, Ulrich et al. 1999; McTiernan, Tworoger et al.
2004; McTiernan, Tworoger et al. 2004). After 3 months, exercisers (n=87) experienced
declines in estrone, estradiol, and free estradiol compared to (n=86) controls (P = 0.03,
0.07, and 0.02, respectively). At 12 months, the direction of effect remained the same,
although the differences were no longer statistically significant (McTiernan, Tworoger et
al. 2004). Also at 12 months, women in the exercise intervention group showed
reductions in total testosterone and free testosterone, compared to the controls. However,
the associations were statistically significant only in women who lost more than 2% of
body fat (p=0.02). The study also noted an increase in SHBG level in the moderate
exercise intervention group at 3 months (p=0.02), which however, was not associated at
12 months (p=0.10).
In a recent cross-sectional population-based study, 2082 postmenopausal women
not using exogenous hormones demonstrated an inverse relationship between usual
physical activity levels and circulating concentrations of estradiol (p=0.02) and
testosterone (p<0.001) (Chan, Dowsett et al. 2007). Consistent with our findings, they
also found a positive association with SHBG concentrations (p=0.03) (Chan, Dowsett et
al. 2007). Another cross-sectional study in 1989 found an inverse relationship between
estrone and estradiol levels and physical activity (Cauley, Gutai et al. 1989). Similarly, in
a longitudinal study that followed participants for 8 years, endurance-trained
postmenopausal women had lower estrogen, specifically estrone, levels compared to
sedentary women (Nelson, Sammel et al. 2008).
29
A reduction in female sex hormones with higher physical activity may impact
breast cancer risk in postmenopausal women. High endogenous estrogen levels have been
associated with elevated breast cancer risk in postmenopausal women (Bernstein and
Ross 1993). Cumulative exposure to estrogen is particularly relevant to breast cancer risk
and is determined by factors such as age at menarche, age at menopause, pregnancy, use
of oral contraception and lactation (Bernstein 2002). Physical activity itself has been
associated with reduced breast cancer risk. To date 33 original reports have investigated
the relationship of physical activity and breast cancer risk; of the 10 of these studies
reporting results separately for postmenopausal women, 9 found an inverse association
(Monninkhof, Peeters et al. 2007). The mechanisms by which physical activity exerts its
beneficial effects are still being explored and debated (Stevenson, Davy et al. 1995;
McTiernan 2008).
The inverse association of physical activity with serum SHBG level may also be
associated with reduced breast cancer risk. SHBG binds estradiol, reducing the
bioavailability of this hormone. Postmenopausal women with low serum SHBG have
increased risk of breast cancer (Chan, Dowsett et al. 2007).
The significant negative associations of physical activity with androgens in the
placebo group may imply a beneficial effect on cardiovascular disease. A large body of
observational evidence suggests a positive association between menopause and
cardiovascular disease risk (Witteman, Grobbee et al. 1989; Hill 1996). Some studies
suggest that an increase in androgens relative to estrogen in healthy postmenopausal
women is associated with an unfavorable cardiovascular risk profile (Rexrode, Manson et
al. 2003). In EPAT androgen levels were affected by as little as 2 hours of weekly
30
moderate or higher physical activity. This association can also be explained partially by
the inverse association of physical activity with serum SHBG level in this population.
SHBG is known to regulate the concentration of bioavailable estrogen and androgens and
has a greater affinity for androgens, especially testosterone (Rosner 1991). Therefore,
physical activity may benefit postmenopausal women not taking any hormonal
intervention by contributing to a reduction in an androgen excess.
Associations in Estradiol-Treated Women
In our cohort of estradiol-treated women, total energy expenditure was not
associated with any significant changes in the hormone or SHBG level. However, in a
categorical analysis estradiol levels decreased with increasing quartiles of energy
expenditure. Estradiol, bioavailable estradiol and free estradiol were inversely associated
with total hours spent in moderate or more physical activity. There was also a statistically
significant increasing trend in serum DHEA and androstenedione level with increasing
tertiles of weekly hours spent in moderate or higher physical activity. Similarly, time
spent in vigorous activity was inversely associated with estradiol, bioavailable estradiol
and free estradiol levels and positively associated with DHEA and androstenedione
levels.
To date, no study has reported on the relationship of physical activity and sex
hormone levels in postmenopausal women taking estradiol or any exogenous female
steroid hormones. The significant decreasing trend of female sex hormones with
increasing physical activity may be beneficial for the added risk of breast cancer
associated with HT in some studies. Among women participating in observational studies
who are current or recent users of estrogen therapy, risk of breast cancer was modestly
31
elevated (Bernstein 2006). In a large population-based case-control study in Germany,
including 3,464 breast cancer cases aged 50-74 at diagnosis and 6,657 population-based
and frequency-matched controls, the risk of breast cancer was slightly higher in women
taking estrogens only therapy (Flesch-Janys, Slanger et al. 2008). Results from the WHI
trial also suggest that those receiving conjugated equine estrogens (CEEs) showed a
lower breast cancer risk compared to the placebo group (Prentice, Chlebowski et al.
2008).
Strengths and Limitations
There are several strengths of the current analysis from EPAT. Having two groups
in our cohort, one taking estradiol and one without any exogenous hormones, we were
able to observe the influence of physical activity in postmenopausal women at both
physiologic and pharmacologic levels of hormones. A comprehensive panel of hormones
was measured, enabling us to investigate a large part of the hormonal milieu in these
postmenopausal women. Moreover, repeated measures hormone and physical activity
allowed us to obtain more representative measure of a woman’s average hormone and
physical activity profile over time than a single measurement. This was especially
applicable for the physical activity measurements since they were very stable over time.
The major limitation of the study is the post hoc nature of the analysis, in that
EPAT was not designed to address our hypothesis. The relatively small sample size may
have compromised our power to achieve statistical significance in some associations. For
example, in the 90 participants receiving HT there were marginally significant decreasing
trends in bioavailable and free estradiol levels with increasing quartiles of total energy
expenditure. In addition, the small sample reduced the statistical power to test
32
interactions (all p-values >0.10), including possible interactions with BMI and race. The
cohort had a very high BMI with a mean of 29 (±5.4) kg/m
2
. As adipose tissue is a major
source of estrogens in postmenopausal women, the amount of physical activity might not
have been sufficient to reduce the hormones to a great extent in this relatively overweight
population of postmenopausal women. This may explain the lack of inverse association
with time spent in moderate or vigorous activity in the placebo group, where only 36% of
the participants reported some time spent in vigorous physical activity and only 9%
reported more than one hour of weekly vigorous physical activity.
Conclusion
This study adds to the limited evidence on the relationship between physical
activity and sex hormone levels in postmenopausal women. Given the fact that this was a
post hoc cross-sectional analysis, we cannot infer a causal relationship between physical
activity and hormone levels. Despite that, the implications of the results warrant further
exploration. To our knowledge, no study has examined the relationship of physical
activity with such a large panel of hormones at both physiologic and pharmacologic
levels. Further longitudinal interventional studies with a larger sample size might help to
further understand the uncertain, largely unexplored, yet intriguing, relationship between
physical activity and sex hormone levels in this population.

