Follicle-stimulating hormone | EPFL Graph Search

Follicle-stimulating hormone (FSH) is a gonadotropin, a glycoprotein polypeptide hormone. FSH is synthesized and secreted by the gonadotropic cells of the anterior pituitary gland and regulates the development, growth, pubertal maturation, and reproductive processes of the body. FSH and luteinizing hormone (LH) work together in the reproductive system. Structure FSH is a 35.5 kDa glycoprotein heterodimer, consisting of two polypeptide units, alpha and beta. Its structure is similar to those of luteinizing hormone (LH), thyroid-stimulating hormone (TSH), and human chorionic gonadotropin (hCG). The alpha subunits of the glycoproteins LH, FSH, TSH, and hCG are identical and consist of 96 amino acids, while the beta subunits vary. Both subunits are required for biological activity. FSH has a beta subunit of 111 amino acids (FSH β), which confers its specific biologic action, and is responsible for interaction with the follicle-stimulating hormone receptor. The sugar portion of

Development of an in vitro microenvironment for maturing oocytes

Jong Eun Ihm

The development of in vitro culture systems, comparable to the in vivo microenvironment in terms of effect on the oocyte growth and development could provide a valuable experimental tool for studying the mechanisms governing folliculogenesis. This tool might serve as well for practical clinical, agricultural, zoological, or biotechnological applications. This thesis reports on the importance of the microenvironment for the ovarian folliculogenesis process. The complexity of such a microenvironment was approached with a strategy based on functionalized PEG-hydrogels. The PEG matrix not only served as a scaffold, but it was also used a "reservoir" of immobilized cues. Tethered integrin-binding peptides in combination with other signaling factors aimed at better understanding the interactions of the oocyte and its surrounding granulosa cells that, most probably, determine the efficiency of the actual and the future in vitro mature oocyte production strategies. This work led also to investigating the lymph angiogenesis and the lymphatic transport in the context of oocyte maturation and their impact on mice fertility. In a first step, the mechanical properties of PEG-hydrogel were optimized for culturing secondary follicles. This report shows that the growth of the follicles was highly dependent on the mechanical properties of the surrounding environment. The optimal elastic modulus was found to be close to 900 Pa. In a second step, the effect of Arg-Gly-Asp (RGD) peptide, the minimal integrin-binding sequence, was studied. RGD presence did not influence the follicle growth rate but it significantly improved the quality of the produced oocytes. These findings demonstrated that approaching, biochemically and mechanically, the complexity of the ovarian extracellular matrix could be a winning strategy. The effect of key soluble factors was also investigated in order 1- to confirm their compatibility with the established 3-D culture system and 2- to further improve qualitatively and quantitatively the produced mature oocytes. Various combinations of gonadotropins such as the follicle-stimulating hormone (FSH) or the luteinizing hormone (LH) were tested. Interestingly, the effects of the gonadotropins in the 3-D PEG system were close to their known effects in vivo. Here the aberrant effects of these hormones in the used 2-D systems appeared clearly. c-Kit ligand (KL) is suspected to be one of the most important factors for the activation of primordial follicles and thus for controlling the exit from the resting pool. Previously, studying the effects of the two forms of KL (soluble and membrane-attached) had to cope with the lack of biologically-relevant immobilization strategies. For overcoming this problem, KL constructs were designed to include a substrate sequence for Factor XIIIa (NQEQVSPL or NQEQVSPLRCG). Thus the produced recombinant KL proteins could be enzymatically crosslinked to the PEG matrix. The different constructs of KL, including a wild type extracellular domain, were successfully cloned and produced from mammalian HEK-293 cells. The identity and the activity of the produced proteins were confirmed. Ovarian tissues from four days-old mice were cultured in PEG-hydrogels functionalized with KL. The results showed that the primordial follicles grew and were activated in the PEG-hydrogels where KL was immobilized but not when the soluble form of KL was preset in the medium. This experiment showed that the membrane-attached and the soluble form of KL play a different role in the rodent folliculogenesis. In parallel it was observed that blocking vascular endothelial growth factor receptor-3 (VEGFR-3) signaling had a critical but still unsuspected role in reproduction. This study demonstrates that variation in lymphangiogenesis is a regular, non-pathological event during folliculogenesis in the ovary; blocking lymphangiogenesis, might have an effect on hormone transport and thus on pregnancy. The reported results demonstrates that the blockade of lymphangiogenesis decreases the progesterone and estradioal levels during pregnancy and in fine results in failed fetal development. In conclusion, this study demonstrates the efficiency and the flexibility of a novel 3-D culture system. Circumventing problems inherent to the "on-plastic" standard culture, such as the loss of the granulosa-oocyte interactions, allowed the emergence of a culture system tailored for investigating fundamental folliculogenesis-related questions. Furthermore, the reported culture system might serve as a platform for developing clinical and biotechnology applications.

