Resistance exercise provides a potent stimulus for muscular adaptations. This process is mediated, at least in part, by an increase in the levels of anabolic hormones including testosterone, growth hormone (GH), dehydroepiandrosterone sulfate (DHEAS), and insulin-like growth factor I (IGF-I) (4,9,17,23). However, resistance exercise-induced anabolic hormone changes are different between men and women. Previous studies reported that a high-intensity and high-volume resistance exercise program with short rest periods induced an increase in GH after exercise in both men and women, whereas testosterone increased only in men (12,22). In addition, the resting serum level of testosterone in women is about 10 times lower than that in men. Riechman et al. (29) showed that acute resistance exercise induces increased DHEAS, a peripheral precursor of testosterone, in both men and women. These results indicated that there are sex differences in basal anabolic hormone levels and responses to exercise.
The hormonal responses to exercise are also modified by ovarian systems in women. It is known that menstrual disorders including oligomenorrhea and amenorrhea are functional disorders characterized by an altered gonadotropin-releasing hormone pulsatility, loss of pulsatile secretion of gonadotropins (FSH and LH), and, in turn, altered ovarian steroidogenesis (26). Waters et al. (31) reported that amenorrheic athletes showed significantly lower (four- to fivefold) GH responses to 50-min submaximal exercise (70% V˙O2max) compared with eumenorrheic athletes. Yahiro et al. (32) reported that serum testosterone levels increased in eumenorrheic runners, but not in amenorrheic runners, after an acute treadmill exercise. However, there is no information about anabolic hormonal responses to resistance exercise in women with menstrual disturbances.
To more accurately examine exercise-induced hormone changes in women, it is important to control for the hormonal fluctuations that occur throughout the menstrual cycle. Previous studies reported that GH concentration in the periovulatory phase was higher than that in the early follicular phase (8,28). Kraemer et al. (19) demonstrated that low-volume resistance exercise induced greater increases in estradiol, GH, and androstenedione during the midluteal phase compared with those during the early follicular phase, although they did not compare the responses within the same individuals. These data suggest that anabolic hormone responses to resistance exercise may be influenced by the phase of the menstrual cycle (e.g., levels of ovarian hormones). Moreover, it is useful to examine the changes of anabolic hormones in response to resistance exercise throughout the menstrual cycle within the same individuals.
We hypothesized that changes in ovarian and anabolic hormones in response to an acute resistance exercise are influenced by menstrual cycle phase and state. In the present study, we examined resistance exercise-induced ovarian and anabolic hormone responses in each menstrual phase (i.e., follicular and luteal phases) in different menstrual states (i.e., eumenorrheic young women and young women with menstrual disorders).
Eight eumenorrheic women (EM) and eight women with menstrual disorders including oligomenorrhea and amenorrhea (OAM) were enrolled in this study. The study was conducted under institutional review board approval (of Tsukuba University of Technology and University of Tsukuba), and written informed consent was obtained from all 16 subjects. All subjects were recreationally active women (not involved in any regular exercise or training at least 6 months before the start of the investigation) aged 18-30 yr with body mass index (BMI) of 18-22 kg·m−2. None of the subjects was parous, smoked, or used oral contraceptives. Subjects from the EM group had regular 25- to 38-d menstrual cycles as determined by questionnaires. We then monitored their menstrual cycles with basal body temperature and ovulation test kits (Do-test LH; ROHTO Pharmaceutical Co., Ltd., Osaka, Japan) for 2-3 months, which also aided in scheduling the experimental test for the respective phase of the menstrual cycle. The menstrual disturbance information in the OAM group (oligomenorrhea: periods at interval exceeding 6 wk; amenorrhea: no bleeding for the last 3 months) was provided by the subjects.
To confirm that each of the subjects did not have an eating disorder, the Eating Attitudes Test (EAT-26) (10) was performed. Body weight was measured with a digital scale (TBF-560; Tanita, Tokyo, Japan). Percent body fat was assessed using a multiple-frequency bioelectrical impendence analyzer (MLT-100; Sekisui Chemical, Tokyo, Japan). Subjects were instructed to maintain their normal food intake for 3 d before the exercise session, and the dietary records were inspected. Daily caloric intakes were calculated using software (Excel Eiyou-kun ver. 3.0; Yoshimura, Kenpakusya, Tokyo, Japan).
