Female Athlete Triad Clinical Trial
Official title:
Physiological and Behavioural Aspects of Insufficient Energy Availability in Females Endurance Athletes - Impact on Metabolism, Recovery and Health
The purpose of this project is to study the impact of energy availability (EA) on female
endurance athletes and the adaptive consequences of insufficient EA with a special reference
to the impact on energy metabolism, reproductive-, vascular- and bone health as well as
exercise capacity, neuromuscular performance, ability to recover from intense exercise and
the genetic interaction.
Specific aims:
- To investigate the effects of EA on energy metabolism
- To investigate the effects of EA on reproductive, vascular- and bone health, and
endocrine functions
- To investigate the effects of EA on exercise capacity
- To investigate the effects of EA on neuromuscular performance
- To investigate the effects of EA on recovery after exercise
- To identify potential dietary factors influencing EA, exercise capacity and
neuromuscular performance
- To identify potential exercise factors influencing EA, exercise capacity and
neuromuscular performance
- To identify potential psychological, behavoural and motivational aspects associated with
insufficient EA
The investigators hypothesize that female athletes with insufficient energy availability have
attenuated energy metabolism, reduced BMD, impaired endothelial function and decreased
ability to recover between exercise bouts compared to matched controls with sufficient energy
and nutrient intake. The investigators also hypothesize that EA influences exercise capacity
and neuromuscular performance.
The study population was recruited through the Danish and Swedish sport federations in
weight-bearing endurance sports, and in local competitive endurance sports clubs, through
flyers, webpage announcements and mailings. Interested athletes received the study protocol
and a self-administered questionnaire, assessing age, sport, training regime, and
self-assessed menstrual function. Athletes returning the questionnaire and who were
interested in further participation were contacted by the research group. Before inclusion,
all subjects were informed orally, and in writing, of all study procedures and signed an
informed consent form. Subjects included and defined as elite endurance athletes were
athletes at national team levels or competitive endurance athletes from regional sports
clubs, 18-38 years of age, training a minimum of five times per week. Exclusion criteria
included clinically verified menstrual dysfunction other than oligomenorrhea/functional
hypothalamic amenorrhea; pregnancy; chronic illness; smoking; use of forms of contraceptives
other than oral, e.g., hormonal coil/patches; inability or being unwilling to stop hormonal
contraceptives for at least 6 weeks prior to investigation; or injuries preventing the
athlete from training ≥ 2 weeks.
Data collection was performed on two consecutive days followed by a 7-day registration period
in the athletes' normal environment. The first day comprised of examinations involving bone
health, blood pressure and reproductive function. The second day included examinations of
energy metabolism, aerobic capacity, as well as assessment of eating disorders. The subjects
were instructed not to exercise for more than 30 min at low or moderate intensity on the day
before, and on the first day of examination, and to arrive at the clinic in a fasted (from
midnight) and rested state.
PROTOCOL OVERVIEW:
DAY 1: Start at 7:45 A.M.
1. Pragnancy test
2. Blood sampling
3. DXA scan
4. Blood pressure
5. Meal
6. Gynecological examination
DAY 2: Start at 7 A.M.
1. RMR
2. Blood sampling
3. Work efficiency
4. Meal
5. EDE interview
6. Incremental exercise test
7. EDI-questionnaire
7 DAY DIETARY AND ACTIVITY RECORD
The timing of the examination and registration period was planned individually for each
subject in order to choose a period reflecting their habitual food habits and exercise
regimes. Dietary intake and training intensity were recorded by the subjects for seven
consecutive days to assess current EA. Energy intake was calculated from a prospective
weighed (Exido 246030 Kitchen Scale, Gothenburg, Sweden) food record, using a nutrient
analysis program, Dankost 2000 (Dankost, Copenhagen, Denmark) for Danish food items, and
Dietist XP (Kost och Näringsdata AB, Bromma, Sweden) for Swedish food items. Subjects were
given in-depth verbal and written instructions and a demonstration of how food and beverages
should be weighed and registered. Subjects were instructed to maintain their normal food
habits and eating patterns. Before entering data in the nutrient analysis program the same
dietician reviewed all completed diet records and asked for supplementary information if
needed, and the mean daily energy intake from the 7-day record was used in the analysis. In
order to identify subjects who provided nutritional data of poor validity, the Goldberg
cutoff was calculated using the equation described by Black (2000). Heart rate monitors
(Polar RS400®, Stockholm, Sweden) and training logs were used to assess exercise energy
expenditure. Subjects were instructed to maintain and to follow their normal training regime.
They were, furthermore, instructed to describe each session in as much detail as possible,
regarding type, duration, and intensity of exercise and to wear the heart rate monitor at all
training sessions (except swimming) and during cycling (training as well as transportation).
Eating behavior was assessed using the Eating Disorder Inventory (EDI-3), a questionnaire
assessing behavior and attitudes related to DE behavior, as well as to overt ED. Subjects
were categorized as having DE behavior when the EDI risk subscale score for Drive for
Thinness (DT) was ≥ 14, and/or a Body Dissatisfaction (BD) risk score ≥ 19 and without the
presence of DSM-IV diagnosed ED. The Eating Disorder Examination (EDE-16) was used to
determine whether subjects met the criteria for ED, according DSM-IVcriteria for anorexia
nervosa, bulimia nervosa, and ED not otherwise specified (EDNOS). All interviews were
performed by the same EDE-certified member of the research team.
