Menstrual Cycle Hormonal Changes: Estradiol-β-17 and Progesterone Interactions on Exercise Fat Oxidation

During the last 50 years, research has demonstrated significant physiologic roles of female sex steroid hormones (SSH) on biological processes beyond reproduction. In the 1970s, findings from animal models began linking how SSHs, especially estrogens, impacted energy producing metabolic pathways [1]. Expansion to human models, where SSH were endogenously and exogenously manipulated, resulted in varied carbohydrate and fat oxidation rates for energy production (i.e., ATP) based on the underlying hormonal status [1]. Overall energy production, and the predominant substrate utilized, can have profound impacts on the ability of an individual to perform exercise [2]. Of late, tremendous public-scientific interest has developed concerning the influences of SSH changes across a woman’s menstrual cycle (MC) to affect their exercise training and sports performance capacity [3]. This interest is warranted since 49.6% of the world’s population is female and the number of women engaging in exercise activities for health or sporting endeavors is growing exponentially [4].

In endurance sporting activities, the ability to enhance fat metabolism, and thus spare carbohydrate energy sources, is viewed as a beneficial adaptive response as it mitigates the onset of fatigue (i.e., carbohydrate stores [glycogen] are limited) [2]. Studies have shown that as women transitions from the follicular phase (FP, reduced SSH) to the luteal phase (LP, elevated SSH) of their MC, a greater reliance on fat metabolism as an exercise energy substrate is observed [5]. Increased fat oxidation during the LP is attributed to changes in estrogen (E₂) concentrations as progesterone (P₄) can exhibit anti-estrogenic actions relative to aspects of energy metabolism [6]. In fact, D’Eon et al. clearly demonstrated the anti-estrogenic effects of P₄ on fat oxidation during exercise in a human study using exogenous SSH administration. These researchers postulated that fat metabolism is dependent on the relative concentrations of E₂ to P₄ (i.e., E₂/P₄ ratio), and that a two-fold or more increase in the ratio is necessary to invoke exercise-related substrate metabolism shifts (fat > carbohydrate) [7]. This is an intriguing postulate, but we questioned its in vivo validity since D’Eon’s hormonal manipulations were exogenous and their sample size was small (n=7). Therefore, we examined the issue further by investigating the in vivo influence of changes in the E₂/P₄ ratio on exercise fat oxidation rates in eumenorrheic women across their MC.

Methods

Healthy, physically active, pre-menopausal adult women (n=32) were recruited for this study (age=23±4 yr, BMI=22.5±2.5 kg/m²; X±SD). Inclusion criteria included: 1) not taking anti-inflammatory medications, oral contraceptives, or other hormonal therapies; 2) eumenorrheic status (≥6 months prior to study); 3) physically active ≥3 times/wk, 45–60 min at a moderate to vigorous intensity for ≥ 1 yr. Participants provided written informed consent in accordance with the Helsinki Declaration.

Each participant completed three laboratory sessions: 1) an orientation session involving determination of maximal oxygen uptake (VO_2max = 3.0±0.7 L/min), 2) and two 1-h submaximal steady-state treadmill run sessions at 65% VO_2max, once during the early FP (days 3–7) and once during the LP (days 19–25) phase of their MC (which ranged in length from 25 to 33 days) in random order. During the 1-h runs, standard non-protein respiratory exchange ratio-energy expenditure values via calorimetry were used to calculate fat oxidation values [8]. Details of the procedures and methods employed in this study appear elsewhere [9].

Resting blood samples were collected following ten minutes of supine rest before each 1-h run sessions. A blood sample was collected using standard venipuncture procedures (K₂-EDTA Vacutainer^® tubes), then immediately put on ice until processed to separate and store plasma −80°C. Blood estradiol-β-17 (E₂) and P₄ were measured using standard immunoassay techniques (Siemens Healthcare Technologies, Los Angeles, CA, USA).

Descriptive statistics were determined for all measurements (SPSS, version 25.0, Chicago, IL, USA). Normality of data distribution was assessed with the Kolmogorov Smirnov test. The FP vs. LP 1-h exercise responses for fat oxidation rates as well as resting E₂, P₄, and E₂/P₄ ratio were analyzed with paired t-tests. Effect size (Hedges g) were determined for all comparisons. Relationships between the E₂/P₄ ratio and oxidation responses were examined with Pearson correlation analyses. Significance was set at α < 0.05.