One-repetition maximum test.
Not <3 d before exercise session, all subjects were assessed for maximal strength using a concentric-only one-repetition maximum (1-RM) on devices for lat pull-down, leg curl, bench press, leg extension, and squat exercises. They were instructed on the proper lifting technique before testing. Subjects were allowed a brief light-resistance warm-up, then they were encouraged to meet their 1-RM within five trials of incrementally increasing resistance. All resistance exercises were performed on variable resistance gym equipment (Senoh Corp., Tokyo, Japan).
Experimental design and exercise protocols.
The EM group subjects participated in two series of exercise sessions, one during the early follicular phase (EF; days 4-7 of the menstrual cycle) and one during the midluteal phase (ML; 7-10 d after ovulation). The two series of exercise sessions were randomized. Subjects from the OAM group participated in a series of exercise session on an arbitrary day. The sessions were organized at the same time of the day (in the morning starting at 07:30 h) to minimize diurnal variations. In addition, subjects were asked to avoid any kind of strenuous exercise 3 d before the sessions and to refrain from eating after 21:00 h on the day before testing.
On arrival in the laboratory for all exercise sessions, subjects were seated quietly for approximately 10 min, and resting blood samples (Pre) were obtained. After a light-resistance warm-up, they performed five resistance exercises using the same equipment as those used for the 1-RM test: lat pull-downs, leg curls, bench presses, leg extensions, and squats. Resistance was established at 75%-80% of 1-RM. Subjects performed three sets of each exercise with 1-min rests between sets. Subjects were instructed to lift until concentric failure for every set and to use a spotter's assistance to complete 10 repetitions. To keep work uniform for each repetition, each lift was timed to metronome signals every 2 s. The time to complete three sessions of all five exercises was approximately 20 min (at 08:20 h, the subjects completed the exercise session). Subjects were then seated for a recovery period. Blood samples were obtained immediately after the end of the resistance exercise (P 0), after 30 min (P 30), and after 60 min (P 60) of recovery. Water intake was allowed ad libitum throughout the exercise protocols and recovery.
Whole blood lactate concentrations were determined via a lactate analyzer (Lactate Pro LT-1710; ARKRAY, Inc., Kyoto, Japan). The remainder of the blood was centrifuged at 3000g for 15 min at 4°C. All serum samples were then distributed to appropriate preservative-containing tubes and stored at −40°C until analysis. Serum concentrations of estradiol, progesterone, and testosterone were measured by radioimmunoassay, GH and cortisol were measured by chemiluminescence enzyme immunoassay, and IGF-I was measured by immunoradiometric assay (Mitsubishi Chemical Medience Corp., Ibaraki, Japan). Serum concentrations of DHEAS were measured by enzyme immunoassay using a commercially available kit (DRG Instruments GmbH, Marburg, Germany). The intra-assay coefficients of variation for the hormone assays were as follows: estradiol < 15%, progesterone < 15%, testosterone < 15%, GH < 15%, cortisol < 10%, IGF-I < 10%, and DHEAS = 5.1%, respectively.
Statistical analyses were performed using SPSS 13.0J for Windows (SPSS, Inc., Chicago, IL). Results are expressed as mean ± SEM. Student's t-test was used to assess comparability of physical, nutritional, and resting hormonal data in the menstrual phases (EF vs ML) and menstrual state (EM vs OAM). Two-way repeated-measures ANOVA were used to determine whether menstrual status (EF vs ML, EF vs OAM) influenced the dependent variables and whether this menstrual status effect differed over time after exercise (group × time interaction). The significance of differences between the values at Pre and in response to exercise was determined by the two-tailed multiple t-test with Dunnett correction. The integrated area under the curve (AUC) was determined after subtracting each subject's baseline (Pre) hormone concentrations, using a computer program, MOMENT (EXCEL), for moment analysis (30). To determine whether an AUC response within each group occurred, a Student's t-test was used to determine whether the AUC was greater or less than zero (19). Statistical significance was set at P < 0.05.
