Circulating profile of Activin-Follistatin-Inhibin Axis in women with hypothalamic amenorrhea in response to leptin treatment

Abstract

Background: Chronic energy deficiency observed in women that exercise strenuously affects reproductive function, often leading to hypothalamic amenorrhea (HA). In such conditions, hypoleptinemia and robust changes in the Activin-Follistatin-Inhibin Axis (AFI) are observed. Treatment with leptin restores menstruation in many (60% responders) but not all (40% non-responders) women, suggesting that leptin is not the only regulator of reproductive function related to energy balance. In this work, we aimed to identify differences in hormonal profiles between leptin responders and non-responders among women with HA, with particular focus on the AFI axis.

Methods: AFI axis and reproductive hormones (LH, FSH, Estradiol, ΑΜΗ) were measured in blood in: a) An open-label interventional study, b) a randomized placebo-controlled trial, both investigating responders versus non-responders/women with HA treated with leptin.

Results: Women with HA that responded to leptin treatment have higher circulating levels/peak values of Inhibin A, Estradiol (E2), higher LH/FSH ratio and a trend to lower AMH compared with non-responders.

Conclusions: Components of the AFI axis are associated with improvement of reproductive function in women with HA treated with leptin. ΑΜΗ may serve as a marker of ovarian recovery under HA treatment.

Trial registration: ClinicalTrials.gov NCT00130117.

Keywords: Anorexia; Energy; Exercise; Fasting; Infertility; Reproduction.

Conflict of interest statement

Declaration of competing interest EB and NP have nothing to declare. CSM is advisor of Ansh Labs LLC.

Copyright © 2020 Elsevier Inc. All rights reserved.

Figures

Figure 1:. Circulating leptin levels and body composition parameters in responders vs non-responders to leptin treatment (study 1 and study 2).

Leptin levels, BMI, total fat mass, and lean mass of responders vs non-responders during leptin treatment in study 1 (a-d) and study 2 (e-h) are demonstrated. Data are reported as mean ± SEM. P-values one-tailed for group, time, and group by time effect were calculated with mixed models and are shown below the corresponding graphs (unadjusted and adjusted for baseline). For all molecules post-hoc Fisher’s LSD test was performed to compare responders vs non-responders for each timepoint. *, **, *** correspond to p≤0.05, 0.01, 0.001 respectively. adj.: adjusted; BMI: Body Mass Index; bsl: baseline; SEM: Standard Error of the Mean; unadj.: unadjusted

Figure 2.. Circulating profile of gonadotropins and E2 in responders vs non-responders to leptin treatment (study 1 and study 2).

Blood concentrations of responders vs non-responders of LH, FSH, LH to FSH ratio, and E2 in study 1 (a-d) and study 2 (e-h) are demonstrated. Data are reported as mean ± SEM. P-values one-tailed for group, time, and group by time effect were calculated with mixed models and are shown below the corresponding graphs (unadjusted and adjusted for baseline). For all molecules post-hoc Fischer LSD test was performed to compare responders vs non-responders for each timepoint. *, **, *** correspond to p≤0.05, 0.01, 0.001 respectively. adj.: adjusted; bsl: baseline; E2: Estradiol; FSH: Follicle-stimulating hormone; LH: Luteinizing hormone; SEM: Standard Error of the Mean; unadj.: unadjusted

Figure 3.. Circulating profile of Inhibins and AMH in responders vs non-responders to leptin treatment (study 1 and study 2).

Blood concentrations of responders vs non-responders of Inhibin A, Inhibin B, and AMH in study 1 (a-c), and study 2 (d-f) are demonstrated. Data are reported as mean ± SEM. P-values one-tailed for group, time, and group by time effect were calculated with mixed models and are shown below the corresponding graphs (unadjusted and adjusted for baseline). For all molecules post-hoc Fischer LSD test was performed to compare responders vs non-responders for each timepoint. *, **, *** correspond to p≤0.05, 0.01, 0.001 respectively. adj.: adjusted; bsl: baseline; AMH: Anti-mullerian hormone; SEM: Standard Error of the Mean; unadj.: unadjusted

Figure 4.. Circulating profile of Activins and Follistatins in responders vs non-responders to leptin treatment (study 1 and study 2).

Blood concentrations of responders vs non-responders of Activin A, Activin B, FST, and FSTL-3 in study 1 (a-d), and study 2 (e-h) are demonstrated. Data are reported as mean ± SEM. P-values one-tailed for group, time, and group by time effect were calculated with mixed models and are shown below the corresponding graphs (unadjusted and adjusted for baseline). For all molecules post-hoc Fischer LSD test was performed to compare responders vs non-responders for each timepoint. *, **, *** correspond to p≤0.05, 0.01, 0.001 respectively. adj.: adjusted; bsl: baseline; FST: Follistatin; FSTL-3: Follistatin-like 3; SEM: Standard Error of the Mean; unadj.: unadjusted

Figure 5:. Mechanism of restoration of reproductive function in women with hypothalamic amenorrhea by leptin.

Women with hypothalamic amenorrhea that respond to leptin treatment and have their reproductive function restored show a distinct hormonal profile when compared to non-responders. This hormonal profile is characterized by higher LH/FSH ratio, apparently due to the central effects of leptin on GnRH secretion in hypothalamus. Responders also have higher peaks of E2 and Inhibin A in blood, which indicates improved ovarian function and restoration of normal regular menstrual cycles. Additionally, circulating AMH, which serves as an additional marker of ovarian recovery, is reduced. AMH: Anti-mullerian hormone; E2: Estradiol; FSH: Follicle-stimulating hormone; GnRH: Gonadotropin-releasing hormone; LH: Luteinizing hormone