Results

Overall fat oxidation rates during exercise in the LP (0.49±0.19 g/min) were significantly (p<0.05) greater than in the FP running sessions (0.41±0.14 g/min; small effect size, 0.45 g). Resting E₂ and P₄ values were significantly (p<0.01) higher in the LP vs. FP sessions (E₂ = 518.5±285.4 vs. 243.8±143.2 pmol/L; and P₄ = 47.9±19.1 vs. 2.9±1.5 nmol/L; large effect size = 1.20–3.11 g, respectively).

Percentage delta (Δ) values representing the changes in fat oxidation and E₂/P₄ ratio (log transformed [10]) were calculated for LP vs. FP session responses and examined for relationships (Δ = LP – FP). A significant effect of Δ responses (r=0.743; p<0.001) for ratio changes was observed, such that greater increases in E₂/P₄ ratio was associated with increased fat oxidation (Figure 1).

Figure 1.

Delta changes (Δ = Luteal phase – Follicular phase) for exercise fat oxidation and E₂/P₄ ratio.

Discussion

Our findings support that greater increases in the E₂/P₄ ratio, such as observed during the LP vs. FP of the MC, is associated with a greater reliance on fat as an energy substrate during exercise. This agrees with the earlier work of D’Eon et al. who manipulated hormone levels exogenously [7] and reported that P₄ has anti-estrogenic effects on substrate oxidation during exercise. Review of Figure 1, however, suggests their premise that an increase in the E₂/P₄ ratio of a specific magnitude was necessary to induce a metabolism shift towards a greater reliance on fat during LP exercise was not clearly supported.

Prior in vivo human work has also demonstrated a greater dependence on fat as an exercise energy substrate in women during the MC LP vs. FP; although not a universal finding [5]. However, many of these prior studies, unlike the current, did not account for the role P₄ may play in influencing substrate metabolism [1,5]. Interestingly, the role of E₂ to augment exercise fat oxidation appears intensity dependent; i.e., as exercise intensities increases (reaching ~ 75% VO_2max) the influence of E₂ becomes markedly attenuated [11]. This was the primary reason we examined a lower intensity exercise, and one commonly associated with aerobic exercise training [11].

We utilized a prospective, cohort study approach (i.e., each participant served as their own control) and examined the association between MC SSH changes and exercise fat oxidation. As such, this research design did not allow us to examine the mechanism of our findings, but prior studies have detected the signaling pathways whereby E₂ promotes increased fat oxidation. In skeletal muscle, E₂ via (E₂ receptor-α) activates gene expression of the nuclear hormone receptors, peroxisome proliferator-activated receptor-α and -Δ to enhance expression of long chain fatty acid (LCFA) transporters, mitochondrial transcription factors, β-oxidation enzymes and kinases that direct metabolism away from carbohydrate and towards fat [5,9,12]. E₂ exposure also increases lipid availability in skeletal muscle by promoting intramuscular lipid storage and by redistributing LCFA away from adipose tissue and toward skeletal muscle decreasing adipose lipid storage, and as such, promoting adipose lipolysis [12,13].

A limitation to our study is the use of calorimetry to assess energy substrate metabolism rather than direct biochemical measurements. However, this non-invasive technique has been well validated and is supported in the research literature for its accuracy/appropriateness based upon the exercise protocol used herein [15]. Additionally, we only examined the FP and LP components of the MC, and exercise fat oxidation has been reported higher at ovulation when circulating E₂ is typically greatest [15]. However, we chose to not assess the ovulation phase due to the logistic difficulties (i.e., extremely short phase time period, ~1 d).

In conclusion, increases in SSH concentrations from the FP to LP in eumenorrheic women are associated with a greater reliance on fat as an exercise energy substrate. Furthermore, the interaction of changes in E₂ and P₄ across the MC do interplay to influence substrate oxidation, in which a greater magnitude of P₄ increase relative to E₂ increase seems to attenuate the extent of the fat oxidation shift between MC phases (FP→LP). This interaction may explain why some studies report no, or minimal energy substrate changes between MC phases in exercising women (i.e., they have a relatively low E₂/P₄ ratio change between MC phases).

Data Availability

The datasets created and/or evaluated during the present study are not currently publicly available. They may be available from the corresponding author on reasonable request.