Physical, gynecological, nutritional, and maximal strength (1-RM) characteristics of the subjects from the EM and OAM groups are presented in Table 1. Physical and nutritional data in the EF phase were expressed as EM data in Table 1 because no differences were found between the EF and ML phase values (body weight = 53.5 ± 1.5 and 53.6 ± 1.5 kg, body fat = 25.4% ± 1.3% and 25.5% ± 0.9%, and daily caloric intake = 1779 ± 140 and 1950 ± 178 kcal, respectively). Height, body weight, body fat, BMI, age of menarche, daily caloric intake, and 1-RM resistance exercises did not differ between the EM and OAM groups. The mean age of the OAM group was significantly younger than that of the EM group.
Resting serum concentrations of estradiol, progesterone, GH, IGF-I, testosterone, DHEAS, and cortisol are presented in Table 2. Resting estradiol and progesterone concentrations in the ML phase (66.0-152.0 pg·mL−1 (mean = 104.0 ± 11.2 pg·mL−1) and 4.8-27.0 ng·mL−1 (mean = 13.0 ± 2.9 ng·mL−1), respectively) were significantly higher than in those in the EF phase (21.0-55.0 pg·mL−1 (mean = 38.6 ± 3.8 pg·mL−1) and 0.7-1.6 ng·mL−1 (mean = 1.1 ± 0.1 ng·mL−1) for estradiol and progesterone, respectively), and these concentrations did not differ between the OAM group (15.0-45.0 pg·mL−1 (mean = 28.1 ± 3.4 pg·mL−1) and 0.9-2.8 ng·mL−1 (mean = 1.4 ± 0.2 ng·mL−1) for estradiol and progesterone, respectively) and the EF phase of the EM group. For all subjects, resting testosterone concentrations were within the reference range (female = 0.06-0.86 ng·mL−1) (Mitsubishi Chemical Medience Corp.), but they were significantly higher in the OAM group compared with the EF phase in the EM group. There were no significant differences between the groups in the resting levels of the other hormones, GH, IGF-I, DHEAS, and cortisol.
Blood lactate concentrations at Pre, P 0, P 30, and P 60 were 0.9 ± 0.1, 6.8 ± 0.7, 3.5 ± 0.5, and 1.9 ± 0.2 mmol·L−1 in the EF phase; 1.3 ± 0.2, 6.5 ± 0.6, 3.1 ± 0.3, and 1.8 ± 0.2 mmol·L−1 in the ML phase; and 1.1 ± 0.2, 6.8 ± 0.8, 3.5 ± 0.5, and 1.7 ± 0.3 mmol·L−1 in the OAM group, respectively. The levels of blood lactate significantly increased at P 0 and P 30 compared with Pre, but there was no difference among the groups.
Changes in ovarian hormone (estradiol and progesterone) concentrations during the prolonged exercise test session are presented in Figure 1. Significant interactions (menstrual cycle phase: EF and ML, by time) were found for both estradiol and progesterone. Estradiol levels in the ML phase at P 0, and progesterone levels in the ML phase at P 0 and P 30 were significantly higher than their corresponding levels at Pre. Estradiol and progesterone concentrations in the EF phase and the OAM group showed no significant increase at P 0 and significant decrease at P 30 and P 60 from Pre.
Changes in GH, IGF-I, testosterone, DHEAS, and cortisol concentrations during the prolonged exercise test session and the corresponding AUC panels are presented in Figure 2. GH concentrations at P 0 in the EF and ML phases demonstrated significant increases from Pre, whereas that in the OAM group was not significant from Pre. The AUC for GH was significantly higher than zero in the ML phase. A slight, but significant increase in IGF-I response to resistance exercise is shown at P 0 in the OAM group. Testosterone concentrations showed no significant increase in response to the resistance exercise protocols in all groups, and significant decrease at P 60 in the EF and ML phases from Pre. However, the AUC for testosterone was significantly lower than zero in the OAM group. A significant interaction (menstrual status: EF and OAM, by time) was found for serum DHEAS. DHEAS concentrations showed no significant increase immediately after the resistance exercise in all the groups; however, there was a significant increase at P 60 in the EF phase, and a significant decrease at P 60 in the OAM group. No significant differences in cortisol change were found between groups. Serum cortisol concentrations showed no significant increase response to the resistance exercise protocols in all groups and significant decrease at P 0 and/or P 30 and P 60 from Pre for each group.
