Ovarian Follicle Biology and the Basis for Gonadotoxicity

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Overview of Ovarian Function

The ovary serves two roles – the production of hormones necessary to support the endocrine health of the individual and the generation of mature oocytes that are able to be fertilized and contribute half of the genetic makeup of a new organism. The ovarian follicle is the functional unit of the ovary that carries out both of these goals. The ovarian follicles within an adult ovary produce steroid and protein hormones in cyclical patterns in response to tropic factors from the hypothalamus and pituitary ( Figure 1.1 ). These hormones then act locally to support follicle and oocyte growth and development, as well as systemically to support reproductive health, bone health and cardiovascular function. Interruption of the endocrine hormones at the level of the brain or gonad due to cancer treatment can result in loss of cyclicity and diminished endocrine health for short periods of time or can render an individual sterile due to loss of ovarian follicles.

Each ovarian follicle consists of one oocyte surrounded by supporting somatic cells, the granulosa and theca cells, and the follicle structure is embedded in an ovarian stroma that provides additional critical signals that determine follicle fate. Follicle development starts with selection of follicles from a pool of dormant primordial follicles – the ovarian reserve. The recruited follicles are activated and undergo a series of changes in morphology and size, developing through primary and secondary stages before acquiring a fluid-filled antral cavity. Although multiple follicles are selected to begin growing and eventually reach the antral stage, many are lost to atresia and only one (or a few depending on the species) reaches the Graffian follicle stage. Ovulation occurs in response to a timed surge of pituitary gonadotropins (see Figure 1.1 ) that triggers the release of a mature oocyte from the surface of the ovary into the fallopian tube, where it can be fertilized. The residual follicle unit (the remaining theca and mural granulosa cells) undergoes transformation to become the corpus luteum.

Ovarian follicle development is a carefully orchestrated event that is directed by both endocrine signalling within the hypothalamic–pituitary–ovarian axis as well as paracrine signalling occurring within and between follicles in the ovary. Hormone-regulated follicular development involves the integration of feedback loops between oestrogen, progesterone, inhibin A and inhibin B from the ovary; follicle-stimulating hormone (FSH) and luteinizing hormone (LH) from the pituitary; and hypothalamic gonadotropin-releasing hormone (GnRH) to direct follicle and oocyte development (see Figure 1.1 ). On the other hand, early follicle development is driven by intrinsic ovarian factors that regulate oocytic and follicular development before the follicles acquire FSH responsiveness. These locally acting factors and their roles in maintaining the dormant follicle pool, activating selected primordial follicles and supporting the earliest stages of follicle development are less well understood and of intense interest in the reproductive field. Understanding the early signalling events associated with maintaining a health-quiescent follicle pool may provide insights into gonadotoxicity and how to mitigate this effect.

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Ovarian Development

The mammalian gonad is formed during embryonic development ( Figure 1.2 ) after migration of primordial germ cells (PGCs) from the hindgut into the genital ridge between 9 to 10.5 days post coitum (dpc) in the mouse and at 6 weeks of gestation in humans. During this process, PGCs undergo mitotic divisions with incomplete cytokinesis resulting in cysts where germ cells remain connected by cytoplasmic bridges. At this stage, the gonad is bipotent. In the presence of a Y chromosome, the expression of the Sry gene (sex-determining region on the Y chromosome) directs male development through a process called sex determination. In contrast, sex determination in females requires the activity of several genes, such as Wnt4 , Rspo1 , β-Catenin , Foxl2 and Fst . Loss of these genes during development causes various degrees of female-to-male sex reversal.

Following female-specific gene activation, the early ovary develops distinct structural features, including the formation of ovigerous cords by interaction of the germ cell clusters with surrounding pregranulosa cells. Around 13.5 dpc, mouse germ cells enter meiosis in an anterior-to-posterior gradient believed to be driven by retinoic acid (RA) ; in the human ovary, this process begins in the medulla and spreads radially into the cortex from 10 weeks gestation. Reductive division (meiosis) begins during embryonic development in the female gonad, and postnatally in the male germ cell, respectively. Although the developing testis is also exposed to RA, the production of CYP26B1 allows degradation of RA and the male germ cells remain arrested in G1/G0. Oogonia enter the first stages of meiosis and arrest in diplotene of prophase I; at this time, they are referred to as oocytes and remain arrested in this stage until the LH surge that triggers ovulation, with the final stages of meiotic division completed upon fertilization.

The newly formed oocytes become individually encapsulated in a single layer of somatic cells to form primordial follicles (see Figure 1.2 ). In contrast to humans and other large mammals, most primordial follicles in rodents are formed a few days after birth, with some being activated soon after formation. This process of primordial follicle formation is also called nest breakdown because the pregranulosa cells invade and degrade the cytoplasmic bridges within the cysts while some germ cells are systematically eliminated. Several factors have been identified that control the timing and extent of nest breakdown, including Figla , Foxl2 , Nobox , c-Kit (oocyte)/ Kit ligand (pregranulosa and granulosa cells), members of the transforming growth factor beta (TGFβ) superfamily, oestrogens and phytoestrogens, progesterone, extracellular matrix factors, neurotrophins, and members of the Notch signalling pathway.

Any insults that occur during ovarian embryonic development may negatively impact adult ovarian function; these include poor nutrition, exposure to environmental pollutants, and exposure to certain drugs. Thus, ovarian development represents a critical window of vulnerability that can influence normal gonadal function in the adult animal.

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