Culture of bovine ovarian follicle wall sections maintained the highly estrogenic profile under basal and chemically defined conditions

Abstract

Follicle cultures reproduce in vitro the functional features observed in vivo. In a search for an ideal model, we cultured bovine antral follicle wall sections (FWS) in a serum-free defined medium (DM) known to induce 17β-estradiol (E2) production, and in a nondefined medium (NDM) containing serum. Follicles were sectioned and cultured in NDM or DM for 24 or 48 h. Morphological features were determined by light microscopy. Gene expression of steroidogenic enzymes and follicle-stimulating hormone (FSH) receptor were determined by RT-PCR; progesterone (P4) and E2 concentrations in the media were measured by radioimmunoassay. DM, but not NDM, maintained an FWS morphology in vitro that was similar to fresh tissue. DM also induced an increase in the expression of all steroidogenic enzymes, except FSH receptor, but NDM did not. In both DM and NDM, there was a gradual increase in P4 throughout the culture period; however, P4 concentration was significantly higher in NDM. In both media, E2 concentration was increased at 24 h, followed by a decrease at 48 h. The E2:P4 ratio was higher in DM than in NDM. These results suggest that DM maintains morphological structure, upregulates the expression of steroidogenic enzyme genes, and maintains steroid production with a high E2:P4 ratio in FWS cultures.

Effects of PI3K and FSH on steroidogenesis, viability and embryo development of the cumulus–oocyte complex after in vitro culture

“…The patented MIV B medium is different from MIV C due to its lack of hormones as shown in a previous analysis (Vasconcelos et al, 2013). MIV C was described as a pro-estrogenic medium in a study in which the wall sections of bovine ovarian follicles were cultured in vitro with medium containing no FSH and supplemented with androstenedione and other hormones (Vasconcelos et al, 2013). The MIV B medium is also estrogenic and sustains COCs in vitro even when FSH is absent (E2:P4 ratio is approximately 1.0).…”

“…The 0FSH medium was able to sustain steroidogenesis based on androstenedione precursor and, as expected, the addition of FSH enhanced its ability. The patented MIV B medium is different from MIV C due to its lack of hormones as shown in a previous analysis (Vasconcelos et al, 2013). MIV C was described as a pro-estrogenic medium in a study in which the wall sections of bovine ovarian follicles were cultured in vitro with medium containing no FSH and supplemented with androstenedione and other hormones (Vasconcelos et al, 2013).…”

In vivo antral follicle wall biopsy: a new research technique to study ovarian function at the cellular and molecular levels

“…Although those ovarian sources provide sufficient materials for various molecular biology techniques, follow-up studies are either impossible or potentially jeopardize animals’ fertility. In this regard, ovaries obtained from slaughterhouses have been used for developing in vitro culture systems for both granulosa and theca cells, and follicle wall culture; those systems have proven to be vital for the understanding of the follicle wall’s cell function, steroidogenesis, and, subsequently, mammalian folliculogenesis [ 20 – 22 ]. However, in most cases when ovaries obtained from post-mortem animals are used, the experimental design is limited and compromised by lack of control of follicle health status.…”

Insulin influences developmental competence of bovine oocytes cultured in α-MEM plus follicle-simulating hormone

“…Some ultrastructural cytoplasmic changes in the oocyte occur after 18 h and extrusion of the polar body occurs 19 h after the luteinizing hormone (LH) surge (Ferreira et al ., 2009). However, the maturation process in vitro is influenced by culture environment; alpha minimum essential medium (α-MEM) defined medium (DCM) has been used to mimic the physiological in vivo conditions as it maintains oestrogen concentrations, aromatase expression in follicular cell walls (Vasconcelos et al ., 2013) and blockage of oocyte nuclear maturation (Oliveira e Silva et al ., 2011).…”