33
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Abstract (if available)

Abstract The crucial yet complex relationships between physical activity and physiologic and pharmacologic sex hormone levels in postmenopausal women have not been investigated sufficiently. In this post hoc cohort analysis of data from Estrogen in the Prevention of Atherosclerosis Trial, physical activity was tested for an association with sex hormone levels, assessed longitudinally over two years in 194 postmenopausal women, 90 randomized to 17β-estradiol (e2) and 104 to placebo. Physical activity (total energy expenditure and weekly hours spent in moderate or more physical activity) was associated with decreased serum levels of estradiol in both groups [Placebo: (p=0.02)

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Asset Metadata

Creator Choudhury, Farzana (author)

Core Title Physical activity and sex hormone levels in postmenopausal women

School Keck School of Medicine

Degree Master of Science

Degree Program Applied Biostatistics

Publication Date 11/17/2008

Defense Date 10/20/2008

Publisher University of Southern California (original), University of Southern California. Libraries (digital)

Tag OAI-PMH Harvest,physical activity,postmenopausal women,sex hormones

Language English

Contributor Electronically uploaded by the author (provenance)

Advisor Mack, Wendy J. (committee chair), Azen, Stanley Paul (committee member), Bernstein, Leslie (committee member), Hodis, Howard Neil (committee member)

Creator Email fchoudhu@usc.edu,neeparr@gmail.com

Permanent Link (DOI) https://doi.org/10.25549/usctheses-m1780

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Rights Choudhury, Farzana

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