Subjects were transported by car to the clinic on the morning of the second test day in order
to minimize physical activity prior to the measurement. RMR was assessed between 7:00 and
8:30 h, after an overnight fast, using a ventilated open hood system (Oxycon Pro 4, Jeager,
Germany). The system was automatically calibrated before each test, and again weekly, by an
alcohol burning test with coefficients of variability (CV) of 0.7% for O2, 1.1% for CO2,
0.8%for the respiratory exchange ratio, and 2% for EE. After voiding, subjects laid down for
15 min before measurements of oxygen consumption (VO2) and carbon dioxide production (VCO2)
over 35 min (the equation is defined under Calculations). Work efficiency was assessed by a
standardized test protocol in the fasted state, initiated by the subject seated on the
bicycle ergometer (Monark 939E, Monark Exercise AB, Vansbro, Sweden) for 6 min, followed by
cycling for 6 min at 0W, 50W and 100W, respectively. An air-tight mask covering the mouth and
nose was used in order to measure respiratory gas exchange (the equations for calculations
are defined under Calculations). In order to calculate daily total EE, HR monitors (Polar
RS400®) were used to assess EE during bicycle transportation, while actigraphy (ActiGraph
GT3X®, Pensacola, FL, USA) and the data analysis software ActiLife 5 (ActiGraph) were used
for assessment of NEAT. Subjects were instructed to wear an accelerometer on the wrist during
sleep, and on the hip from getting up in the morning until bedtime, and only to take it off
during showering, swimming, bicycle transportation, and training.
Two hours after a standardized breakfast, an incremental exercise test on the bicycle
ergometer was performed, initiated by cycling for 6 min at 50W, followed by an increase in
workload of 12-14 W/min until exhaustion. An air-tight mask covering the mouth and nose was
used in order to measure VO2peak and respiratory exchange ratio, and HR (Polar RS400®) was
measured.
Subjects using hormonal contraceptives were requested to stop for a minimum of 6 weeks prior
to examination in order to secure a sufficientwashout period for exogenous estrogen and
progesterone. Subjects not recovering their menstrual bleeding within the 6 weeks were
contacted monthly by the research team during a follow-up period of a minimum of 3 months
before gynecological assessment. Menstruating athletes were examined in the early follicular
phase, on the third to fifth day of menstruation. A pregnancy test was performed on arrival
at the hospital, and menstrual function was examined by an experienced gynecologist who
performed a transvaginal ultrasound examination (Ultrasound Scanner, Class 1 type B, B-K
Medical REF TYPE 2202, Bedfordshire, UK). The maximum number of ovarian follicles present in
a single plane was counted, and total volume was assessed. Sex hormone status [estrogen,
progesterone, LH, follicle stimulating hormone (FSH), sexual hormone binding globulin (SHBG),
prolactin, dehydroepiandrosteron sulfate (DHEA-S), androstendion, and total testosterone] and
anamnestic assessment, e.g., age of menarche, previous menstrual irregularities, use of
hormonal contraceptives, and number of menstrual cycles during the last year, were recorded
using the low EAin females questionnaire (LEAF-Q) (Melin et al., 2014). Subjects were then
classified with eumenorrhea (menstrual cycles of 28 days ± 7 days and sex hormones within the
normal range); oligomenorrhea (menstrual cycles > 35 days where other causes than
hypothalamic suppression had been ruled out); FHA (either primary: no menarche after 15 years
of age, or secondary: absence of at least three consecutive menstrual cycles where other
causes than hypothalamic suppression had been ruled out) (Nattiv et al., 2007); other MDs
(anatomic defects, hyperprolactinemia or other dysfunctional ovarian conditions); PCOS
involving at least two of the following: (a) enlarged ovaries with a volume greater than 10
mL and/or ≥ one ovary demonstrating ≥ 12 follicles in one plane; (b) irregular or absence of
bleeding; and (c) elevated androgen level, or otherwise androgen stigmatized.
Body weight was measured with an accuracy of 0.01 kg in underwear on an electronic scale
(Lindeltronic 8000, Samhall Lavi AB, Kristianstad, Sweden). Height was measured without shoes
and standing with legs together against a wall, using a fixed stadiometer (Hultafors AB,
Hultafors, Sweden). After resting in a supine position for 7 min, HR and BP were measured
three times (the mean was used) using an electronic sphygmomanometer (Microlife BP A100,
Widnau, Switzerland). Hypotension was defined as a systolic BP < 90 mmHg and/or diastolic BP
< 60 mmHg. Dual-energy X-ray absorptiometry (DXA) (Hologic, Model Discovery 2009, Hologic
Inc.,Waltham, MA, USA) was used to determine fat-free, fat, and bone mass, respectively. BMD
was determined for whole body, lumbar spine (L1-L4) and hip. All measurements and scans were
performed in the fasted and resting state between 7:30 and 9:00 h, and were assessed by the
same technician, and performed on the same scanner. Calibration of the Hologic Discovery 2009
was performed weekly, using a phantom provided by the manufacturer. Subjects were classified
as having normal BMD: Z-scores > −1 in all measured sites, low BMD: Z-score −1 to −2 in at
least one site, and osteoporosis: Z-score < −2 in at least one site together with minimum one
secondary risk factor such as low EA, ED and oligomenorrhea/FHA.
Blood samples were drawn following an overnight fast from an antecubital vein on the first as
well as on the second day, between 8:30 and 8:50 h, in a rested state.
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