We investigated changes in ovarian and anabolic hormones after acute resistance exercise in the EF and ML phases of healthy women and women with menstrual disorders (OAM). Serum estradiol and progesterone in the ML phase increased after the exercise but did not change in the EF phase or in the OAM group. Serum GH increased after exercise in both the EF and ML phases. Women with a menstrual disturbance with low estradiol and progesterone levels exhibited an attenuated anabolic hormonal response to acute resistance exercise.
In the present study, anabolic hormonal responses to acute resistance exercise were different between menstrual phases. Consistent with a previous investigation (19), the total secretion (AUC) of GH was significantly increased in the ML phase, but not in the other groups, although serum GH levels were increased after the acute resistance exercise in both the EF and ML phases. Hornum et al. (14) also demonstrated that total secretion of GH after a high-intensity cycling exercise was greater in the periovulatory phase than the follicular phase, during which plasma levels of endogenous estradiol were relatively high (1112 ± 407 pmol·L−1) and low (272 ± 59 pmol·L−1), respectively. These findings indicate that menstrual cycle variations in circulating estradiol may affect exercise-induced GH secretion.
Interestingly, the level of serum GH in women with menstrual disorders was not changed by the resistance exercise in this study. A previous study reported that secretion of GH by a 50-min submaximal exercise bout (70% V˙O2max) was significantly lower in amenorrheic athletes (31). The neuroendocrine mechanisms of exercise-induced GH release are not fully understood; however, the secretion of GH is controlled by hypothalamic hormones. These hormones include GH-releasing hormone, which exerts positive feedback, and somatostatin, which exerts negative feedback, on GH secretion (11,15). Because central neuroregulation of the secretion of many pituitary hormones is disturbed in hypothalamic amenorrhea (1,31), it is possible that the attenuated GH secretion in response to acute resistance exercise in the OAM group was caused by a disturbance of the hypothalamic-pituitary function in hypothalamic amenorrhea.
In the current study, resting levels of estradiol and progesterone in the EF phase and in the OAM group were lower than those in the ML phase. Moreover, serum estradiol and progesterone in both groups did not change after the exercise but increased in the ML phase. On the other hand, changes in GH response to the exercise were different between the EF phase and the OAM group, although circulating ovarian hormone levels were similar in both groups. Previous studies suggested that estradiol stimulates GH secretion because hormone (estrogen) replacement therapy increased GH secretion in postmenopausal women and prepubertal girls with Turner syndrome (18,25). This effect of estrogen could be brought about by withdrawal of somatostatin's inhibitory tone, amplification of endogenous GH-releasing hormone release or its pituitary actions, and/or recruitment of other mechanisms that stimulate GH release (11). In addition, Kraemer et al. (20) and Kanaley et al. (18) demonstrated that GH responses to endurance exercise are higher in postmenopausal women receiving hormone replacement therapy than in those without treatment. Specific endocrine differences between eumenorrheic women and women with menstrual disorders are cyclic fluctuations of estrogen and progesterone controlled by the hypothalamic-pituitary-gonadal axis. In this study, the subjects in the OAM group were estrogen deficient for more than 6 wk, but the EM group had normal fluctuations of ovarian hormones throughout the menstrual cycle. The insufficient estrogen and progesterone feedback causes a disturbance of the hypothalamic-pituitary responses (6). Thus, differences in ovarian hormone secretion status, that is, differences in hypothalamic-pituitary function between women with menstrual disorders and eumenorrheic women may influence the GH response to acute resistance exercise.
