No abstract

Ovarian follicle steroidogenesis associated with embryo quality results in a successful pregnancy. Each follicle consists of an oocyte surrounded by granulosa cells, which secrete several steroid and peptide hormones. Follicles harvested from women who conceived after assisted reproductive therapy (ART) had significantly higher estradiol levels in follicular fluids than the follicles from women who failed to conceive after ART. The higher follicular estradiol levels correlate well with successful fertilization following ART. Mitochondria are the central sites for steroid hormone biosynthesis. The first and rate-limiting step in the biosynthesis of steroid hormones occurs in the mitochondria of granulosa cells. In the present study, we hypothesized that the mitochondria in granulosa cells are critical for maintaining oocyte quality and fertility capacity. This study aims to clarify the relationship between mitochondrial function and granulosa cell steroidogenesis, and the relationship between hormone levels and fertility capacity. Sera, follicular fluids and granulosa cells were obtained from individuals undergoing IVF-ET treatment. The oocyte numbers, oocyte quality, fertilization rate, and pregnancy rate were also recorded. The patients who provided the granulosa cells were further classified into four groups: endometriosis, ovarian endometrioma, endometriosis without ovarian endometrioma, and polycystic ovary syndrome (PCOS); patients with other female factor infertility and male factor infertility were used as controls. We measured the levels of estradiol (E2) by radioimmunoassay. Concurrently, we analyzed the mitochondrial mass and membrane potential, and apoptosis by flow cytometry using nonyl acridine orange, TMRE, Annexin V-FITC and PI. Mitochondrial morphology was visualized by transfection with pLV-mitoDsRed. In addition, we assessed the protein levels of steroidogenic enzymes, steroidogenic acute regulatory protein (StAR) and 3β-hydroxysteroid dehydrogenase (3β-HSD) by Western blot. The results showed significantly decreased serum E2 and follicular E2 levels, and decreased IVF outcomes, in the patients with endometriosis. Reduced mitochondrial mass and decreased mitochondrial membrane potential were correlated with lower E2. Furthermore, a significant decrease in StAR and 3β-HSD was found in patients with ovarian endometrioma. The enzyme levels of StAR and 3β-HSD were highly correlated with E2 levels. Finally, elevated cumulus cell apoptosis was found in the patient group with ovarian endometrioma and PCOS. In conclusion, mitochondrial dysfunction of human granulosa cells may contribute to the decline of steroidogenesis, decreased fertilization rate, oocyte maturation rate, and oocyte quality, and it can ultimately jeopardize fertility.

17α-estradiol (αE2), an endogenous stereoisomer of the hormone 17β-estradiol (E2), is capable of binding to estrogen receptors (ER). We aimed to mathematically describe, using experimental data, the possible interactions between αE2 and sperm ER during the process of sperm capacitation and to develop a kinetic model. The goal was to compare the suggested kinetic model with previously published results of ER interactions with E2 and 17α-ethynylestradiol (EE2). The HPLC-MS/MS method was developed to monitor the changes of αE2 concentration during capacitation. The calculated relative concentrations Bt were used for kinetic analysis. Rate constants k and molar ratio n were optimized and used for the construction of theoretical B(t) curves. Modifications in αE2–ER interactions were discovered during comparison with models for E2 and EE2. These new interactions displayed autocatalytic formation of an unstable adduct between the hormone and the cytoplasmic receptors. αE2 accumulates between the plasma membrane lipid bilayer with increasing potential, and when the critical level is reached, αE2 penetrates through the inner layer of the plasma membrane into the cytoplasm. It then rapidly reacts with the ER and creates an unstable adduct. The revealed dynamics of αE2–ER action may contribute to understanding tissue rejuvenation and the cancer-related physiology of αE2 signaling.