EPFL2008

NR5A2 Regulates Lhb and Fshb Transcription in Gonadotrope-Like Cells In Vitro, but Is Dispensable for Gonadotropin Synthesis and Fertility In Vivo

Johan Auwerx, Vikas Kumar, Kristina Schoonjans, Ying Wang, Xin Zhou

Successful mammalian reproduction depends on proper synthesis of the pituitary-derived glycoprotein hormones, luteinizing hormone (LH) and follicle-stimulating hormone (FSH). Several transcription factors cooperate to activate cell-specific and hormone-regulated expression of the gonadotropin beta subunits (Lhb and Fshb). Among these, NR5A1 (steroidogenic factor 1; SF1) has been shown to directly bind to the Lhb promoter, mediate basal and gonadotropin-releasing hormone (GnRH)-stimulated Lhb transcription, and possibly directly regulate Fshb expression. Recently, the closely-related NR5A2 was shown to activate the rat Lhb promoter in vitro. Here, we further characterized the role of NR5A2 in regulating gonadotropin synthesis. Ectopically expressed NR5A2 directly activated the murine Lhb promoter in a manner identical to that of NR5A1, whereas neither factor activated the murine Fshb promoter. In L beta T2 gonadotrope-like cells, depletion of endogenous NR5A1 or NR5A2 impaired basal and GnRH-stimulated Lhb and Fshb transcription. To analyze the physiological role of NR5A2 in gonadotropes in vivo, we generated mice with a gonadotrope-specific deletion of Nr5a2. In contrast with our in vitro data, these mice had normal pituitary Lhb and Fshb expression and intact fertility. Together, our data establish that NR5A2 can act in a non-redundant manner to regulate Lhb and Fshb transcription in vitro, but is dispensable in vivo.

Public Library of Science2013

Murine ovarian follicle culture in PEG-hydrogel: Effects of mechanical properties and the hormones FSH and LH on development

Jeffrey Alan Hubbell, Jong Eun Ihm, Seung Tae Lee

The development of an in vitro culture system, comparable to the in vivo environment in terms of effects on oocyte growth and development, could provide a valuable experimental tool for studying the mechanisms governing oocyte development as well as practical clinical, agricultural, zoological, and biotechnological applications. This study reports on the importance of the microenvironment for the ovarian folliculogenesis process. The complexity of such a microenvironment was approached with a strategy based on functionalized PEG-hydrogels. The PEG matrix not only serves as a scaffold, but it is also used a reservoir of immobilized cues. Using tethered integrin-binding peptides in combination with other signaling factors, we aimed at better understanding the interactions of the oocyte and its surrounding granulosa that may determine the efficiency of the actual and the future in vitro mature oocyte production. In a first step, the mechanical properties of PEG-hydrogel were optimized for producing secondary follicles, in which the oocyte is surrounded by two layers of granulosa cells. Follicle growth was highly dependent on the mechanical properties of the surrounding environment, with the optimal elastic modulus being approx. 1 kPa. The effects of key soluble factors were also investigated to confirm their compatibility with the established 3-D culture system and to further qualitatively and quantitatively improve the produced mature oocytes. Accordingly, various combinations of the gonadotropins follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were tested. Interestingly, the effects of the gonadotropins in the 3-D PEG system were close to their known in vivo effects. In conclusion, this study demonstrates the efficiency and the flexibility of a novel 3-D culture system, PEG-hydrogel. Circumventing problems inherent to the "on-plastic" standard culture, such as the loss of the granulosa-oocyte interactions, allowed the emergence of a culture system tailored for investigating fundamental folliculogenesis-related questions. Furthermore, the reported culture system may serve as a platform for developing clinical and biotechnology applications.