It is suggested that many factors (e.g., sex, age, fitness level, nutritional status, exercise variables) seem to influence the hormonal responses to resistance exercise (4,23). In the present study, we controlled for most of these characteristics as well as menstrual cycle phase and status of the subjects. Resting testosterone concentration was significantly higher in the OAM group than in the EF phase. These data might suggest that the subjects with menstrual disorder included those with hyperandrogenism. Although we did not systematically evaluate clinical symptoms of hyperandrogenism, all eight subjects in the OAM group showed a reference range of resting concentration of testosterone (0.28-0.46 ng·mL−1). The change in blood lactate through the exercise did not differ in all groups, suggesting that exercise intensity was similar among all groups.
Serum DHEAS levels increased in the EF phase at P 60, but they decreased in the OAM group at P 60. There is little information available on acute responses of DHEAS to resistance exercise in women. Riechman et al. (29) reported that acute resistance exercise increased blood DHEAS levels. The exercise-induced increases in DHEAS concentrations have been attributed to an increased rate of secretion from the adrenal cortex in response to ACTH stimulation (16). Meczekalski et al. (26) investigated the hypothalamic-pituitary-adrenal axis in hypothalamic amenorrhea and reported that the response of ACTH to corticotrophin-releasing hormone was significantly lower in amenorrheic women compared with healthy controls. In this study, DHEAS levels were not significantly increased immediately after resistance exercise in any of the groups, but the response at P 60 was different between the EF phase of the EM group and the OAM group. Reportedly, short-term exercise does not induce increased adrenal steroid production in response to ACTH secretion (7). It is possible that a higher-volume resistance exercise program for a longer period could induce an increase in DHEAS. Although resting levels of blood DHEAS in the EM group (EF and ML phases) seems to be a bit higher than that in the OAM group, these differences were not significant. Some previous studies (6,27) reported that resting levels of DHEAS in amenorrheic women seem to be similar to those in eumenorrheic women. In females, the synthesis and secretion of DHEAS occur in the adrenal cortex in response to ACTH. This may suggest that the large variability of DHEAS level can be attributed to altered pituitary hormones and/or conversion of androgens from the adrenal cortex.
IGF-I and testosterone concentrations were not significantly changed in response to the exercise, regardless of menstrual phase and menstrual disorder. This is consistent with previous studies (3,5,12,13,21). Cortisol, a catabolic hormone, also did not increase after the resistance exercise in the three groups. Kraemer et al. (24) have shown significant elevations in cortisol after acute resistance exercise in men and women. In contrast, Häkkinen and Pakarinen (12) reported no change in cortisol levels after acute resistance exercise in women. It is possible that the difference in cortisol response to exercise is affected by training status of subjects (24) and/or daily fluctuation, that is, the level of cortisol peaks in early morning and then declines sharply (2). All subjects in this study were not involved in any regular exercise or training, and all the resistance exercise was performed in the morning.
The responses of anabolic hormones to acute resistance exercise were different depending on the menstrual cycle state, suggesting that menstrual cycle state may influence the exercise training-induced skeletal muscular adaptation. Thus, it would be possible that training programs for eumenorrheic women might be timed in accordance with the menstrual cycle to maximize anabolic effects. In contrast, it is suggested that anabolic effects of resistance exercise are reduced in women with menstrual disturbances. Further studies will be needed to demonstrate the short-term and long-term effects on skeletal muscle by changes in hormonal responses to resistance exercise in different menstrual states. In conclusion, we demonstrated that the responses of anabolic hormones to acute resistance exercise are different in menstrual states among young women. Menstrual disturbance with low estradiol and progesterone status attenuated GH response to acute resistance exercise, suggesting that menstrual disorder accompanying with low ovarian hormone status affects exercise-induced change in anabolic hormones in women.
This study was supported in part by funds from Tsukuba University of Technology and University of Tsukuba.
The authors thank the volunteers who participated in this study. The assistance of Fumie Murai and the other technicians who collected blood samples is greatly appreciated. The authors also thank Dr. Takayuki Akimoto for his helpful comments on this article.
The results of the current study do not constitute endorsement by the American College of Sports Medicine.
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